WorldWideScience

Sample records for high-affinity branched-chain amino

  1. Branched-chain fatty acid biosynthesis in a branched-chain amino acid aminotransferase mutant of Staphylococcus carnosus

    DEFF Research Database (Denmark)

    Beck, Hans Christian

    2005-01-01

    Fatty acid biosynthesis by a mutant strain of Staphylococcus carnosus deficient in branched-chain amino acid aminotransferase (IlvE) activity was analysed. This mutant was unable to produce the appropriate branched-chain alpha-ketoacid precursors for branched-chain fatty acid biosynthesis from...... for 2-methylpropanoic acid production, revealing that the IlvE protein plays an important, but not essential role in the biosynthesis of branched-chain fatty acids and secondary metabolites in S. carnosus....

  2. Role of mitochondrial transamination in branched chain amino acid metabolism

    Energy Technology Data Exchange (ETDEWEB)

    Hutson, S.M.; Fenstermacher, D.; Mahar, C.

    1988-03-15

    Oxidative decarboxylation and transamination of 1-/sup 14/C-branched chain amino and alpha-keto acids were examined in mitochondria isolated from rat heart. Transamination was inhibited by aminooxyacetate, but not by L-cycloserine. At equimolar concentrations of alpha-ketoiso(1-/sup 14/C)valerate (KIV) and isoleucine, transamination was increased by disrupting the mitochondria with detergent which suggests transport may be one factor affecting the rate of transamination. Next, the subcellular distribution of the aminotransferase(s) was determined. Branched chain aminotransferase activity was measured using two concentrations of isoleucine as amino donor and (1-/sup 14/C)KIV as amino acceptor. The data show that branched chain aminotransferase activity is located exclusively in the mitochondria in rat heart. Metabolism of extramitochondrial branched chain alpha-keto acids was examined using 20 microM (1-/sup 14/C)KIV and alpha-ketoiso(1-/sup 14/C)caproate (KIC). There was rapid uptake and oxidation of labeled branched chain alpha-keto acid, and, regardless of the experimental condition, greater than 90% of the labeled keto acid substrate was metabolized during the 20-min incubation. When a branched chain amino acid (200 microM) or glutamate (5 mM) was present, 30-40% of the labeled keto acid was transaminated while the remainder was oxidized. Provision of an alternate amino acceptor in the form of alpha-keto-glutarate (0.5 mM) decreased transamination of the labeled KIV or KIC and increased oxidation. Metabolism of intramitochondrially generated branched chain alpha-keto acids was studied using (1-/sup 14/C)leucine and (1-/sup 14/C)valine. Essentially all of the labeled branched chain alpha-keto acid produced by transamination of (1-/sup 14/C)leucine or (1-/sup 14/C)valine with a low concentration of unlabeled branched chain alpha-keto acid (20 microM) was oxidized.

  3. Branched-chain Amino Acid Metabolon: INTERACTION OF GLUTAMATE DEHYDROGENASE WITH THE MITOCHONDRIAL BRANCHED-CHAIN AMINOTRANSFERASE (BCATm)*

    OpenAIRE

    Islam, Mohammad Mainul; Nautiyal, Manisha; Wynn, R. Max; Mobley, James A.; Chuang, David T.; Hutson, Susan M.

    2009-01-01

    The catabolic pathway for branched-chain amino acids includes deamination followed by oxidative decarboxylation of the deaminated product branched-chain α-keto acids, catalyzed by the mitochondrial branched-chain aminotransferase (BCATm) and branched-chain α-keto acid dehydrogenase enzyme complex (BCKDC). We found that BCATm binds to the E1 decarboxylase of BCKDC, forming a metabolon that allows channeling of branched-chain α-keto acids from BCATm to E1. The protein complex also contains glut...

  4. Branched-chain amino acid metabolon: interaction of glutamate dehydrogenase with the mitochondrial branched-chain aminotransferase (BCATm).

    Science.gov (United States)

    Islam, Mohammad Mainul; Nautiyal, Manisha; Wynn, R Max; Mobley, James A; Chuang, David T; Hutson, Susan M

    2010-01-01

    The catabolic pathway for branched-chain amino acids includes deamination followed by oxidative decarboxylation of the deaminated product branched-chain alpha-keto acids, catalyzed by the mitochondrial branched-chain aminotransferase (BCATm) and branched-chain alpha-keto acid dehydrogenase enzyme complex (BCKDC). We found that BCATm binds to the E1 decarboxylase of BCKDC, forming a metabolon that allows channeling of branched-chain alpha-keto acids from BCATm to E1. The protein complex also contains glutamate dehydrogenase (GDH1), 4-nitrophenylphosphatase domain and non-neuronal SNAP25-like protein homolog 1, pyruvate carboxylase, and BCKDC kinase. GDH1 binds to the pyridoxamine 5'-phosphate (PMP) form of BCATm (PMP-BCATm) but not to the pyridoxal 5'-phosphate-BCATm and other metabolon proteins. Leucine activates GDH1, and oxidative deamination of glutamate is increased further by addition of PMP-BCATm. Isoleucine and valine are not allosteric activators of GDH1, but in the presence of 5'-phosphate-BCATm, they convert BCATm to PMP-BCATm, stimulating GDH1 activity. Sensitivity to ADP activation of GDH1 was unaffected by PMP-BCATm; however, addition of a 3 or higher molar ratio of PMP-BCATm to GDH1 protected GDH1 from GTP inhibition by 50%. Kinetic results suggest that GDH1 facilitates regeneration of the form of BCATm that binds to E1 decarboxylase of the BCKDC, promotes metabolon formation, branched-chain amino acid oxidation, and cycling of nitrogen through glutamate.

  5. Branched chain amino acids requirements and metabolism in pigs

    DEFF Research Database (Denmark)

    Assadi Soumeh, Elham

    2015-01-01

    according to the ideal protein profile that is compatible with the animal AA demand for normal body function. During the past decades, it has been tried to understand and characterize branched chain amino acids (BCAA) requirements, biological importance, and mode of actions. This is interesting for two...... reasons: first, BCAA share the same enzymes in their catabolic pathways, and there is an interaction among them in a way that excess Leu for example increases the catabolism of them all and changes the requirements. Second, BCAA are not only building blocks of protein biosynthesis, but are also involved...... in important regulatory mechanisms and biological functions, e.g. muscle protein synthesis, chronic diseases, neurotransmitter biosynthesis, and so on. Identifying biomarkers of the BCAA status may help to understand their biological effects. The objectives of the current study were first to estimate Ile, Val...

  6. Early administration of branched-chain amino acid granules

    Institute of Scientific and Technical Information of China (English)

    Toru Ishikawa

    2012-01-01

    The effect of malnutrition on survival in patients with decompensated liver cirrhosis has not been well defined.Nutritional intervention with branched-chain amino acid (BCAA) can increase serum albumin concentration in patients with decompensated cirrhosis but its effects on survival are unclear.The BCAA to tyrosine ratio (BTR) is a surrogate marker (the normal range of BTR is between 4.41 and 10.05,and a Fischer's ratio of 1.8 corresponds to a BTR of 3.5) in patients with decompensated liver cirrhosis,and BCAA inhibits hepatic carcinogenesis in patients with compensated cirrhosis.This review discusses data regarding the effect of early administration of BCAA granules based on the ratio of BCAA to BTR on prognosis in patients with cirrhosis.

  7. Branched chain amino acid profile in early chronic kidney disease

    Directory of Open Access Journals (Sweden)

    M Anil Kumar

    2012-01-01

    Full Text Available The nutritional status in chronic kidney disease (CKD patients is a predictor of prognosis during the first period of dialysis. Serum albumin is the most commonly used nutritional marker. Another index is plasma amino acid profile. Of these, the plasma levels of branched chain amino acids (BCAA, especially valine and leucine, correlate well with nutritional status. Plasma BCAAs were evaluated along with albumin and C-reactive protein in 15 patients of early stages of CKD and 15 age- and sex-matched healthy controls. A significant decrease in plasma valine, leucine and albumin levels was observed in CKD patients when compared with the controls (P <0.05. No significant difference in C-reactive protein (CRP levels was observed between the two groups. Malnutrition seen in our CKD patients in the form of hypoalbuminemia and decreased concentrations of BCAA points to the need to evaluate the nutritional status in the early stages itself. Simple measures in the form of amino acid supplementation should be instituted early to decrease the morbidity and mortality before start of dialysis in these patients.

  8. Catabolism of Branched Chain Amino Acids Supports Respiration but Not Volatile Synthesis in Tomato Fruits

    Institute of Scientific and Technical Information of China (English)

    Andrej Kochevenko; Wagner L.Araújo; Gregory S.Maloney; Denise M.Tieman; Phuc Thi Do; Mark G.Taylor; Harry J.Klee; Alisdair R.Fernie

    2012-01-01

    The branched-chain amino acid transaminases (BCATs) have a crucial role in metabolism of the branched-chain amino acids leucine,isoleucine,and valine.These enzymes catalyze the last step of synthesis and the initial step of degradation of these amino acids.Although the biosynthetic pathways of branched chain amino acids in plants have been extensively investigated and a number of genes have been characterized,their catabolism in plants is not yet completely understood.We previously characterized the branched chain amino acid transaminase gene family in tomato,revealing both the subcellular localization and kinetic properties of the enzymes encoded by six genes.Here,we examined possible functions of the enzymes during fruit development.We further characterized transgenic plants differing in the expression of branched chain amino acid transaminases 1 and 3,evaluating the rates of respiration in fruits deficient in BCAT1 and the levels of volatiles in lines overexpressing either BCAT1 or BCAT3.We quantitatively tested,via precursor and isotope feeding experiments,the importance of the branched chain amino acids and their corresponding keto acids in the formation of fruit volatiles.Our results not only demonstrate for the first time the importance of branched chain amino acids in fruit respiration,but also reveal that keto acids,rather than amino acids,are the likely precursors for the branched chain flavor volatiles.

  9. Branched-chain amino acids for hepatic encephalopathy. Protocol for Cochrane Review

    DEFF Research Database (Denmark)

    Gluud, C; Koretz, RL

    2000-01-01

    Hepatic encephalopathy may be caused by a decreased plasma ratio of branched-chain amino acids (BCAA) to aromatic amino acids. Treatment with BCAA may therefore have a beneficial effect on patients with hepatic encephalopathy.......Hepatic encephalopathy may be caused by a decreased plasma ratio of branched-chain amino acids (BCAA) to aromatic amino acids. Treatment with BCAA may therefore have a beneficial effect on patients with hepatic encephalopathy....

  10. Mathematical Modeling of the Process for Microbial Production of Branched Chained Amino Acids

    OpenAIRE

    Todorov K.; Georgiev T.; Ratkov A.

    2009-01-01

    This article deals with modelling of branched chained amino acids production. One of important branched chained amino acid is L-valine. The aim of the article is synthesis of dynamic unstructured model of fed-batch fermentation process with intensive droppings for L-valine production. The presented approach of the investigation includes the following main procedures: description of the process by generalized stoichiometric equations; preliminary data processing and calculation of specific rat...

  11. Oral branched-chain amino acids decrease whole-body proteolysis

    Science.gov (United States)

    Ferrando, A. A.; Williams, B. D.; Stuart, C. A.; Lane, H. W.; Wolfe, R. R.

    1995-01-01

    BACKGROUND: This study reports the effects of ingesting branched-chain amino acids (leucine, valine, and isoleucine) on protein metabolism in four men. METHODS: To calculate leg protein synthesis and breakdown, we used a new model that utilized the infusion of L-[ring-13C6]phenylalanine and the sampling of the leg arterial-venous difference and muscle biopsies. In addition, protein-bound enrichments provided for the direct calculation of muscle fractional synthetic rate. Four control subjects ingested an equivalent amount of essential amino acids (threonine, methionine, and histidine) to discern the effects of branched-chain amino acid nitrogen vs the effects of essential amino acid nitrogen. Each drink also included 50 g of carbohydrate. RESULTS: Consumption of the branched-chain and the essential amino acid solutions produced significant threefold and fourfold elevations in their respective arterial concentrations. Protein synthesis and breakdown were unaffected by branched-chain amino acids, but they increased by 43% (p < .05) and 36% (p < .03), respectively, in the group consuming the essential amino acids. However, net leg balance of phenylalanine was unchanged by either drink. Direct measurement of protein synthesis by tracer incorporation into muscle protein (fractional synthetic rate) revealed no changes within or between drinks. Whole-body phenylalanine flux was significantly suppressed by each solution but to a greater extent by the branched-chain amino acids (15% and 20%, respectively) (p < .001). CONCLUSIONS: These results suggest that branched-chain amino acid ingestion suppresses whole-body proteolysis in tissues other than skeletal muscle in normal men.

  12. A pilot, short-term dietary manipulation of branched chain amino acids has modest influence on fasting levels of branched chain amino acids

    OpenAIRE

    Cavallaro, Nicole Landa; Garry, Jamie; Shi, Xu; Gerszten, Robert E.; Anderson, Ellen J.; Walford, Geoffrey A.

    2016-01-01

    Background: Elevated fasting levels of branched chain amino acids (BCAAs: valine, isoleucine, leucine) in venous blood are associated with a variety of metabolic impairments, including increased risk of type 2 diabetes (T2D). Fasting BCAA levels are influenced by non-dietary factors. However, it is unknown whether fasting BCAAs can be altered through manipulation of dietary intake alone. Objective: To test whether a specific dietary intervention, using differences in BCAA intake, alters fasti...

  13. Mathematical Modeling of the Process for Microbial Production of Branched Chained Amino Acids

    Directory of Open Access Journals (Sweden)

    Todorov K.

    2009-12-01

    Full Text Available This article deals with modelling of branched chained amino acids production. One of important branched chained amino acid is L-valine. The aim of the article is synthesis of dynamic unstructured model of fed-batch fermentation process with intensive droppings for L-valine production. The presented approach of the investigation includes the following main procedures: description of the process by generalized stoichiometric equations; preliminary data processing and calculation of specific rates for main kinetic variables; identification of the specific rates takes into account the dissolved oxygen tension; establishment and optimisation of dynamic model of the process; simulation researches. MATLAB is used as a research environment.

  14. Branched-chain amino acids for people with hepatic encephalopathy

    DEFF Research Database (Denmark)

    Gluud, Lise Lotte; Dam, Gitte; Les, Iñigo

    2017-01-01

    -chain amino acids (BCAA) versus control interventions has evaluated if BCAA may benefit people with hepatic encephalopathy. OBJECTIVES: To evaluate the beneficial and harmful effects of BCAA versus any control intervention for people with hepatic encephalopathy. SEARCH METHODS: We identified trials through...

  15. Branched-chain amino acids for people with hepatic encephalopathy

    DEFF Research Database (Denmark)

    Gluud, Lise Lotte; Dam, Gitte; Les, Iñigo;

    2015-01-01

    -chain amino acids (BCAA) versus control interventions has evaluated if BCAA may benefit people with hepatic encephalopathy. OBJECTIVES: To evaluate the beneficial and harmful effects of BCAA versus any control intervention for people with hepatic encephalopathy. SEARCH METHODS: We identified trials through...

  16. Do branched chain amino acids improve hepatic encephalopathy in cirrhosis?

    Directory of Open Access Journals (Sweden)

    Maximiliano Vergara

    2016-12-01

    Full Text Available Resumen Existe controversia sobre si los aminoácidos de cadena ramificada son efectivos para el tratamiento de la encefalopatía hepática. Utilizando la base de datos Epistemonikos, la cual es mantenida mediante búsquedas en múltiples bases de datos, identificamos siete revisiones sistemáticas que en conjunto incluyen 32 estudios aleatorizados, de los cuales 30 responden la pregunta de este resumen. Extrajimos los resultados, realizamos un metanálisis y preparamos tablas de resumen de los resultados utilizando el método GRADE. Concluimos que los aminoácidos de cadena ramificada podrían llevar a una mejoría sintomática en la encefalopatía hepática, pero probablemente tienen poco o nulo efecto sobre la mortalidad.

  17. Do branched chain amino acids improve hepatic encephalopathy in cirrhosis?

    OpenAIRE

    Maximiliano Vergara; Victoria Castro-Gutiérrez; Gabriel Rada

    2016-01-01

    Resumen Existe controversia sobre si los aminoácidos de cadena ramificada son efectivos para el tratamiento de la encefalopatía hepática. Utilizando la base de datos Epistemonikos, la cual es mantenida mediante búsquedas en múltiples bases de datos, identificamos siete revisiones sistemáticas que en conjunto incluyen 32 estudios aleatorizados, de los cuales 30 responden la pregunta de este resumen. Extrajimos los resultados, realizamos un metanálisis y preparamos tablas de resumen de los r...

  18. Do branched chain amino acids improve hepatic encephalopathy in cirrhosis?

    OpenAIRE

    Maximiliano Vergara; Victoria Castro-Gutiérrez; Gabriel Rada

    2016-01-01

    Resumen Existe controversia sobre si los aminoácidos de cadena ramificada son efectivos para el tratamiento de la encefalopatía hepática. Utilizando la base de datos Epistemonikos, la cual es mantenida mediante búsquedas en múltiples bases de datos, identificamos siete revisiones sistemáticas que en conjunto incluyen 32 estudios aleatorizados, de los cuales 30 responden la pregunta de este resumen. Extrajimos los resultados, realizamos un metanálisis y preparamos tablas de resumen de los r...

  19. Branched-chain amino acids in metabolic signaling and insulin resistance

    Science.gov (United States)

    Branched-chain amino acids (BCAAs) are important directly- and indirectly-acting nutrient signals. Frequently, their actions have been reported to be anti-obesity in nature, especially in rodent models. Yet, circulating BCAAs tend to be elevated in obesity, and even associated with poorer metaboli...

  20. Enzymology of the branched-chain amino acid oxidation disorders: the valine pathway.

    Science.gov (United States)

    Wanders, Ronald J A; Duran, Marinus; Loupatty, Ference J

    2012-01-01

    Valine is one of the three branched-chain amino acids which undergoes oxidation within mitochondria. In this paper, we describe the current state of knowledge with respect to the enzymology of the valine oxidation pathway and the different disorders affecting oxidation.

  1. Branched-chain amino acids for people with hepatic encephalopathy

    DEFF Research Database (Denmark)

    Gluud, Lise Lotte; Dam, Gitte; Les, Iñigo

    2015-01-01

    -chain amino acids (BCAA) versus control interventions has evaluated if BCAA may benefit people with hepatic encephalopathy. OBJECTIVES: To evaluate the beneficial and harmful effects of BCAA versus any control intervention for people with hepatic encephalopathy. SEARCH METHODS: We identified trials through...... randomised clinical trials including 827 participants with hepatic encephalopathy classed as overt (12 trials) or minimal (four trials). Eight trials assessed oral BCAA supplements and seven trials assessed intravenous BCAA. The control groups received placebo/no intervention (two trials), diets (10 trials...... between BCAA and controls (risk ratio (RR) 0.88, 95% confidence interval (CI) 0.69 to 1.11; 760 participants; 15 trials; moderate quality of evidence). We found no evidence of small-study effects. Sensitivity analyses of trials with a low risk of bias found no beneficial or detrimental effect of BCAA...

  2. Adipose Tissue Branched Chain Amino Acid (BCAA) Metabolism Modulates Circulating BCAA Levels*

    OpenAIRE

    Herman, Mark A.; She, Pengxiang; Peroni, Odile D.; Lynch, Christopher J.; Kahn, Barbara B.

    2010-01-01

    Whereas the role of adipose tissue in glucose and lipid homeostasis is widely recognized, its role in systemic protein and amino acid metabolism is less well-appreciated. In vitro and ex vivo experiments suggest that adipose tissue can metabolize substantial amounts of branched chain amino acids (BCAAs). However, the role of adipose tissue in regulating BCAA metabolism in vivo is controversial. Interest in the contribution of adipose tissue to BCAA metabolism has been renewed with recent obse...

  3. Metformin inhibits Branched Chain Amino Acid (BCAA) derived ketoacidosis and promotes metabolic homeostasis in MSUD.

    Science.gov (United States)

    S Sonnet, Davis; N O'Leary, Monique; A Gutierrez, Mark; M Nguyen, Steven; Mateen, Samiha; Hsu, Yuehmei; P Mitchell, Kylie; J Lopez, Antonio; Vockley, Jerry; K Kennedy, Brian; Ramanathan, Arvind

    2016-07-04

    Maple Syrup Urine Disease (MSUD) is an inherited disorder caused by the dysfunction in the branched chain keto-acid dehydrogenase (BCKDH) enzyme. This leads to buildup of branched-chain keto-acids (BCKA) and branched-chain amino acids (BCAA) in body fluids (e.g. keto-isocaproic acid from the BCAA leucine), leading to numerous clinical features including a less understood skeletal muscle dysfunction in patients. KIC is an inhibitor of mitochondrial function at disease relevant concentrations. A murine model of intermediate MSUD (iMSUD) shows significant skeletal muscle dysfunction as by judged decreased muscle fiber diameter. MSUD is an orphan disease with a need for novel drug interventions. Here using a 96-well plate (liquid chromatography- mass spectrometry (LC-MS) based drug-screening platform we show that Metformin, a widely used anti-diabetic drug, reduces levels of KIC in patient-derived fibroblasts by 20-50%. This Metformin-mediated effect was conserved in vivo; Metformin-treatment significantly reduced levels of KIC in the muscle (by 69%) and serum (by 56%) isolated from iMSUD mice, and restored levels of mitochondrial metabolites (e.g. AMP and other TCA). The drug also decreased the expression of mitochondrial branched chain amino transferase (BCAT) which produces KIC in skeletal muscle. This suggests that Metformin can restore skeletal muscle homeostasis in MSUD by decreasing mitochondrial KIC production.

  4. Metformin inhibits Branched Chain Amino Acid (BCAA) derived ketoacidosis and promotes metabolic homeostasis in MSUD

    Science.gov (United States)

    S. Sonnet, Davis; N. O’Leary, Monique; A. Gutierrez, Mark; M. Nguyen, Steven; Mateen, Samiha; Hsu, Yuehmei; P. Mitchell, Kylie; J. Lopez, Antonio; Vockley, Jerry; K. Kennedy, Brian; Ramanathan, Arvind

    2016-01-01

    Maple Syrup Urine Disease (MSUD) is an inherited disorder caused by the dysfunction in the branched chain keto-acid dehydrogenase (BCKDH) enzyme. This leads to buildup of branched-chain keto-acids (BCKA) and branched-chain amino acids (BCAA) in body fluids (e.g. keto-isocaproic acid from the BCAA leucine), leading to numerous clinical features including a less understood skeletal muscle dysfunction in patients. KIC is an inhibitor of mitochondrial function at disease relevant concentrations. A murine model of intermediate MSUD (iMSUD) shows significant skeletal muscle dysfunction as by judged decreased muscle fiber diameter. MSUD is an orphan disease with a need for novel drug interventions. Here using a 96-well plate (liquid chromatography- mass spectrometry (LC-MS) based drug-screening platform we show that Metformin, a widely used anti-diabetic drug, reduces levels of KIC in patient-derived fibroblasts by 20–50%. This Metformin-mediated effect was conserved in vivo; Metformin-treatment significantly reduced levels of KIC in the muscle (by 69%) and serum (by 56%) isolated from iMSUD mice, and restored levels of mitochondrial metabolites (e.g. AMP and other TCA). The drug also decreased the expression of mitochondrial branched chain amino transferase (BCAT) which produces KIC in skeletal muscle. This suggests that Metformin can restore skeletal muscle homeostasis in MSUD by decreasing mitochondrial KIC production. PMID:27373929

  5. Evolution of the biosynthesis of the branched-chain amino acids

    Science.gov (United States)

    Keefe, Anthony D.; Lazcano, Antonio; Miller, Stanley L.

    1995-06-01

    The origin of the biosynthetic pathways for the branched-chain amino acids cannot be understood in terms of the backwards development of the present acetolactate pathway because it contains unstable intermediates. We propose that the first biosynthesis of the branched-chain amino acids was by the reductive carboxylation of short branched chain fatty acids giving keto acids which were then transaminated. Similar reaction sequences mediated by nonspecific enzymes would produce serine and threonine from the abundant prebiotic compounds glycolic and lactic acids. The aromatic amino acids may also have first been synthesized in this way, e.g. tryptophan from indole acetic acid. The next step would have been the biosynthesis of leucine from α-ketoisovaleric acid. The acetolactate pathway developed subsequently. The first version of the Krebs cycle, which was used for amino acid biosynthesis, would have been assembled by making use of the reductive carboxylation and leucine biosynthesis enzymes, and completed with the development of a single new enzyme, succinate dehydrogenase. This evolutionary scheme suggests that there may be limitations to inferring the origins of metabolism by a simple back extrapolation of current pathways.

  6. Evolution of the biosynthesis of the branched-chain amino acids

    Science.gov (United States)

    Keefe, Anthony D.; Lazcano, Antonio; Miller, Stanley L.

    1995-01-01

    The origins of the biosynthetic pathways for the branched-chain amino acids cannot be understood in terms of the backwards development of the present acetolactate pathway because it contains unstable intermediates. We propose that the first biosynthesis of the branched-chain amino acids was by the reductive carboxylation of short branched chain fatty acids giving keto acids which were then transaminated. Similar reaction sequences mediated by nonspecific enzymes would produce serine and threomine from the abundant prebiotic compounds glycolic and lactic acids. The aromatic amino acids may also have first been synthesized in this way, e.g. tryptophan from indole acetic acid. The next step would have been the biosynthesis of leucine from alpha-ketoisovalerc acid. The acetolactate pathway developed subsequently. The first version of the Krebs cycle, which was used for amino acid biosynthesis, would have been assembled by making use fo the reductive carboxylation and leucine biosynthesis enzymes, and completed with the development of a single new enzyme, succinate dehydrogenase. This evolutionary scheme suggests that there may be limitations to inferring the origins of metabolism by a simple back extrapolation of current pathways.

  7. Plasma branched chain amino acid abnormalities in sake-treated rats.

    Directory of Open Access Journals (Sweden)

    Watanabe,Akiharu

    1985-02-01

    Full Text Available Plasma amino acid abnormalities in rats treated with large doses of sake and whisky for 3 days were investigated under adequate nutritional conditions. A significant decrease in plasma branched-chain amino acid (BCAA levels was observed in sake- but not whisky-treated rats. However, known factors affecting BCAA levels, such as serum insulin and plasma glucagon levels ahd BCAA-metabolizing enzyme (BCAA transaminase and branched chain alpha-ketoacid dehydrogenase activities in the liver and skeletal muscle, were not significantly altered in the sake group. Furthermore, ethanol-metabolizing enzyme (alcohol and aldehyde dehydrogenases and the microsomal ethanol-oxidizing system activities in the liver were not altered in the sake group. Other mechanisms need to be considered for explaining the diminished levels of plasma BCAA in sake-treated rats.

  8. Sodium Phenylbutyrate Decreases Plasma Branched-Chain Amino Acids in Patients with Urea Cycle Disorders

    OpenAIRE

    Burrage, Lindsay C.; Jain, Mahim; Gandolfo, Laura; Lee, Brendan H.; Nagamani, Sandesh CS

    2014-01-01

    Sodium phenylbutyrate (NaPBA) is a commonly used medication for the treatment of patients with urea cycle disorders (UCDs). Previous reports involving small numbers of patients with UCDs have shown that NaPBA treatment can result in lower plasma levels of the branched-chain amino acids (BCAA) but this has not been studied systematically. From a large cohort of patients (n=553) with UCDs enrolled in Longitudinal Study of Urea Cycle Disorders, a collaborative multicenter study of the Urea Cycle...

  9. Novel metabolic and physiological functions of branched chain amino acids: a review

    OpenAIRE

    Zhang, Shihai; Zeng, Xiangfang; Ren, Man; Mao, Xiangbing; Qiao, Shiyan

    2017-01-01

    It is widely known that branched chain amino acids (BCAA) are not only elementary components for building muscle tissue but also participate in increasing protein synthesis in animals and humans. BCAA (isoleucine, leucine and valine) regulate many key signaling pathways, the most classic of which is the activation of the mTOR signaling pathway. This signaling pathway connects many diverse physiological and metabolic roles. Recent years have witnessed many striking developments in determining ...

  10. Branched-chain amino acid aminotransferase and methionine formation in Mycobacterium tuberculosis

    OpenAIRE

    Radford Cynthia L; Knodel Marvin H; Venos Erik S; Berger Bradley J

    2004-01-01

    Abstract Background Tuberculosis remains a major world-wide health threat which demands the discovery and characterisation of new drug targets in order to develop future antimycobacterials. The regeneration of methionine consumed during polyamine biosynthesis is an important pathway present in many microorganisms. The final step of this pathway, the conversion of ketomethiobutyrate to methionine, can be performed by aspartate, tyrosine, or branched-chain amino acid aminotransferases depending...

  11. Imaging of branched chain amino acid metabolism in tumors with hyperpolarized 13C ketoisocaproate.

    Science.gov (United States)

    Karlsson, Magnus; Jensen, Pernille R; in 't Zandt, René; Gisselsson, Anna; Hansson, Georg; Duus, Jens Ø; Meier, Sebastian; Lerche, Mathilde H

    2010-08-01

    Powerful analytical tools are vital for characterizing the complex molecular changes underlying oncogenesis and cancer treatment. This is particularly true, if information is to be collected in vivo by noninvasive approaches. In the recent past, hyperpolarized (13)C magnetic resonance (MR) spectroscopy has been employed to quickly collect detailed spectral information on the chemical fate of tracer molecules in different tissues at high sensitivity. Here, we report a preclinical study showing that alpha-ketoisocaproic acid (KIC) can be used to assess molecular signatures of tumors with hyperpolarized MR spectroscopy. KIC is metabolized to leucine by the enzyme branched chain amino acid transferase (BCAT), which is found upregulated in some tumors. BCAT is a putative marker for metastasis and a target of the proto-oncogene c-myc. Very different fluxes through the BCAT-catalyzed reaction can be detected for murine lymphoma (EL4) and rat mammary adenocarcinoma (R3230AC) tumors in vivo. EL4 tumors show a more than 7-fold higher hyperpolarized (13)C leucine signal relative to the surrounding healthy tissue. In R3230AC tumor on the other hand branched chain amino acid metabolism is not enhanced relative to surrounding tissues. The distinct molecular signatures of branched chain amino acid metabolism in EL4 and R3230AC tumors correlate well with ex vivo assays of BCAT activity.

  12. Branched-chain amino acid aminotransferase and methionine formation in Mycobacterium tuberculosis

    Directory of Open Access Journals (Sweden)

    Radford Cynthia L

    2004-10-01

    Full Text Available Abstract Background Tuberculosis remains a major world-wide health threat which demands the discovery and characterisation of new drug targets in order to develop future antimycobacterials. The regeneration of methionine consumed during polyamine biosynthesis is an important pathway present in many microorganisms. The final step of this pathway, the conversion of ketomethiobutyrate to methionine, can be performed by aspartate, tyrosine, or branched-chain amino acid aminotransferases depending on the particular species examined. Results The gene encoding for branched-chain amino acid aminotransferase in Mycobacterium tuberculosis H37Rv has been cloned, expressed, and characterised. The enzyme was found to be a member of the aminotransferase IIIa subfamily, and closely related to the corresponding aminotransferase in Bacillus subtilis, but not to that found in B. anthracis or B. cereus. The amino donor preference for the formation of methionine from ketomethiobutyrate was for isoleucine, leucine, valine, glutamate, and phenylalanine. The enzyme catalysed branched-chain amino acid and ketomethiobutyrate transamination with a Km of 1.77 – 7.44 mM and a Vmax of 2.17 – 5.70 μmol/min/mg protein, and transamination of ketoglutarate with a Km of 5.79 – 6.95 mM and a Vmax of 11.82 – 14.35 μmol/min/mg protein. Aminooxy compounds were examined as potential enzyme inhibitors, with O-benzylhydroxylamine, O-t-butylhydroxylamine, carboxymethoxylamine, and O-allylhydroxylamine yielding mixed-type inhibition with Ki values of 8.20 – 21.61 μM. These same compounds were examined as antimycobacterial agents against M. tuberculosis and a lower biohazard M. marinum model system, and were found to completely prevent cell growth. O-Allylhydroxylamine was the most effective growth inhibitor with an MIC of 78 μM against M. marinum and one of 156 ��M against M. tuberculosis. Conclusion Methionine formation from ketomethiobutyrate is catalysed by a

  13. Metabolism of branched-chain amino acids in leg muscles from tail-cast suspended intact and adrenalectomized rats

    Science.gov (United States)

    Jaspers, Stephen R.; Henriksen, Erik; Jacob, Stephan; Tischler, Marc E.

    1989-01-01

    The effects of muscle unloading, adrenalectomy, and cortisol treatment on the metabolism of branched-chain amino acids in the soleus and extensor digitorum longus of tail-cast suspended rats were investigated using C-14-labeled lucine, isoleucine, and valine in incubation studies. It was found that, compared to not suspended controls, the degradation of branched-chain amino acids in hind limb muscles was accelerated in tail-cast suspended rats. Adrenalectomy was found to abolish the aminotransferase flux and to diminish the dehydrogenase flux in the soleus. The data also suggest that cortisol treatment increases the rate of metabolism of branched-chain amino acids at the dehydrogenase step.

  14. Branched chain amino acid transaminase and branched chain alpha-ketoacid dehydrogenase activity in the brain, liver and skele­tal muscle of acute hepatic failure rats

    Directory of Open Access Journals (Sweden)

    Takei,Nobuyuki

    1985-02-01

    Full Text Available Branched chain amino acid (BCAA transaminase activity increased in both the mitochondrial and supernatant fractions of brain from hepatic failure rats, in which a partial hepatectomy was performed 24h following carbon tetrachloride (CCl4 administration, although the activity of liver and skeletal muscle was the same as in control rats. The elevation of mitochondrial BCAA transaminase activity in liver-injured rats was partly due to increased activity of brain specific Type III isozyme. Branched chain alpha-ketoacid (BCKA dehydrogenase in the brain homogenates was not significantly altered in acute hepatic failure rats, while the liver enzyme activity was markedly diminished. BCKA dehydrogenase activity in the brain homogenates was inhibited by adding ATP to the assay system, and was activated in vitro by preincubating the brain homogenate at 37 degrees C for 15 min. These findings suggest that brain BCAA catabolism is accelerated in acute hepatic failure rats.

  15. Regulation of adipose branched chain amino acid catabolism enzyme expression and cross-adipose amino acid flux in human obesity

    Science.gov (United States)

    Elevated blood branched chain amino acids (BCAA) are often associated with insulin resistance and type 2 diabetes. One possibility is that under these conditions there is a reduced cellular utilization and/or lower complete oxidation of BCAAs. White adipose tissue (WAT) has become appreciated as a...

  16. Impaired adiponectin signaling contributes to disturbed catabolism of branched-chain amino acids in diabetic mice.

    Science.gov (United States)

    Lian, Kun; Du, Chaosheng; Liu, Yi; Zhu, Di; Yan, Wenjun; Zhang, Haifeng; Hong, Zhibo; Liu, Peilin; Zhang, Lijian; Pei, Haifeng; Zhang, Jinglong; Gao, Chao; Xin, Chao; Cheng, Hexiang; Xiong, Lize; Tao, Ling

    2015-01-01

    The branched-chain amino acids (BCAA) accumulated in type 2 diabetes are independent contributors to insulin resistance. The activity of branched-chain α-keto acid dehydrogenase (BCKD) complex, rate-limiting enzyme in BCAA catabolism, is reduced in diabetic states, which contributes to elevated BCAA concentrations. However, the mechanisms underlying decreased BCKD activity remain poorly understood. Here, we demonstrate that mitochondrial phosphatase 2C (PP2Cm), a newly identified BCKD phosphatase that increases BCKD activity, was significantly downregulated in ob/ob and type 2 diabetic mice. Interestingly, in adiponectin (APN) knockout (APN(-/-)) mice fed with a high-fat diet (HD), PP2Cm expression and BCKD activity were significantly decreased, whereas BCKD kinase (BDK), which inhibits BCKD activity, was markedly increased. Concurrently, plasma BCAA and branched-chain α-keto acids (BCKA) were significantly elevated. APN treatment markedly reverted PP2Cm, BDK, BCKD activity, and BCAA and BCKA levels in HD-fed APN(-/-) and diabetic animals. Additionally, increased BCKD activity caused by APN administration was partially but significantly inhibited in PP2Cm knockout mice. Finally, APN-mediated upregulation of PP2Cm expression and BCKD activity were abolished when AMPK was inhibited. Collectively, we have provided the first direct evidence that APN is a novel regulator of PP2Cm and systematic BCAA levels, suggesting that targeting APN may be a pharmacological approach to ameliorating BCAA catabolism in the diabetic state.

  17. Branched-chain amino acid metabolism in rat muscle: abnormal regulation in acidosis

    Energy Technology Data Exchange (ETDEWEB)

    May, R.C.; Hara, Y.; Kelly, R.A.; Block, K.P.; Buse, M.G.; Mitch, W.E.

    1987-06-01

    Branched-chain amino acid (BCAA) metabolism is frequently abnormal in pathological conditions accompanied by chronic metabolic acidosis. To study how metabolic acidosis affects BCAA metabolism in muscle, rats were gavage fed a 14% protein diet with or without 4 mmol NH/sub 4/Cl x 100 g body wt/sup -1/ x day/sup -1/. Epitrochlearis muscles were incubated with L-(1-/sup 14/C)-valine and L-(1-/sup 14/C)leucine, and rates of decarboxylation, net transamination, and incorporation into muscle protein were measured. Plasma and muscle BCAA levels were lower in acidotic rats. Rates of valine and leucine decarboxylation and net transamination were higher in muscles from acidotic rats; these differences were associated with a 79% increase in the total activity of branched-chain ..cap alpha..-keto acid dehydrogenase and a 146% increase in the activated form of the enzyme. They conclude that acidosis affects the regulation of BCAA metabolism by enhancing flux through the transaminase and by directly stimulating oxidative catabolism through activation of branched-chain ..cap alpha..-keto acid dehydrogenase.

  18. Progress in application of branched-chain amino acids in patients with liver cirrhosis

    Directory of Open Access Journals (Sweden)

    LYU Zheng

    2015-03-01

    Full Text Available The metabolism of amino acids mainly takes place in the liver, and patients with liver cirrhosis may develop metabolic disorders of proteins, carbohydrates, fats, and amino acids, which in turn causes dysfunction of multiple organs and systems, as well as complications like hepatic encephalopathy, esophageal variceal bleeding, and ascites, resulting in high mortality. This paper summarizes the metabolic characteristics of amino acids and the application of branched-chain amino acids (BCAAs in the treatment of liver cirrhosis, and it points out the significance of BCAAs in regulating serum aminogram, increasing the ratio of BCAAs to aromatic amino acids, preventing complications of liver cirrhosis, and improving the quality of life for patients with the disease.

  19. Interactions in the Metabolism of Glutamate and the Branched-Chain Amino Acids and Ketoacids in the CNS.

    Science.gov (United States)

    Yudkoff, Marc

    2017-01-01

    Glutamatergic neurotransmission entails a tonic loss of glutamate from nerve endings into the synapse. Replacement of neuronal glutamate is essential in order to avoid depletion of the internal pool. In brain this occurs primarily via the glutamate-glutamine cycle, which invokes astrocytic synthesis of glutamine and hydrolysis of this amino acid via neuronal phosphate-dependent glutaminase. This cycle maintains constancy of internal pools, but it does not provide a mechanism for inevitable losses of glutamate N from brain. Import of glutamine or glutamate from blood does not occur to any appreciable extent. However, the branched-chain amino acids (BCAA) cross the blood-brain barrier swiftly. The brain possesses abundant branched-chain amino acid transaminase activity which replenishes brain glutamate and also generates branched-chain ketoacids. It seems probable that the branched-chain amino acids and ketoacids participate in a "glutamate-BCAA cycle" which involves shuttling of branched-chain amino acids and ketoacids between astrocytes and neurons. This mechanism not only supports the synthesis of glutamate, it also may constitute a mechanism by which high (and potentially toxic) concentrations of glutamate can be avoided by the re-amination of branched-chain ketoacids.

  20. PGC-1α-mediated branched-chain amino acid metabolism in the skeletal muscle.

    Directory of Open Access Journals (Sweden)

    Yukino Hatazawa

    Full Text Available Peroxisome proliferator-activated receptor (PPAR γ coactivator 1α (PGC-1α is a coactivator of various nuclear receptors and other transcription factors, which is involved in the regulation of energy metabolism, thermogenesis, and other biological processes that control phenotypic characteristics of various organ systems including skeletal muscle. PGC-1α in skeletal muscle is considered to be involved in contractile protein function, mitochondrial function, metabolic regulation, intracellular signaling, and transcriptional responses. Branched-chain amino acid (BCAA metabolism mainly occurs in skeletal muscle mitochondria, and enzymes related to BCAA metabolism are increased by exercise. Using murine skeletal muscle overexpressing PGC-1α and cultured cells, we investigated whether PGC-1α stimulates BCAA metabolism by increasing the expression of enzymes involved in BCAA metabolism. Transgenic mice overexpressing PGC-1α specifically in the skeletal muscle had increased the expression of branched-chain aminotransferase (BCAT 2, branched-chain α-keto acid dehydrogenase (BCKDH, which catabolize BCAA. The expression of BCKDH kinase (BCKDK, which phosphorylates BCKDH and suppresses its enzymatic activity, was unchanged. The amount of BCAA in the skeletal muscle was significantly decreased in the transgenic mice compared with that in the wild-type mice. The amount of glutamic acid, a metabolite of BCAA catabolism, was increased in the transgenic mice, suggesting the activation of muscle BCAA metabolism by PGC-1α. In C2C12 cells, the overexpression of PGC-1α significantly increased the expression of BCAT2 and BCKDH but not BCKDK. Thus, PGC-1α in the skeletal muscle is considered to significantly contribute to BCAA metabolism.

  1. PGC-1α-mediated branched-chain amino acid metabolism in the skeletal muscle.

    Science.gov (United States)

    Hatazawa, Yukino; Tadaishi, Miki; Nagaike, Yuta; Morita, Akihito; Ogawa, Yoshihiro; Ezaki, Osamu; Takai-Igarashi, Takako; Kitaura, Yasuyuki; Shimomura, Yoshiharu; Kamei, Yasutomi; Miura, Shinji

    2014-01-01

    Peroxisome proliferator-activated receptor (PPAR) γ coactivator 1α (PGC-1α) is a coactivator of various nuclear receptors and other transcription factors, which is involved in the regulation of energy metabolism, thermogenesis, and other biological processes that control phenotypic characteristics of various organ systems including skeletal muscle. PGC-1α in skeletal muscle is considered to be involved in contractile protein function, mitochondrial function, metabolic regulation, intracellular signaling, and transcriptional responses. Branched-chain amino acid (BCAA) metabolism mainly occurs in skeletal muscle mitochondria, and enzymes related to BCAA metabolism are increased by exercise. Using murine skeletal muscle overexpressing PGC-1α and cultured cells, we investigated whether PGC-1α stimulates BCAA metabolism by increasing the expression of enzymes involved in BCAA metabolism. Transgenic mice overexpressing PGC-1α specifically in the skeletal muscle had increased the expression of branched-chain aminotransferase (BCAT) 2, branched-chain α-keto acid dehydrogenase (BCKDH), which catabolize BCAA. The expression of BCKDH kinase (BCKDK), which phosphorylates BCKDH and suppresses its enzymatic activity, was unchanged. The amount of BCAA in the skeletal muscle was significantly decreased in the transgenic mice compared with that in the wild-type mice. The amount of glutamic acid, a metabolite of BCAA catabolism, was increased in the transgenic mice, suggesting the activation of muscle BCAA metabolism by PGC-1α. In C2C12 cells, the overexpression of PGC-1α significantly increased the expression of BCAT2 and BCKDH but not BCKDK. Thus, PGC-1α in the skeletal muscle is considered to significantly contribute to BCAA metabolism.

  2. Decreased Consumption of Branched-Chain Amino Acids Improves Metabolic Health

    Directory of Open Access Journals (Sweden)

    Luigi Fontana

    2016-07-01

    Full Text Available Protein-restricted (PR, high-carbohydrate diets improve metabolic health in rodents, yet the precise dietary components that are responsible for these effects have not been identified. Furthermore, the applicability of these studies to humans is unclear. Here, we demonstrate in a randomized controlled trial that a moderate PR diet also improves markers of metabolic health in humans. Intriguingly, we find that feeding mice a diet specifically reduced in branched-chain amino acids (BCAAs is sufficient to improve glucose tolerance and body composition equivalently to a PR diet via metabolically distinct pathways. Our results highlight a critical role for dietary quality at the level of amino acids in the maintenance of metabolic health and suggest that diets specifically reduced in BCAAs, or pharmacological interventions in this pathway, may offer a translatable way to achieve many of the metabolic benefits of a PR diet.

  3. Branched-chain amino acids in metabolic signalling and insulin resistance

    Science.gov (United States)

    Lynch, Christopher J.; Adams, Sean H.

    2015-01-01

    Branched-chain amino acids (BCAAs) are important nutrient signals that have direct and indirect effects. Frequently, BCAAs have been reported to mediate antiobesity effects, especially in rodent models. However, circulating levels of BCAAs tend to be increased in individuals with obesity and are associated with worse metabolic health and future insulin resistance or type 2 diabetes mellitus (T2DM). A hypothesized mechanism linking increased levels of BCAAs and T2DM involves leucine-mediated activation of the mammalian target of rapamycin complex 1 (mTORC1), which results in uncoupling of insulin signalling at an early stage. A BCAA dysmetabolism model proposes that the accumulation of mitotoxic metabolites (and not BCAAs per se) promotes β-cell mitochondrial dysfunction, stress signalling and apoptosis associated with T2DM. Alternatively, insulin resistance might promote aminoacidaemia by increasing the protein degradation that insulin normally suppresses, and/or by eliciting an impairment of efficient BCAA oxidative metabolism in some tissues. Whether and how impaired BCAA metabolism might occur in obesity is discussed in this Review. Research on the role of individual and model-dependent differences in BCAA metabolism is needed, as several genes (BCKDHA, PPM1K, IVD and KLF15) have been designated as candidate genes for obesity and/or T2DM in humans, and distinct phenotypes of tissue-specific branched chain ketoacid dehydrogenase complex activity have been detected in animal models of obesity and T2DM. PMID:25287287

  4. A pilot, short-term dietary manipulation of branched chain amino acids has modest influence on fasting levels of branched chain amino acids

    Directory of Open Access Journals (Sweden)

    Nicole Landa Cavallaro

    2016-01-01

    Full Text Available Background: Elevated fasting levels of branched chain amino acids (BCAAs: valine, isoleucine, leucine in venous blood are associated with a variety of metabolic impairments, including increased risk of type 2 diabetes (T2D. Fasting BCAA levels are influenced by non-dietary factors. However, it is unknown whether fasting BCAAs can be altered through manipulation of dietary intake alone. Objective: To test whether a specific dietary intervention, using differences in BCAA intake, alters fasting BCAA levels independent of other factors. Design: Five healthy male volunteers underwent 4 days of a low and 4 days of a high BCAA content dietary intervention (ClinicalTrials.gov [NCT02110602]. All food and supplements were provided. Fasting BCAAs were measured from venous blood samples by mass spectrometry at baseline and after each intervention. Results: Diets were isocaloric; contained equal percentages of calories from carbohydrate, fats, and protein; and differed from each other in BCAA content (1.5±0.1 vs. 14.0±0.6 g for valine; 4.5±0.9 g vs. 13.8±0.5 g for isoleucine; 2.1±0.2 g vs. 27.1±1.0 g for leucine; p<0.0001 for all. Fasting valine was significantly lower (p=0.02 and fasting isoleucine and leucine were numerically lower following the low BCAA content vs. the high BCAA content diet levels. The inter-individual response to the dietary interventions was variable and not explained by adherence. Conclusion: Short-term dietary manipulation of BCAA intake led to modest changes in fasting levels of BCAAs. The approach from our pilot study can be expanded to test the metabolic implications of dietary BCAA manipulation.

  5. Defective branched chain amino acid catabolism contributes to cardiac dysfunction and remodeling following myocardial infarction.

    Science.gov (United States)

    Wang, Wei; Zhang, Fuyang; Xia, Yunlong; Zhao, Shihao; Yan, Wenjun; Wang, Helin; Lee, Yan; Li, Congye; Zhang, Ling; Lian, Kun; Gao, Erhe; Cheng, Hexiang; Tao, Ling

    2016-11-01

    Cardiac metabolic remodeling is a central event during heart failure (HF) development following myocardial infarction (MI). It is well known that myocardial glucose and fatty acid dysmetabolism contribute to post-MI cardiac dysfunction and remodeling. However, the role of amino acid metabolism in post-MI HF remains elusive. Branched chain amino acids (BCAAs) are an important group of essential amino acids and function as crucial nutrient signaling in mammalian animals. The present study aimed to determine the role of cardiac BCAA metabolism in post-MI HF progression. Utilizing coronary artery ligation-induced murine MI models, we found that myocardial BCAA catabolism was significantly impaired in response to permanent MI, therefore leading to an obvious elevation of myocardial BCAA abundance. In MI-operated mice, oral BCAA administration further increased cardiac BCAA levels, activated the mammalian target of rapamycin (mTOR) signaling, and exacerbated cardiac dysfunction and remodeling. These data demonstrate that BCAAs act as a direct contributor to post-MI cardiac pathologies. Furthermore, these BCAA-mediated deleterious effects were improved by rapamycin cotreatment, revealing an indispensable role of mTOR in BCAA-mediated adverse effects on cardiac function/structure post-MI. Of note, pharmacological inhibition of branched chain ketoacid dehydrogenase kinase (BDK), a negative regulator of myocardial BCAA catabolism, significantly improved cardiac BCAA catabolic disorders, reduced myocardial BCAA levels, and ameliorated post-MI cardiac dysfunction and remodeling. In conclusion, our data provide the evidence that impaired cardiac BCAA catabolism directly contributes to post-MI cardiac dysfunction and remodeling. Moreover, improving cardiac BCAA catabolic defects may be a promising therapeutic strategy against post-MI HF.

  6. Purification and characterization of a branched-chain amino acid aminotransferase from Lactobacillus paracasei subsp paracasei CHCC 2115

    DEFF Research Database (Denmark)

    Thage, B.V.; Rattray, F.P.; Laustsen, M.W.;

    2004-01-01

    Purification and characterization of an aminotransferase (AT) specific for the degradation of branched-chain amino acids from Lactobacillus paracasei subsp. paracasei CHCC 2115. Methods and Results: The purification protocol consisted of anion exchange chromatography, affinity chromatography...... of other metal ions, thiol- and carbonyl-binding agents. The N-terminal sequence of the enzyme was SVNIDWNNLGFDYMQLPYRYVAHXKDGVXD, and had at the amino acid level, 60 and 53% identity to a branched-chain amino acid AT of Lact. plantarum and Lactococcus lactis, respectively. Conclusions: The results suggest...

  7. Study on color identification for monitoring and controlling fermentation process of branched chain amino acid

    Science.gov (United States)

    Ma, Lei; Wang, Yizhong; Chen, Ning; Liu, Tiegen; Xu, Qingyang; Kong, Fanzhi

    2008-12-01

    In this paper, a new method for monitoring and controlling fermentation process of branched chain amino acid (BCAA) was proposed based on color identification. The color image of fermentation broth of BCAA was firstly taken by a CCD camera. Then, it was changed from RGB color model to HIS color model. Its histograms of hue H and saturation S were calculated, which were used as the input of a designed BP network. The output of the BP network was the description of the color of fermentation broth of BCAA. After training, the color of fermentation broth was identified by the BP network according to the histograms of H and S of a fermentation broth image. Along with other parameters, the fermentation process of BCAA was monitored and controlled to start the stationary phase of fermentation soon. Experiments were conducted with satisfied results to show the feasibility and usefulness of color identification of fermentation broth in fermentation process control of BCAA.

  8. Genetic evidence of a causal effect of insulin resistance on branched-chain amino acid levels

    DEFF Research Database (Denmark)

    Mahendran, Yuvaraj; Jonsson, Anna; Have, Christian T

    2017-01-01

    AIMS/HYPOTHESIS: Fasting plasma levels of branched-chain amino acids (BCAAs) are associated with insulin resistance, but it remains unclear whether there is a causal relation between the two. We aimed to disentangle the causal relations by performing a Mendelian randomisation study using genetic...... variants associated with circulating BCAA levels and insulin resistance as instrumental variables. METHODS: We measured circulating BCAA levels in blood plasma by NMR spectroscopy in 1,321 individuals from the ADDITION-PRO cohort. We complemented our analyses by using previously published genome...... variable for insulin resistance. A GRS of three variants increasing circulating BCAA levels was used as an instrumental variable for circulating BCAA levels. RESULTS: Fasting plasma BCAA levels were associated with higher HOMA-IR in ADDITION-PRO (β 0.137 [95% CI 0.08, 0.19] p = 6 × 10(-7)). However...

  9. Fermentative production of branched chain amino acids: a focus on metabolic engineering.

    Science.gov (United States)

    Park, Jin Hwan; Lee, Sang Yup

    2010-01-01

    The branched chain amino acids (BCAAs), L-valine, L-leucine, and L-isoleucine, have recently been attracting much attention as their potential to be applied in various fields, including animal feed additive, cosmetics, and pharmaceuticals, increased. Strategies for developing microbial strains efficiently producing BCAAs are now in transition toward systems metabolic engineering from random mutagenesis. The metabolism and regulatory circuits of BCAA biosynthesis need to be thoroughly understood for designing system-wide metabolic engineering strategies. Here we review the current knowledge on BCAAs including their biosynthetic pathways, regulations, and export and transport systems. Recent advances in the development of BCAA production strains are also reviewed with a particular focus on L-valine production strain. At the end, the general strategies for developing BCAA overproducers by systems metabolic engineering are suggested.

  10. The dietary branched chain amino acid requirements of hybrid striped bass(Morone chrysops x M. saxatilis)

    Science.gov (United States)

    The requirements for branched chain amino acids (BCAAs) are unknown in hybrid striped bass and necessary for formulating efficient and nutritious diets. Moreover, the dietary balance among these three amino acids can substantially influence the performance of meat animals fed those diets. The diet...

  11. Removal of branched-chain amino acids and alpha-ketoisocaproate by haemofiltration and haemodiafiltration.

    Science.gov (United States)

    Gouyon, J B; Semama, D; Prévot, A; Desgres, J

    1996-01-01

    Venovenous haemofiltration (VVHF) and haemodiafiltration (VVHDF) were performed with a neonatal haemo(dia)filter (Miniflow 10, Hospal) on 8 anaesthetized rabbits infused with branched-chain amino acids (leucine, isoleucine and valine) and alpha-ketoisocaproate. The branched-chain amino acids (BCAA) and alpha-ketoisocaproate blood levels were close to those previously observed in neonates with maple syrup urine disease when extracorporeal blood purification was required. VVHF and VVHDF performances were assessed with two different blood flows (Qb = 8.3 and 16.6 ml/min). VVHDF was performed with four dialysate flow rates (Qd = 0.5, 1.0, 2.0 and 3.0 L/h). Within each period, clearances of the three BCAA were strictly similar. BCAA clearances obtained by VVHF were similar to ultrafiltration rates (respectively, 0.78 +/- 0.14 and 1.79 +/- 0.28 ml/min at high and low Qb; p < 0.05). The alpha-ketoisocaproate clearances obtained by VVHF were 0.39 +/- 0.17 and 0.92 +/- 0.43 ml/min at low and high Qb (not significantly different). Whatever the Qd value, the VVHDF procedures always allowed higher BCAA and alpha-ketoisocaproate clearances as compared with the corresponding VVHF period with similar Qb. BCAA clearances obtained by VVHDF with a 0.5 L/h dialysate flow were 4.1 +/- 0.5 and 5.4 +/- 0.5 mL/min at low and high Qb, respectively. The concurrent alpha-ketoisocaproate clearances were 2.5 +/- 0.8 and 2.9 +/- 1.0 ml/min.

  12. Branched-chain and aromatic amino acid catabolism into aroma volatiles in Cucumis melo L. fruit.

    Science.gov (United States)

    Gonda, Itay; Bar, Einat; Portnoy, Vitaly; Lev, Shery; Burger, Joseph; Schaffer, Arthur A; Tadmor, Ya'akov; Gepstein, Shimon; Giovannoni, James J; Katzir, Nurit; Lewinsohn, Efraim

    2010-02-01

    The unique aroma of melons (Cucumis melo L., Cucurbitaceae) is composed of many volatile compounds biosynthetically derived from fatty acids, carotenoids, amino acids, and terpenes. Although amino acids are known precursors of aroma compounds in the plant kingdom, the initial steps in the catabolism of amino acids into aroma volatiles have received little attention. Incubation of melon fruit cubes with amino acids and alpha-keto acids led to the enhanced formation of aroma compounds bearing the side chain of the exogenous amino or keto acid supplied. Moreover, L-[(13)C(6)]phenylalanine was also incorporated into aromatic volatile compounds. Amino acid transaminase activities extracted from the flesh of mature melon fruits converted L-isoleucine, L-leucine, L-valine, L-methionine, or L-phenylalanine into their respective alpha-keto acids, utilizing alpha-ketoglutarate as the amine acceptor. Two novel genes were isolated and characterized (CmArAT1 and CmBCAT1) encoding 45.6 kDa and 42.7 kDa proteins, respectively, that displayed aromatic and branched-chain amino acid transaminase activities, respectively, when expressed in Escherichia coli. The expression of CmBCAT1 and CmArAT1 was low in vegetative tissues, but increased in flesh and rind tissues during fruit ripening. In addition, ripe fruits of climacteric aromatic cultivars generally showed high expression of CmBCAT1 and CmArAT1 in contrast to non-climacteric non-aromatic fruits. The results presented here indicate that in melon fruit tissues, the catabolism of amino acids into aroma volatiles can initiate through a transamination mechanism, rather than decarboxylation or direct aldehyde synthesis, as has been demonstrated in other plants.

  13. Branched-chain amino acid supplementation promotes aerobic growth of Salmonella Typhimurium under nitrosative stress conditions.

    Science.gov (United States)

    Park, Yoon Mee; Lee, Hwa Jeong; Jeong, Jae-Ho; Kook, Joong-Ki; Choy, Hyon E; Hahn, Tae-Wook; Bang, Iel Soo

    2015-12-01

    Nitric oxide (NO) inactivates iron-sulfur enzymes in bacterial amino acid biosynthetic pathways, causing amino acid auxotrophy. We demonstrate that exogenous supplementation with branched-chain amino acids (BCAA) can restore the NO resistance of hmp mutant Salmonella Typhimurium lacking principal NO-metabolizing enzyme flavohemoglobin, and of mutants further lacking iron-sulfur enzymes dihydroxy-acid dehydratase (IlvD) and isopropylmalate isomerase (LeuCD) that are essential for BCAA biosynthesis, in an oxygen-dependent manner. BCAA supplementation did not affect the NO consumption rate of S. Typhimurium, suggesting the BCAA-promoted NO resistance independent of NO metabolism. BCAA supplementation also induced intracellular survival of ilvD and leuCD mutants at wild-type levels inside RAW 264.7 macrophages that produce constant amounts of NO regardless of varied supplemental BCAA concentrations. Our results suggest that the NO-induced BCAA auxotrophy of Salmonella, due to inactivation of iron-sulfur enzymes for BCAA biosynthesis, could be rescued by bacterial taking up exogenous BCAA available in oxic environments.

  14. Genetic analysis of pathway regulation for enhancing branched-chain amino acid biosynthesis in plants

    KAUST Repository

    Chen, Hao

    2010-08-01

    The branched-chain amino acids (BCAAs) valine, leucine and isoleucine are essential amino acids that play critical roles in animal growth and development. Animals cannot synthesize these amino acids and must obtain them from their diet. Plants are the ultimate source of these essential nutrients, and they synthesize BCAAs through a conserved pathway that is inhibited by its end products. This feedback inhibition has prevented scientists from engineering plants that accumulate high levels of BCAAs by simply over-expressing the respective biosynthetic genes. To identify components critical for this feedback regulation, we performed a genetic screen for Arabidopsis mutants that exhibit enhanced resistance to BCAAs. Multiple dominant allelic mutations in the VALINE-TOLERANT 1 (VAT1) gene were identified that conferred plant resistance to valine inhibition. Map-based cloning revealed that VAT1 encodes a regulatory subunit of acetohydroxy acid synthase (AHAS), the first committed enzyme in the BCAA biosynthesis pathway. The VAT1 gene is highly expressed in young, rapidly growing tissues. When reconstituted with the catalytic subunit in vitro, the vat1 mutant-containing AHAS holoenzyme exhibits increased resistance to valine. Importantly, transgenic plants expressing the mutated vat1 gene exhibit valine tolerance and accumulate higher levels of BCAAs. Our studies not only uncovered regulatory characteristics of plant AHAS, but also identified a method to enhance BCAA accumulation in crop plants that will significantly enhance the nutritional value of food and feed. © 2010 Blackwell Publishing Ltd.

  15. Branched-chain amino acid supplementation during bed rest: effect on recovery

    Science.gov (United States)

    Stein, T. P.; Donaldson, M. R.; Leskiw, M. J.; Schluter, M. D.; Baggett, D. W.; Boden, G.

    2003-01-01

    Bed rest is associated with a loss of protein from the weight-bearing muscle. The objectives of this study are to determine whether increasing dietary branched-chain amino acids (BCAAs) during bed rest improves the anabolic response after bed rest. The study consisted of a 1-day ambulatory period, 14 days of bed rest, and a 4-day recovery period. During bed rest, dietary intake was supplemented with either 30 mmol/day each of glycine, serine, and alanine (group 1) or with 30 mmol/day each of the three BCAAs (group 2). Whole body protein synthesis was determined with U-(15)N-labeled amino acids, muscle, and selected plasma protein synthesis with l-[(2)H(5)]phenylalanine. Total glucose production and gluconeogenesis from alanine were determined with l-[U-(13)C(3)]alanine and [6,6-(2)H(2)]glucose. During bed rest, nitrogen (N) retention was greater with BCAA feeding (56 +/- 6 vs. 26 +/- 12 mg N. kg(-1). day(-1), P BCAA supplementation on either whole body, muscle, or plasma protein synthesis or the rate of 3-MeH excretion. Muscle tissue free amino acid concentrations were increased during bed rest with BCAA (0.214 +/- 0.066 vs. 0.088 +/- 0.12 nmol/mg protein, P BCAA group in the recovery phase. In conclusion, the improved N retention during bed rest is due, at least in part, to accretion of amino acids in the tissue free amino acid pools. The amount accreted is not enough to impact protein kinetics in the recovery phase but does improve N retention by providing additional essential amino acids in the early recovery phase.

  16. Branched-chain and aromatic amino acids, insulin resistance and liver specific ectopic fat storage in overweight to obese subjects.

    Science.gov (United States)

    Haufe, S; Witt, H; Engeli, S; Kaminski, J; Utz, W; Fuhrmann, J C; Rein, D; Schulz-Menger, J; Luft, F C; Boschmann, M; Jordan, J

    2016-07-01

    Amino acids may interfere with insulin action, particularly in obese individuals. We hypothesized that increased circulating branched-chain and aromatic amino acids herald insulin resistance and ectopic fat storage, particularly hepatic fat accumulation. We measured fasting branched-chain and aromatic amino acids (tryptophan, tyrosine, and phenylalanine) by mass spectrometry in 111 overweight to obese subjects. We applied abdominal magnetic resonance imaging and spectroscopy to assess adipose tissue distribution and ectopic fat storage, respectively. Plasma branched-chain amino acids concentrations were related to insulin sensitivity and intrahepatic fat independent from adiposity, age and gender, but not to abdominal adipose tissue or intramyocellular fat. In weight stable overweight and obese individuals, branched-chain amino acid concentrations are specifically associated with hepatic fat storage and insulin resistance. Copyright © 2016 The Italian Society of Diabetology, the Italian Society for the Study of Atherosclerosis, the Italian Society of Human Nutrition, and the Department of Clinical Medicine and Surgery, Federico II University. Published by Elsevier B.V. All rights reserved.

  17. Attenuation of the protein wasting associated with bed rest by branched-chain amino acids

    Science.gov (United States)

    Stein, T. P.; Schluter, M. D.; Leskiw, M. J.; Boden, G.

    1999-01-01

    Bed rest is generally accepted as being an appropriate ground-based model for human spaceflight. The objectives of this study were to test the hypothesis that increasing the amount of branched-chain amino acids (BCAAs) in the diet could attenuate the protein loss associated with bed rest. Nineteen healthy subjects were randomized into two groups according to diet. During the 6 d of bed rest, the diets were supplemented with either 30 mmol/d each of three non-essential amino acids, glycine, serine, and alanine (control group), or with 30 mmol/d each of the BCAAs, leucine, isoleucine, and valine (BCAA group). Nutrition was supplied as a commercially available defined formula diet at a rate of 1.3 x REE. Nitrogen (N) balance and urinary 3-MeH excretion were determined for the 6 d. In our results, the urine-based estimate of N balance was 22.2 +/- 14.4 (n = 9) mg N.kg-1.d-1 and 60.5 +/- 10.1 mg (n = 8) N.kg-1.d-1 for the control and BCAA-supplemented groups, respectively (P < 0.05). Urinary 3-MeH excretion was unchanged in both groups with bed rest. We conclude that BCAA supplementation attenuates the N loss during short-term bed rest.

  18. Branched-chain amino acid catabolism fuels adipocyte differentiation and lipogenesis.

    Science.gov (United States)

    Green, Courtney R; Wallace, Martina; Divakaruni, Ajit S; Phillips, Susan A; Murphy, Anne N; Ciaraldi, Theodore P; Metallo, Christian M

    2016-01-01

    Adipose tissue plays important roles in regulating carbohydrate and lipid homeostasis, but less is known about the regulation of amino acid metabolism in adipocytes. Here we applied isotope tracing to pre-adipocytes and differentiated adipocytes to quantify the contributions of different substrates to tricarboxylic acid (TCA) metabolism and lipogenesis. In contrast to proliferating cells, which use glucose and glutamine for acetyl-coenzyme A (AcCoA) generation, differentiated adipocytes showed increased branched-chain amino acid (BCAA) catabolic flux such that leucine and isoleucine from medium and/or from protein catabolism accounted for as much as 30% of lipogenic AcCoA pools. Medium cobalamin deficiency caused methylmalonic acid accumulation and odd-chain fatty acid synthesis. Vitamin B12 supplementation reduced these metabolites and altered the balance of substrates entering mitochondria. Finally, inhibition of BCAA catabolism compromised adipogenesis. These results quantitatively highlight the contribution of BCAAs to adipocyte metabolism and suggest that BCAA catabolism has a functional role in adipocyte differentiation.

  19. The Emerging Role of Branched-Chain Amino Acids in Insulin Resistance and Metabolism

    Directory of Open Access Journals (Sweden)

    Mee-Sup Yoon

    2016-07-01

    Full Text Available Insulin is required for maintenance of glucose homeostasis. Despite the importance of insulin sensitivity to metabolic health, the mechanisms that induce insulin resistance remain unclear. Branched-chain amino acids (BCAAs belong to the essential amino acids, which are both direct and indirect nutrient signals. Even though BCAAs have been reported to improve metabolic health, an increased BCAA plasma level is associated with a high risk of metabolic disorder and future insulin resistance, or type 2 diabetes mellitus (T2DM. The activation of mammalian target of rapamycin complex 1 (mTORC1 by BCAAs has been suggested to cause insulin resistance. In addition, defective BCAA oxidative metabolism might occur in obesity, leading to a further accumulation of BCAAs and toxic intermediates. This review provides the current understanding of the mechanism of BCAA-induced mTORC1 activation, as well as the effect of mTOR activation on metabolic health in terms of insulin sensitivity. Furthermore, the effects of impaired BCAA metabolism will be discussed in detail.

  20. Novel metabolic and physiological functions of branched chain amino acids: a review.

    Science.gov (United States)

    Zhang, Shihai; Zeng, Xiangfang; Ren, Man; Mao, Xiangbing; Qiao, Shiyan

    2017-01-01

    It is widely known that branched chain amino acids (BCAA) are not only elementary components for building muscle tissue but also participate in increasing protein synthesis in animals and humans. BCAA (isoleucine, leucine and valine) regulate many key signaling pathways, the most classic of which is the activation of the mTOR signaling pathway. This signaling pathway connects many diverse physiological and metabolic roles. Recent years have witnessed many striking developments in determining the novel functions of BCAA including: (1) Insufficient or excessive levels of BCAA in the diet enhances lipolysis. (2) BCAA, especially isoleucine, play a major role in enhancing glucose consumption and utilization by up-regulating intestinal and muscular glucose transporters. (3) Supplementation of leucine in the diet enhances meat quality in finishing pigs. (4) BCAA are beneficial for mammary health, milk quality and embryo growth. (5) BCAA enhance intestinal development, intestinal amino acid transportation and mucin production. (6) BCAA participate in up-regulating innate and adaptive immune responses. In addition, abnormally elevated BCAA levels in the blood (decreased BCAA catabolism) are a good biomarker for the early detection of obesity, diabetes and other metabolic diseases. This review will provide some insights into these novel metabolic and physiological functions of BCAA.

  1. Attenuation of the protein wasting associated with bed rest by branched-chain amino acids

    Science.gov (United States)

    Stein, T. P.; Schluter, M. D.; Leskiw, M. J.; Boden, G.

    1999-01-01

    Bed rest is generally accepted as being an appropriate ground-based model for human spaceflight. The objectives of this study were to test the hypothesis that increasing the amount of branched-chain amino acids (BCAAs) in the diet could attenuate the protein loss associated with bed rest. Nineteen healthy subjects were randomized into two groups according to diet. During the 6 d of bed rest, the diets were supplemented with either 30 mmol/d each of three non-essential amino acids, glycine, serine, and alanine (control group), or with 30 mmol/d each of the BCAAs, leucine, isoleucine, and valine (BCAA group). Nutrition was supplied as a commercially available defined formula diet at a rate of 1.3 x REE. Nitrogen (N) balance and urinary 3-MeH excretion were determined for the 6 d. In our results, the urine-based estimate of N balance was 22.2 +/- 14.4 (n = 9) mg N.kg-1.d-1 and 60.5 +/- 10.1 mg (n = 8) N.kg-1.d-1 for the control and BCAA-supplemented groups, respectively (P BCAA supplementation attenuates the N loss during short-term bed rest.

  2. Sodium phenylbutyrate decreases plasma branched-chain amino acids in patients with urea cycle disorders.

    Science.gov (United States)

    Burrage, Lindsay C; Jain, Mahim; Gandolfo, Laura; Lee, Brendan H; Nagamani, Sandesh C S

    2014-01-01

    Sodium phenylbutyrate (NaPBA) is a commonly used medication for the treatment of patients with urea cycle disorders (UCDs). Previous reports involving small numbers of patients with UCDs have shown that NaPBA treatment can result in lower plasma levels of the branched-chain amino acids (BCAA) but this has not been studied systematically. From a large cohort of patients (n=553) with UCDs enrolled in the Longitudinal Study of Urea Cycle Disorders, a collaborative multicenter study of the Urea Cycle Disorders Consortium, we evaluated whether treatment with NaPBA leads to a decrease in plasma BCAA levels. Our analysis shows that NaPBA use independently affects the plasma BCAA levels even after accounting for multiple confounding covariates. Moreover, NaPBA use increases the risk for BCAA deficiency. This effect of NaPBA seems specific to plasma BCAA levels, as levels of other essential amino acids are not altered by its use. Our study, in an unselected population of UCD subjects, is the largest to analyze the effects of NaPBA on BCAA metabolism and potentially has significant clinical implications. Our results indicate that plasma BCAA levels should to be monitored in patients treated with NaPBA since patients taking the medication are at increased risk for BCAA deficiency. On a broader scale, these findings could open avenues to explore NaPBA as a therapy in maple syrup urine disease and other common complex disorders with dysregulation of BCAA metabolism.

  3. Branched-chain amino acids and ammonia metabolism in liver disease: therapeutic implications.

    Science.gov (United States)

    Holecek, Milan

    2013-10-01

    The rationale for recommendation of branched-chain amino acids (BCAA; valine, leucine, and isoleucine) in treatment of liver failure is based on their unique pharmacologic properties, stimulatory effect on ammonia detoxification to glutamine (GLN), and decreased concentrations in liver cirrhosis. Multiple lines of evidence have shown that the main cause of the BCAA deficiency in liver cirrhosis is their consumption in skeletal muscle for synthesis of glutamate, which acts as a substrate for ammonia detoxification to GLN and that the BCAA administration to patients with liver failure may exert a number of positive effects that may be more pronounced in patients with marked depression of BCAA levels. On the other hand, due to the stimulatory effect of BCAA on GLN synthesis, BCAA supplementation may lead to enhanced ammonia production from GLN breakdown in the intestine and the kidneys and thus exert harmful effects on the development of hepatic encephalopathy. Therefore, to enhance therapeutic effectiveness of the BCAA in patients with liver injury, their detrimental effect on ammonia production, which is negligible in healthy people and/or patients with other disorders, should be avoided. In treatment of hepatic encephalopathy, simultaneous administration of the BCAA (to correct amino acid imbalance and promote ammonia detoxification to GLN) with α-ketoglutarate (to inhibit GLN breakdown to ammonia in enterocytes) and/or phenylbutyrate (to enhance GLN excretion by the kidneys) is suggested. Attention should be given to the type of liver injury, gastrointestinal bleeding, signs of inflammation, and the dose of BCAA.

  4. Defects in muscle branched-chain amino acid oxidation contribute to impaired lipid metabolism

    DEFF Research Database (Denmark)

    Lerin, Carles; Goldfine, Allison B; Boes, Tanner;

    2016-01-01

    OBJECTIVE: Plasma levels of branched-chain amino acids (BCAA) are consistently elevated in obesity and type 2 diabetes (T2D) and can also prospectively predict T2D. However, the role of BCAA in the pathogenesis of insulin resistance and T2D remains unclear. METHODS: To identify pathways related t...... catabolism may contribute to the development of insulin resistance by perturbing both amino acid and fatty acid metabolism and suggest that targeting BCAA metabolism may hold promise for prevention or treatment of T2D....... methylmalonyl-CoA mutase (Mut) and assessed the effects of altered BCAA flux on lipid and glucose homeostasis. RESULTS: Our data demonstrate perturbed BCAA metabolism and fatty acid oxidation in muscle from insulin resistant humans. Experimental alterations in BCAA flux in cultured cells similarly modulate...... fatty acid oxidation. Mut heterozygosity in mice alters muscle lipid metabolism in vivo, resulting in increased muscle triglyceride accumulation, increased plasma glucose, hyperinsulinemia, and increased body weight after high-fat feeding. CONCLUSIONS: Our data indicate that impaired muscle BCAA...

  5. Branched-chain amino acids enhance cyst development in autosomal dominant polycystic kidney disease.

    Science.gov (United States)

    Yamamoto, Junya; Nishio, Saori; Hattanda, Fumihiko; Nakazawa, Daigo; Kimura, Toru; Sata, Michio; Makita, Minoru; Ishikawa, Yasunobu; Atsumi, Tatsuya

    2017-03-21

    Autosomal dominant polycystic kidney disease (ADPKD) is characterized by the progressive development of kidney and liver cysts. The mammalian target of rapamycin (mTOR) cascade is one of the important pathways regulating cyst growth in ADPKD. Branched-chain amino acids (BCAAs), including leucine, play a crucial role to activate mTOR pathway. Therefore, we administered BCAA dissolved in the drinking water to Pkd1(flox/flox):Mx1-Cre (cystic) mice from four to 22 weeks of age after polyinosinic-polycytidylic acid-induced conditional Pkd1 knockout at two weeks of age. The BCAA group showed significantly greater kidney/body weight ratio and higher cystic index in both the kidney and liver compared to the placebo-treated mice. We found that the L-type amino acid transporter 1 that facilitates BCAA entry into cells is strongly expressed in cells lining the cysts. We also found increased cyst-lining cell proliferation and upregulation of mTOR and mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) pathways in the BCAA group. In vitro, we cultured renal epithelial cell lines from Pkd1 null mice with or without leucine. Leucine was found to stimulate cell proliferation, as well as activate mTOR and MAPK/ERK pathways in these cells. Thus, BCAA accelerated disease progression by mTOR and MAPK/ERK pathways. Hence, BCAA may be harmful to patients with ADPKD.

  6. Cloning and inactivation of a branched-chain-amino-acid aminotransferase gene from Staphylococcus carnosus and characterization of the enzyme

    DEFF Research Database (Denmark)

    Madsen, Søren M; Beck, Hans Christian; Ravn, Peter

    2002-01-01

    Staphylococcus carnosus and Staphylococcus xylosus are widely used as aroma producers in the manufacture of dried fermented sausages. Catabolism of branched-chain amino acids (BCAAs) by these strains contributes to aroma formation by production of methyl-branched aldehydes and carboxy acids. The ...

  7. Insulin resistance and the metabolism of branched-chain amino acids in humans.

    Science.gov (United States)

    Adeva, María M; Calviño, Jesús; Souto, Gema; Donapetry, Cristóbal

    2012-07-01

    Peripheral resistance to insulin action is the major mechanism causing the metabolic syndrome and eventually type 2 diabetes mellitus. The metabolic derangement associated with insulin resistance is extensive and not restricted to carbohydrates. The branched-chain amino acids (BCAAs) are particularly responsive to the inhibitory insulin action on amino acid release by skeletal muscle and their metabolism is profoundly altered in conditions featuring insulin resistance, insulin deficiency, or both. Obesity, the metabolic syndrome and diabetes mellitus display a gradual increase in the plasma concentration of BCAAs, from the obesity-related low-grade insulin-resistant state to the severe deficiency of insulin action in diabetes ketoacidosis. Obesity-associated hyperinsulinemia succeeds in maintaining near-normal or slightly elevated plasma concentration of BCAAs, despite the insulin-resistant state. The low circulating levels of insulin and/or the deeper insulin resistance occurring in diabetes mellitus are associated with more marked elevation in the plasma concentration of BCAAs. In diabetes ketoacidosis, the increase in plasma BCAAs is striking, returning to normal when adequate metabolic control is achieved. The metabolism of BCAAs is also disturbed in other situations typically featuring insulin resistance, including kidney and liver dysfunction. However, notwithstanding the insulin-resistant state, the plasma level of BCAAs in these conditions is lower than in healthy subjects, suggesting that these organs are involved in maintaining BCAAs blood concentration. The pathogenesis of the decreased BCAAs plasma level in kidney and liver dysfunction is unclear, but a decreased afflux of these amino acids into the blood stream has been observed.

  8. Branched-chain Amino Acid Biosensing Using Fluorescent Modified Engineered Leucine/Isoleucine/Valine Binding Protein

    Directory of Open Access Journals (Sweden)

    Koji Sode

    2007-06-01

    Full Text Available A novel fluorescence sensing system for branched-chain amino acids (BCAAswas developed based on engineered leucine/isoleucine/valine-binding proteins (LIVBPsconjugated with environmentally sensitive fluorescence probes. LIVBP was cloned fromEscherichia coli and Gln149Cys, Gly227Cys, and Gln254Cys mutants were generated bygenetic engineering. The mutant LIVBPs were then modified with environmentallysensitive fluorophores. Based on the fluorescence intensity change observed upon thebinding of the ligands, the MIANS-conjugated Gln149Cys mutant (Gln149Cys-M showedthe highest and most sensitive response. The BCAAs Leu, Ile, and Val can each bemonitored at the sub-micromolar level using Gln149Cys-M. Measurements were alsocarried out on a mixture of BCAFAs and revealed that Gln149Cys-M-based measurementis not significantly affected by the change in the molar ratio of Leu, Ile and Val in thesample. Its high sensitivity and group-specific molecular recognition ability make the newsensing system ideally suited for the measurement of BCAAs and the determination of theFischer ratio, an indicator of hepatic disease involving metabolic dysfunction.

  9. Efficacy, dosage and duration of action of branched chain amino acid therapy for traumatic brain injury

    Directory of Open Access Journals (Sweden)

    Jaclynn eElkind

    2015-03-01

    Full Text Available Traumatic brain injury (TBI results in long-lasting cognitive impairments for which there is currently no accepted treatment. A well-established mouse model of mild to moderate TBI, lateral fluid percussion injury (FPI, shows changes in network excitability in the hippocampus including a decrease in net synaptic efficacy in area CA1 and an increase in net synaptic efficacy in dentate gyrus. Previous studies identified a novel therapy consisting of branched chain amino acids (BCAAs which restored normal mouse hippocampal responses and ameliorated cognitive impairment following FPI. However, the optimal BCAA dose and length of treatment needed to improve cognitive recovery is unknown. In the current study mice underwent FPI then consumed 100 mM BCAA supplemented water ad libitum for 2, 3, 4, 5 and 10 days. BCAA therapy ameliorated cognitive impairment at 5 and 10 days duration. Neither BCAA supplementation at 50 mM, nor BCAAs when dosed 5 days on then 5 days off, was sufficient to ameliorate cognitive impairment. These results suggest that brain injury causes alterations in hippocampal function which underlie and contribute to hippocampal cognitive impairment which are reversible with at least 5 days of BCAA treatment, and that sustaining this effect is dependent on continuous treatment. Our findings have profound implications for the clinical investigation of TBI therapy.

  10. Branched Chain Amino Acids Cause Liver Injury in Obese/Diabetic Mice by Promoting Adipocyte Lipolysis and Inhibiting Hepatic Autophagy

    OpenAIRE

    Fuyang Zhang; Shihao Zhao; Wenjun Yan; Yunlong Xia; Xiyao Chen; Wei Wang; Jinglong Zhang; Chao Gao; Cheng Peng; Feng Yan; Huishou Zhao; Kun Lian; Yan Lee; Ling Zhang; Wayne Bond Lau

    2016-01-01

    The Western meat-rich diet is both high in protein and fat. Although the hazardous effect of a high fat diet (HFD) upon liver structure and function is well recognized, whether the co-presence of high protein intake contributes to, or protects against, HF-induced hepatic injury remains unclear. Increased intake of branched chain amino acids (BCAA, essential amino acids compromising 20% of total protein intake) reduces body weight. However, elevated circulating BCAA is associated with non-alco...

  11. Bolus ingestion of individual branched-chain amino acids alters plasma amino acid profiles in young healthy men.

    Science.gov (United States)

    Matsumoto, Takuya; Nakamura, Koichi; Matsumoto, Hideki; Sakai, Ryosei; Kuwahara, Tomomi; Kadota, Yoshihiro; Kitaura, Yasuyuki; Sato, Juichi; Shimomura, Yoshiharu

    2014-01-01

    Physiological conditions in humans affect plasma amino acid profiles that might have potential for medical use. Because the branched-chain amino acids (BCAAs) leucine, isoleucine and valine are used as medicines and supplements, we investigated the acute effects of individual BCAAs (10-90 mg/kg body weight) or mixed BCAAs ingested as a bolus on plasma amino acid profiles in young healthy men. Plasma leucine levels rapidly increased and peaked around 30 min after leucine ingestion. Concentrations of plasma isoleucine, valine and phenylalanine subsequently decreased after ingestion, and those of methionine and tyrosine tended to decrease. The effects of ingested leucine on other plasma amino acids were biphasic, being higher at lower doses (10-20 mg/kg body weight). Isoleucine or valine intake also caused corresponding plasma amino acid concentrations to rapidly elevate, and peaks at 30-40 min after ingestion were much higher than that of plasma leucine after leucine ingestion. However, the increase in plasma isoleucine and valine concentrations essentially did not affect those of other plasma amino acids. The rate of decline among peak plasma BCAA concentrations was the highest for leucine, followed by isoleucine and valine. Oral mixed BCAAs promoted the decline in plasma isoleucine and valine concentrations. These results suggest that plasma leucine is a regulator of the plasma concentrations of BCAAs, methionine and aromatic amino acids.

  12. Adipose tissue branched chain amino acid (BCAA) metabolism modulates circulating BCAA levels.

    Science.gov (United States)

    Herman, Mark A; She, Pengxiang; Peroni, Odile D; Lynch, Christopher J; Kahn, Barbara B

    2010-04-09

    Whereas the role of adipose tissue in glucose and lipid homeostasis is widely recognized, its role in systemic protein and amino acid metabolism is less well-appreciated. In vitro and ex vivo experiments suggest that adipose tissue can metabolize substantial amounts of branched chain amino acids (BCAAs). However, the role of adipose tissue in regulating BCAA metabolism in vivo is controversial. Interest in the contribution of adipose tissue to BCAA metabolism has been renewed with recent observations demonstrating down-regulation of BCAA oxidation enzymes in adipose tissue in obese and insulin-resistant humans. Using gene set enrichment analysis, we observe alterations in adipose-tissue BCAA enzyme expression caused by adipose-selective genetic alterations in the GLUT4 glucose-transporter expression. We show that the rate of adipose tissue BCAA oxidation per mg of tissue from normal mice is higher than in skeletal muscle. In mice overexpressing GLUT4 specifically in adipose tissue, we observe coordinate down-regulation of BCAA metabolizing enzymes selectively in adipose tissue. This decreases BCAA oxidation rates in adipose tissue, but not in muscle, in association with increased circulating BCAA levels. To confirm the capacity of adipose tissue to modulate circulating BCAA levels in vivo, we demonstrate that transplantation of normal adipose tissue into mice that are globally defective in peripheral BCAA metabolism reduces circulating BCAA levels by 30% (fasting)-50% (fed state). These results demonstrate for the first time the capacity of adipose tissue to catabolize circulating BCAAs in vivo and that coordinate regulation of adipose-tissue BCAA enzymes may modulate circulating BCAA levels.

  13. Regulation of indole-3-acetic acid biosynthesis by branched-chain amino acids in Enterobacter cloacae UW5.

    Science.gov (United States)

    Parsons, Cassandra V; Harris, Danielle M M; Patten, Cheryl L

    2015-09-01

    The soil bacterium Enterobacter cloacae UW5 produces the rhizosphere signaling molecule indole-3-acetic acid (IAA) via the indolepyruvate pathway. Expression of indolepyruvate decarboxylase, a key pathway enzyme encoded by ipdC, is upregulated by the transcription factor TyrR in response to aromatic amino acids. Some members of the TyrR regulon may also be controlled by branched-chain amino acids and here we show that expression from the ipdC promoter and production of IAA are downregulated by valine, leucine and isoleucine. Regulation of the IAA synthesis pathway by both aromatic and branched-chain amino acids suggests a broader role for this pathway in bacterial physiology, beyond plant interactions.

  14. Plasma branched-chain amino acids and incident cardiovascular disease in the PREDIMED trial

    Science.gov (United States)

    Ruiz-Canela, Miguel; Toledo, Estefania; Clish, Clary B.; Hruby, Adela; Liang, Liming; Salas-Salvadó, Jordi; Razquin, Cristina; Corella, Dolores; Estruch, Ramón; Ros, Emilio; Fitó, Montserrat; Gómez-Gracia, Enrique; Arós, Fernando; Fiol, Miquel; Lapetra, José; Serra-Majem, Lluis; Martínez-González, Miguel A.; Hu, Frank B.

    2016-01-01

    Background The role of branched-chain amino acids (BCAAs) in cardiovascular disease (CVD) remains poorly understood. We hypothesized that baseline BCAA concentrations predict future risk of CVD and that a Mediterranean Diet (MedDiet) intervention may counteract this effect. Methods We developed a case-cohort study within the “PREvención con DIeta MEDiterránea” (PREDIMED), with 226 incident CVD cases and 781 non-cases. We used LC-MS/MS to measure plasma BCAAs (leucine, isoleucine and valine), both at baseline and after 1-year follow-up. The primary outcome was a composite of incident stroke, myocardial infarction, or cardiovascular death. Results After adjustment for potential confounders, baseline leucine and isoleucine concentrations were associated with higher CVD risk: the hazard ratios (HRs) for the highest vs. lowest quartile were 1.70 (95% confidence interval, 1.05–2.76) and 2.09 (1.27–3.44), respectively. Stronger associations were found for stroke. For both CVD and stroke, we found higher HRs across successive quartiles of BCAAs in the control group than in the MedDiet groups. Using stroke as the outcome, a significant interaction (P=0.009) between the baseline BCAA score and the intervention with MedDiet was observed. No significant effect of the intervention on 1-yr changes in BCAAs nor any association between 1-year changes in BCAAs and CVD were observed. Conclusions Higher concentrations of baseline BCAAs were associated with increased risk of CVD, especially stroke, in a high cardiovascular risk population. A Mediterranean-style diet had a negligible effect on 1-year changes in BCAAs, but it may counteract the harmful effects of BCAAs on stroke. PMID:26888892

  15. Inhibitory effect of aroma on the bitterness of branched-chain amino acid solutions.

    Science.gov (United States)

    Mukai, Junji; Tokuyama, Emi; Ishizaka, Toshihiko; Okada, Sachie; Uchida, Takahiro

    2007-11-01

    Nutritional products for patients with liver failure available on the Japanese market contain many branched-chain amino acids (BCAAs) such as L-leucine, L-isoleucine, and L-valine, which not only have a bitter taste but also strong, unpleasant odours, leading to low palatability. The palatability of these nutritional products can be significantly improved by the addition of flavoured powders containing various kinds of tastants (sucrose, citric acid, etc.) and odourants (fruit, coffee aromas, etc.). The specific effects of the aroma of flavoured powders have not yet been clearly evaluated. In the present article, the inhibitory effect of aroma on the bitterness of BCAA solutions was examined. The bitterness intensity of a BCAA solution at the same concentration as Aminoleban EN was defined as 3.5 (measured by a previously described gustatory sensation method). The bitterness threshold of a BCAA standard solution without added aroma was estimated to be 1.87, while those of BCAA solutions containing green-tea, coffee, apple, vanilla, or strawberry aromas were 2.02, 1.98, 2.35, 2.40 and 2.87, respectively, when evaluated by the probit method. This shows that the addition of an aroma can elevate the bitterness threshold in human volunteers. The green-tea and coffee aromas predominantly evoked bitterness, while the vanilla aroma predominantly evoked sweetness. Apple and strawberry aromas evoked both sweetness and sourness, with the apple aroma having stronger sourness and the strawberry aroma stronger sweetness. Thus, a 'sweet' aroma suppresses the bitterness of BCAA, with coexisting sourness also participating in the bitterness inhibition.

  16. Comparative Genomics of Regulation of Fatty Acid and Branched-chain Amino Acid Utilization in Proteobacteria

    Energy Technology Data Exchange (ETDEWEB)

    Kazakov, Alexey E.; Rodionov, Dmitry A.; Arkin, Adam Paul; Dubchak, Inna; Gelfand, Mikhail S.; Alm, Eric

    2008-10-31

    Bacteria can use branched-chain amino acids (ILV, i.e. isoleucine, leucine, valine) and fatty acids (FA) as sole carbon and energy sources convering ILV into acetyl-CoA, propanoyl-CoA and propionyl-CoA, respectively. In this work, we used the comparative genomic approach to identify candidate transcriptional factors and DNA motifs that control ILV and FA utilization pathways in proteobacteria. The metabolic regulons were characterized based on the identification and comparison of candidate transcription factor binding sites in groups of phylogenetically related genomes. The reconstructed ILV/FA regulatory network demonstrates considerable variability and involves six transcriptional factors from the MerR, TetR and GntR families binding to eleven distinct DNA motifs. The ILV degradation genes in gamma- and beta-proteobacteria are mainly regulated by anovel regulator from the MerR family (e.g., LiuR in Pseudomonas aeruginosa) (40 species), in addition, the TetR-type regulator LiuQ was identified in some beta-proteobacteria (8 species). Besides the core set of ILV utilization genes, the LiuR regulon in some lineages is expanded to include genes from other metabolic pathways, such as the glyoxylate shunt and glutamate synthase in the Shewanella species. The FA degradation genes are controlled by four regulators including FadR in gamma-proteobacteria (34 species), PsrA in gamma- and beta-proteobacteria (45 species), FadP in beta-proteobacteria (14 species), and LiuR orthologs in alpha-proteobacteria (22 species). The remarkable variability of the regulatory systems associated with the FA degradation pathway is discussed from the functional and evolutionary points of view.

  17. Cross-talk between branched-chain amino acids and hepatic mitochondria is compromised in nonalcoholic fatty liver disease

    OpenAIRE

    Sunny, Nishanth E.; Kalavalapalli, Srilaxmi; Bril, Fernando; Garrett, Timothy J; Nautiyal, Manisha; Mathew, Justin T.; Caroline M Williams; Cusi, Kenneth

    2015-01-01

    Elevated plasma branched-chain amino acids (BCAA) in the setting of insulin resistance have been relevant in predicting type 2 diabetes mellitus (T2DM) onset, but their role in the etiology of hepatic insulin resistance remains uncertain. We determined the link between BCAA and dysfunctional hepatic tricarboxylic acid (TCA) cycle, which is a central feature of hepatic insulin resistance and nonalcoholic fatty liver disease (NAFLD). Plasma metabolites under basal fasting and euglycemic hyperin...

  18. Is administrating branched-chain amino acid-enriched nutrition achieved symptom-free in malnourished cirrhotic patients?

    OpenAIRE

    Tsuda, Yasuhiro; Fukui, Hideo; Sujishi, Tetsuya; Ohama, Hideko; Tsuchimoto, Yusuke; Asai, Akira; Fukunisi, Shinya; Higuchi, Kazuhide

    2013-01-01

    Administration of branched-chain amino acids (BCAA) has been reported to improve liver function, quality of life (QOL). However, in some malnourished patients, serum albumin levels do not improve in response to BCAA granules. In this study, we examined the effects of BCAA-enriched enteral nutrition in patients unresponsive to BCAA granules. Thirty-two decompensated cirrhotic patients at Osaka Medical College were enrolled in this study. Since all patients showed no improvement in serum albumi...

  19. Branched-Chain Amino Acids Are Required for the Survival and Virulence of Actinobacillus pleuropneumoniae in Swine▿

    OpenAIRE

    Subashchandrabose, Sargurunathan; LeVeque, Rhiannon M.; Wagner, Trevor K.; Kirkwood, Roy N; Kiupel, Matti; Mulks, Martha H.

    2009-01-01

    In Actinobacillus pleuropneumoniae, which causes porcine pleuropneumonia, ilvI was identified as an in vivo-induced (ivi) gene and encodes the enzyme acetohydroxyacid synthase (AHAS) required for branched-chain amino acid (BCAA) biosynthesis. ilvI and 7 of 32 additional ivi promoters were upregulated in vitro when grown in chemically defined medium (CDM) lacking BCAA. Based on these observations, we hypothesized that BCAA would be found at limiting concentrations in pulmonary secretions and t...

  20. Branched-Chain Amino Acid Ingestion Stimulates Muscle Myofibrillar Protein Synthesis following Resistance Exercise in Humans

    Directory of Open Access Journals (Sweden)

    Sarah R. Jackman

    2017-06-01

    Full Text Available The ingestion of intact protein or essential amino acids (EAA stimulates mechanistic target of rapamycin complex-1 (mTORC1 signaling and muscle protein synthesis (MPS following resistance exercise. The purpose of this study was to investigate the response of myofibrillar-MPS to ingestion of branched-chain amino acids (BCAAs only (i.e., without concurrent ingestion of other EAA, intact protein, or other macronutrients following resistance exercise in humans. Ten young (20.1 ± 1.3 years, resistance-trained men completed two trials, ingesting either 5.6 g BCAA or a placebo (PLA drink immediately after resistance exercise. Myofibrillar-MPS was measured during exercise recovery with a primed, constant infusion of L-[ring13C6] phenylalanine and collection of muscle biopsies pre and 4 h-post drink ingestion. Blood samples were collected at time-points before and after drink ingestion. Western blotting was used to measure the phosphorylation status of mTORC1 signaling proteins in biopsies collected pre, 1-, and 4 h-post drink. The percentage increase from baseline in plasma leucine (300 ± 96%, isoleucine (300 ± 88%, and valine (144 ± 59% concentrations peaked 0.5 h-post drink in BCAA. A greater phosphorylation status of S6K1Thr389 (P = 0.017 and PRAS40 (P = 0.037 was observed in BCAA than PLA at 1 h-post drink ingestion. Myofibrillar-MPS was 22% higher (P = 0.012 in BCAA (0.110 ± 0.009%/h than PLA (0.090 ± 0.006%/h. Phenylalanine Ra was ~6% lower in BCAA (18.00 ± 4.31 μmol·kgBM−1 than PLA (21.75 ± 4.89 μmol·kgBM−1; P = 0.028 after drink ingestion. We conclude that ingesting BCAAs alone increases the post-exercise stimulation of myofibrillar-MPS and phosphorylation status mTORC1 signaling.

  1. Associations between plasma branched-chain amino acids, β-aminoisobutyric acid and body composition.

    Science.gov (United States)

    Rietman, Annemarie; Stanley, Takara L; Clish, Clary; Mootha, Vamsi; Mensink, Marco; Grinspoon, Steven K; Makimura, Hideo

    2016-01-01

    Plasma branched-chain amino acids (BCAA) are elevated in obesity and associated with increased cardiometabolic risk. β-Aminoisobutyric acid (B-AIBA), a recently identified small molecule metabolite, is associated with decreased cardiometabolic risk. Therefore, we investigated the association of BCAA and B-AIBA with each other and with detailed body composition parameters, including abdominal visceral adipose tissue (VAT) and subcutaneous adipose tissue (SAT). A cross-sectional study was carried out with lean (n 15) and obese (n 33) men and women. Detailed metabolic evaluations, including measures of body composition, insulin sensitivity and plasma metabolomics were completed. Plasma BCAA were higher (1·6 (se 0·08) (×10(7)) v. 1·3 (se 0·06) (×10(7)) arbitrary units; P = 0·005) in obese v. lean subjects. BCAA were positively associated with VAT (R 0·49; P = 0·0006) and trended to an association with SAT (R 0·29; P = 0·052). The association between BCAA and VAT, but not SAT, remained significant after controlling for age, sex and race on multivariate modelling (P BCAA were also associated with parameters of insulin sensitivity (Matsuda index: R -0·50, P = 0·0004; glucose AUC: R 0·53, P BCAA were not associated with B-AIBA (R -0·04; P = 0·79). B-AIBA was negatively associated with SAT (R -0·37; P = 0·01) but only trended to an association with VAT (R 0·27; P = 0·07). However, neither relationship remained significant after multivariate modelling (P > 0·05). Plasma B-AIBA was associated with parameters of insulin sensitivity (Matsuda index R 0·36, P = 0·01; glucose AUC: R -0·30, P = 0·04). Plasma BCAA levels were positively correlated with VAT and markers of insulin resistance. The results suggest a possible complex role of adipose tissue in BCAA homeostasis and insulin resistance.

  2. Branched-chain amino acid treatment before transcatheter arterial chemoembolization for hepatocellular carcinoma

    Institute of Scientific and Technical Information of China (English)

    Hiroki Nishikawa; Yukio Osaki; Tadashi Inuzuka; Haruhiko Takeda; Jun Nakajima; Fumihiro Matsuda; Shin-ichiro Henmi

    2012-01-01

    AIM:To examine the significance of branched-chain amino acid (BCAA) treatment before transcatheter arterial chemoembolization (TACE) for hepatocellular carcinoma (HCC).METHODS:This study included 99 patients who underwent TACE therapy for HCC at our hospital and were followed up without treatment for at least 6 mo between January 2004 and January 2010.They were divided into 2 groups:those receiving BCAA granules (n =40) or regular diet (n =59,control).Data obtained were retrospectively analyzed (prior to TACE,and 1 wk,1,3,and 6 mo after TACE) in terms of nutritional condition and clinical laboratory parameters (serum albumin level and Child-Pugh score),both of which are determinants of hepatic functional reserve.RESULTS:The BCAA group comprised 27 males and 13 females with a mean age of 69.9 ± 8.8 years.The patients of the BCAA group were classified as follows:Child-Pugh A/B/C in 22/15/3 patients,and Stage Ⅱ/Ⅲ/IVA HCC in 12/23/5 patients,respectively.The control group comprised 32 males and 27 females with a mean age of 73.2 ± 10.1 years.In the control group,9 patients had chronic hepatitis,Child-Pugh A/B/C in 39/10/1 patients,and Stage I/Ⅱ /Ⅲ/IVA HCC in 1/11/35/12 patients,respectively.Overall,both serum albumin level and Child-Pugh score improved significantly in the BCAA group as compared with the control 3 and 6 mo after TACE (P < 0.05).Further analysis was performed by the following categorization:(1) child-Pugh classification; (2) liver cirrhosis subgroup with a serum albumin level > 3.5 g/dL; and (3) epirubicin dose.A similar trend indicating a significant improvement of all variables in the BCAA group was noted (P < 0.05).CONCLUSION:Treatment with BCAA granules in patients who have undergone TACE for HCC is considered useful to maintain their hepatic functional reserve.

  3. Long-term treatment of latent portosystemic encephalopathy with branched-chain amino acids. A double-blind placebo-controlled crossover study.

    Science.gov (United States)

    Plauth, M; Egberts, E H; Hamster, W; Török, M; Müller, P H; Brand, O; Fürst, P; Dölle, W

    1993-03-01

    This trial was undertaken to assess the safety and efficacy of long-term oral supplementation with branched-chain amino acids as an adjunct to conventional therapy in patients with stable cirrhosis and latent encephalopathy. Latent encephalopathy was diagnosed by psychometric testing, used to assess automobile driving capacity. Seventeen patients with impaired driving capacity received either branched-chain amino acids or placebo for 8 weeks before being crossed over to the other regimen for an equal period. Branched-chain amino acids but not placebo significantly improved psychomotor disturbances (p automobile driving capacity associated with latent portosystemic encephalopathy.

  4. The Effects of Herbicides Targeting Aromatic and Branched Chain Amino Acid Biosynthesis Support the Presence of Functional Pathways in Broomrape.

    Science.gov (United States)

    Dor, Evgenia; Galili, Shmuel; Smirnov, Evgeny; Hacham, Yael; Amir, Rachel; Hershenhorn, Joseph

    2017-01-01

    It is not clear why herbicides targeting aromatic and branched-chain amino acid biosynthesis successfully control broomrapes-obligate parasitic plants that obtain all of their nutritional requirements, including amino acids, from the host. Our objective was to reveal the mode of action of imazapic and glyphosate in controlling the broomrape Phelipanche aegyptiaca and clarify if this obligatory parasite has its own machinery for the amino acids biosynthesis. P. aegyptiaca callus was studied to exclude the indirect influence of the herbicides on the parasite through the host plant. Using HRT - tomato plants resistant to imidazolinone herbicides, it was shown that imazapic is translocated from the foliage of treated plants to broomrape attachments on its roots and controls the parasite. Both herbicides inhibited P. aegyptiaca callus growth and altered the free amino acid content. Blasting of Arabidopsis thaliana 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) and acetolactate synthase (ALS) cDNA against the genomic DNA of P. aegyptiaca yielded a single copy of each homolog in the latter, with about 78 and 75% similarity, respectively, to A. thaliana counterparts at the protein level. We also show for the first time that both EPSPS and ALS are active in P. aegyptiaca callus and flowering shoots and are inhibited by glyphosate and imazapic, respectively. Thus leading to deficiency of those amino acids in the parasite tissues and ultimately, death of the parasite, indicating the ability of P. aegyptiaca to synthesize branched-chain and aromatic amino acids through the activity of ALS and EPSPS, respectively.

  5. The branched-chain amino acid aminotransferase encoded by ilvE is involved in acid tolerance in Streptococcus mutans.

    Science.gov (United States)

    Santiago, Brendaliz; MacGilvray, Matthew; Faustoferri, Roberta C; Quivey, Robert G

    2012-04-01

    The ability of Streptococcus mutans to produce and tolerate organic acids from carbohydrate metabolism represents a major virulence factor responsible for the formation of carious lesions. Pyruvate is a key metabolic intermediate that, when rerouted to other metabolic pathways such as amino acid biosynthesis, results in the alleviation of acid stress by reducing acid end products and aiding in maintenance of intracellular pH. Amino acid biosynthetic genes such as ilvC and ilvE were identified as being upregulated in a proteome analysis of Streptococcus mutans under acid stress conditions (A. C. Len, D. W. Harty, and N. A. Jacques, Microbiology 150:1353-1366, 2004). In Lactococcus lactis and Staphylococcus carnosus, the ilvE gene product is involved with biosynthesis and degradation of branched-chain amino acids, as well as in the production of branched-chain fatty acids (B. Ganesan and B. C. Weimer, Appl. Environ. Microbiol. 70:638-641, 2004; S. M. Madsen et al., Appl. Environ. Microbiol. 68:4007-4014, 2002; and M. Yvon, S. Thirouin, L. Rijnen, D. Fromentier, and J. C. Gripon, Appl. Environ. Microbiol. 63:414-419, 1997). Here we constructed and characterized an ilvE deletion mutant of S. mutans UA159. Growth experiments revealed that the ilvE mutant strain has a lag in growth when nutritionally limited for branched-chain amino acids. We further demonstrated that the loss of ilvE causes a decrease in acid tolerance. The ilvE strain exhibits a defect in F(1)-F(o) ATPase activity and has reduced catabolic activity for isoleucine and valine. Results from transcriptional studies showed that the ilvE promoter is upregulated during growth at low pH. Collectively, the results of this investigation show that amino acid metabolism is a component of the acid-adaptive repertoire of S. mutans.

  6. Exercise-induced muscle damage is reduced in resistance-trained males by branched chain amino acids: a randomized, double-blind, placebo controlled study

    National Research Council Canada - National Science Library

    Howatson, Glyn; Hoad, Michael; Goodall, Stuart; Tallent, Jamie; Bell, Phillip G; French, Duncan N

    2012-01-01

    .... Branched-chain amino acid (BCAA) supplementation has been shown to increase protein synthesis and decrease muscle protein breakdown, however, the effects of BCAAs on recovery from damaging resistance training are unclear...

  7. Genetic evidence of a causal effect of insulin resistance on branched-chain amino acid levels.

    Science.gov (United States)

    Mahendran, Yuvaraj; Jonsson, Anna; Have, Christian T; Allin, Kristine H; Witte, Daniel R; Jørgensen, Marit E; Grarup, Niels; Pedersen, Oluf; Kilpeläinen, Tuomas O; Hansen, Torben

    2017-05-01

    Fasting plasma levels of branched-chain amino acids (BCAAs) are associated with insulin resistance, but it remains unclear whether there is a causal relation between the two. We aimed to disentangle the causal relations by performing a Mendelian randomisation study using genetic variants associated with circulating BCAA levels and insulin resistance as instrumental variables. We measured circulating BCAA levels in blood plasma by NMR spectroscopy in 1,321 individuals from the ADDITION-PRO cohort. We complemented our analyses by using previously published genome-wide association study (GWAS) results from the Meta-Analyses of Glucose and Insulin-related traits Consortium (MAGIC) (n = 46,186) and from a GWAS of serum BCAA levels (n = 24,925). We used a genetic risk score (GRS), calculated using ten established fasting serum insulin associated variants, as an instrumental variable for insulin resistance. A GRS of three variants increasing circulating BCAA levels was used as an instrumental variable for circulating BCAA levels. Fasting plasma BCAA levels were associated with higher HOMA-IR in ADDITION-PRO (β 0.137 [95% CI 0.08, 0.19] p = 6 × 10(-7)). However, the GRS for circulating BCAA levels was not associated with fasting insulin levels or HOMA-IR in ADDITION-PRO (β -0.011 [95% CI -0.053, 0.032] p = 0.6 and β -0.011 [95% CI -0.054, 0.031] p = 0.6, respectively) or in GWAS results for HOMA-IR from MAGIC (β for valine-increasing GRS -0.012 [95% CI -0.069, 0.045] p = 0.7). By contrast, the insulin-resistance-increasing GRS was significantly associated with increased BCAA levels in ADDITION-PRO (β 0.027 [95% CI 0.005, 0.048] p = 0.01) and in GWAS results for serum BCAA levels (β 1.22 [95% CI 0.71, 1.73] p = 4 × 10(-6), β 0.96 [95% CI 0.45, 1.47] p = 3 × 10(-4), and β 0.67 [95% CI 0.16, 1.18] p = 0.01 for isoleucine, leucine and valine levels, respectively) and instrumental variable analyses in ADDITION

  8. YjeH Is a Novel Exporter of l-Methionine and Branched-Chain Amino Acids in Escherichia coli.

    Science.gov (United States)

    Liu, Qian; Liang, Yong; Zhang, Yun; Shang, Xiuling; Liu, Shuwen; Wen, Jifu; Wen, Tingyi

    2015-11-01

    Amino acid efflux transport systems have important physiological functions and play vital roles in the fermentative production of amino acids. However, no methionine exporter has yet been identified in Escherichia coli. In this study, we identified a novel amino acid exporter, YjeH, in E. coli. The yjeH overexpression strain exhibited high tolerance to the structural analogues of l-methionine and branched-chain amino acids, decreased intracellular amino acid levels, and enhanced export rates in the presence of a Met-Met, Leu-Leu, Ile-Ile, or Val-Val dipeptide, suggesting that YjeH functions as an exporter of l-methionine and the three branched-chain amino acids. The export of the four amino acids in the yjeH overexpression strain was competitively inhibited in relation to each other. The expression of yjeH was strongly induced by increasing cytoplasmic concentrations of substrate amino acids. Green fluorescent protein (GFP)-tagged YjeH was visualized by total internal reflection fluorescence microscopy to confirm the plasma membrane localization of YjeH. Phylogenetic analysis of transporters indicated that YjeH belongs to the amino acid efflux family of the amino acid/polyamine/organocation (APC) superfamily. Structural modeling revealed that YjeH has the typical "5 + 5" transmembrane α-helical segment (TMS) inverted-repeat fold of APC superfamily transporters, and its binding sites are strictly conserved. The enhanced capacity of l-methionine export by the overexpression of yjeH in an l-methionine-producing strain resulted in a 70% improvement in titer. This study supplements the transporter classification and provides a substantial basis for the application of the methionine exporter in metabolic engineering.

  9. The antifungal eugenol perturbs dual aromatic and branched-chain amino acid permeases in the cytoplasmic membrane of yeast.

    Science.gov (United States)

    Darvishi, Emad; Omidi, Mansoor; Bushehri, Ali Akbar Shahnejat; Golshani, Ashkan; Smith, Myron L

    2013-01-01

    Eugenol is an aromatic component of clove oil that has therapeutic potential as an antifungal drug, although its mode of action and precise cellular target(s) remain ambiguous. To address this knowledge gap, a chemical-genetic profile analysis of eugenol was done using ∼4700 haploid Saccharomyces cerevisiae gene deletion mutants to reveal 21 deletion mutants with the greatest degree of susceptibility. Cellular roles of deleted genes in the most susceptible mutants indicate that the main targets for eugenol include pathways involved in biosynthesis and transport of aromatic and branched-chain amino acids. Follow-up analyses showed inhibitory effects of eugenol on amino acid permeases in the yeast cytoplasmic membrane. Furthermore, phenotypic suppression analysis revealed that eugenol interferes with two permeases, Tat1p and Gap1p, which are both involved in dual transport of aromatic and branched-chain amino acids through the yeast cytoplasmic membrane. Perturbation of cytoplasmic permeases represents a novel antifungal target and may explain previous observations that exposure to eugenol results in leakage of cell contents. Eugenol exposure may also contribute to amino acid starvation and thus holds promise as an anticancer therapeutic drug. Finally, this study provides further evidence of the usefulness of the yeast Gene Deletion Array approach in uncovering the mode of action of natural health products.

  10. Repeated sprint ability is not enhanced by caffeine, arginine, and branched-chain amino acids in moderately trained soccer players.

    Science.gov (United States)

    Ermolao, Andrea; Zanotto, Tobia; Carraro, Nicolò; Fornasier, Tommaso; Zaccaria, Marco; Neunhaeuserer, Daniel; Bergamin, Marco

    2017-02-01

    The aim was to investigate the effect of a dietary supplementation on the repeated sprint ability (RSA) performance in recreationally trained team sports athletes. Twelve young men underwent a RSA exercise protocol in five trials, in which participants ingested carbohydrates (CHO) plus caffeine (Caf), CHO plus arginine (Arg), CHO plus branched-chain amino acids (BCAA), CHO plus Caf, Arg, and BCAA (ALL), and CHO only. Heart rate, oxygen saturation, hematic lactate, ratings of perceived exertion, average sprint time, total time, best sprint time, peak power, and average power were taken. Data revealed no significant effects neither on physiological nor performance parameters with any of the supplements.

  11. Volatile profiling reveals intracellular metabolic changes in Aspergillus parasiticus: veA regulates branched chain amino acid and ethanol metabolism

    Directory of Open Access Journals (Sweden)

    Roze Ludmila V

    2010-08-01

    Full Text Available Abstract Background Filamentous fungi in the genus Aspergillus produce a variety of natural products, including aflatoxin, the most potent naturally occurring carcinogen known. Aflatoxin biosynthesis, one of the most highly characterized secondary metabolic pathways, offers a model system to study secondary metabolism in eukaryotes. To control or customize biosynthesis of natural products we must understand how secondary metabolism integrates into the overall cellular metabolic network. By applying a metabolomics approach we analyzed volatile compounds synthesized by Aspergillus parasiticus in an attempt to define the association of secondary metabolism with other metabolic and cellular processes. Results Volatile compounds were examined using solid phase microextraction - gas chromatography/mass spectrometry. In the wild type strain Aspergillus parasiticus SU-1, the largest group of volatiles included compounds derived from catabolism of branched chain amino acids (leucine, isoleucine, and valine; we also identified alcohols, esters, aldehydes, and lipid-derived volatiles. The number and quantity of the volatiles produced depended on media composition, time of incubation, and light-dark status. A block in aflatoxin biosynthesis or disruption of the global regulator veA affected the volatile profile. In addition to its multiple functions in secondary metabolism and development, VeA negatively regulated catabolism of branched chain amino acids and synthesis of ethanol at the transcriptional level thus playing a role in controlling carbon flow within the cell. Finally, we demonstrated that volatiles generated by a veA disruption mutant are part of the complex regulatory machinery that mediates the effects of VeA on asexual conidiation and sclerotia formation. Conclusions 1 Volatile profiling provides a rapid, effective, and powerful approach to identify changes in intracellular metabolic networks in filamentous fungi. 2 VeA coordinates the

  12. Repeated sprint ability is not enhanced by caffeine, arginine, and branched-chain amino acids in moderately trained soccer players

    Science.gov (United States)

    Ermolao, Andrea; Zanotto, Tobia; Carraro, Nicolò; Fornasier, Tommaso; Zaccaria, Marco; Neunhaeuserer, Daniel; Bergamin, Marco

    2017-01-01

    The aim was to investigate the effect of a dietary supplementation on the repeated sprint ability (RSA) performance in recreationally trained team sports athletes. Twelve young men underwent a RSA exercise protocol in five trials, in which participants ingested carbohydrates (CHO) plus caffeine (Caf), CHO plus arginine (Arg), CHO plus branched-chain amino acids (BCAA), CHO plus Caf, Arg, and BCAA (ALL), and CHO only. Heart rate, oxygen saturation, hematic lactate, ratings of perceived exertion, average sprint time, total time, best sprint time, peak power, and average power were taken. Data revealed no significant effects neither on physiological nor performance parameters with any of the supplements. PMID:28349034

  13. Repletion of branched chain amino acids reverses mTORC1 signaling but not improved metabolism during dietary protein dilution

    DEFF Research Database (Denmark)

    Maida, Adriano; Chan, Jessica S K; Sjøberg, Kim Anker

    2017-01-01

    dietary dilution of specific amino acids (AAs) is also required. Circulating branched chain amino acids (BCAAs) are sensitive to protein intake, elevated in the serum of obese humans and mice and thought to promote insulin resistance. We tested whether replenishment of dietary BCAAs to an AA-diluted (AAD......) diet is sufficient to reverse the glucoregulatory benefits of dietary PD. METHODS: We conducted AA profiling of serum from healthy humans and lean and high fat-fed or New Zealand obese (NZO) mice following dietary PD. We fed wildtype and NZO mice one of three amino acid defined diets: control, total......, while elevated in serum from hyperphagic NZO, were consistently reduced by dietary PD in humans and murine models. Repletion of dietary BCAAs modestly attenuated insulin sensitivity and metabolic efficiency in wildtype mice but did not restore hyperglycemia in NZO mice. While hepatic markers of the ISR...

  14. The Effects of Herbicides Targeting Aromatic and Branched Chain Amino Acid Biosynthesis Support the Presence of Functional Pathways in Broomrape

    Directory of Open Access Journals (Sweden)

    Evgenia Dor

    2017-05-01

    Full Text Available It is not clear why herbicides targeting aromatic and branched-chain amino acid biosynthesis successfully control broomrapes—obligate parasitic plants that obtain all of their nutritional requirements, including amino acids, from the host. Our objective was to reveal the mode of action of imazapic and glyphosate in controlling the broomrape Phelipanche aegyptiaca and clarify if this obligatory parasite has its own machinery for the amino acids biosynthesis. P. aegyptiaca callus was studied to exclude the indirect influence of the herbicides on the parasite through the host plant. Using HRT – tomato plants resistant to imidazolinone herbicides, it was shown that imazapic is translocated from the foliage of treated plants to broomrape attachments on its roots and controls the parasite. Both herbicides inhibited P. aegyptiaca callus growth and altered the free amino acid content. Blasting of Arabidopsis thaliana 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS and acetolactate synthase (ALS cDNA against the genomic DNA of P. aegyptiaca yielded a single copy of each homolog in the latter, with about 78 and 75% similarity, respectively, to A. thaliana counterparts at the protein level. We also show for the first time that both EPSPS and ALS are active in P. aegyptiaca callus and flowering shoots and are inhibited by glyphosate and imazapic, respectively. Thus leading to deficiency of those amino acids in the parasite tissues and ultimately, death of the parasite, indicating the ability of P. aegyptiaca to synthesize branched-chain and aromatic amino acids through the activity of ALS and EPSPS, respectively.

  15. Involvement of branched-chain amino acid aminotransferases in the production of fusel alcohols during fermentation in yeast

    Energy Technology Data Exchange (ETDEWEB)

    Eden, A.; Drukker, M.; Benvenisty, N. [Hebrew Univ., Jerusalem (Israel). Dept. of Genetics; Nedervelde, L. van; Debourg, A. [Haute Ecole Lucia de Brouckere, Brussels (Belgium). Dept. of Brewing Sciences and Fermentation Technology

    2001-07-01

    Organoleptic compounds produced by yeast during the fermentation of wort have a great impact on beer smell and taste. Among them, fusel alcohols are the major abundant volatile compounds. The availability of Saccharomyces cerevisiae mutants in which the genes coding for the two branched-chain amino acid aminotransferases have been deleted offers the possibility of further defining the role of these enzymes in the formation of higher alcohols. Comparing the production profiles of different strains, it is clear that they are not all influenced in the same way by branched-chain amino acid aminostransferase mutations. First of all, as propanol is synthesised from {alpha}-ketobutyrate, the first metabolic intermediate in the anabolic pathway of isoleucine, neither the eca39 nor eca40 mutations have any effect on the production of this higher alcohol. On the other hand, it can be concluded that the eca40 mutation has a drastic effect on the production of isobutanol. To a certain extent, the same conclusion can be made for the production of active amyl alcohol and isoamyl alcohol, although the results suggest that another route could lead to the formation of these two higher alcohols. (orig.)

  16. First structure of archaeal branched-chain amino acid aminotransferase from Thermoproteus uzoniensis specific for L-amino acids and R-amines.

    Science.gov (United States)

    Boyko, Konstantin M; Stekhanova, Tatiana N; Nikolaeva, Alena Yu; Mardanov, Andrey V; Rakitin, Andrey L; Ravin, Nikolai V; Bezsudnova, Ekaterina Yu; Popov, Vladimir O

    2016-03-01

    The gene TUZN1299 from the genome of the hyperthermophilic archaeon Thermoproteus uzoniensis encoding a new 32.8 kDa branched-chain amino acid aminotransferase (BCAT) was expressed in Escherichia coli. The recombinant protein TUZN1299 was purified to homogeneity in the PLP-bound form. TUZN1299 was active towards branched-chain amino acids (L-Val, L-Leu, L-Ile) and showed low but detectable activity toward (R)-alpha-methylbenzylamine. The enzyme exhibits high-temperature optimum, thermal stability, and tolerance to organic solvents. The structure of an archaeal BCAT called TUZN1299 was solved for the first time (at 2.0 Å resolution). TUZN1299 has a typical BCAT type IV fold, and the organization of its active site is similar to that of bacterial BCATs. However, there are some differences in the amino acid composition of the active site.

  17. Cloning and inactivation of a branched-chain-amino-acid aminotransferase gene from Staphylococcus carnosus and characterization of the enzyme

    DEFF Research Database (Denmark)

    Madsen, Søren M; Beck, Hans Christian; Ravn, Peter

    2002-01-01

    Staphylococcus carnosus and Staphylococcus xylosus are widely used as aroma producers in the manufacture of dried fermented sausages. Catabolism of branched-chain amino acids (BCAAs) by these strains contributes to aroma formation by production of methyl-branched aldehydes and carboxy acids....... The first step in the catabolism is most likely a transamination reaction catalyzed by BCAA aminotransferases (IlvE proteins). In this study, we cloned the ilvE gene from S. carnosus by using degenerate oligonucleotides and PCR. We found that the deduced amino acid sequence was 80% identical......-branched carboxy acids, 2-methylpropanoic acid, 2-methylbutanoic acid, and 3-methylbutanoic acid, which derived from the BCAA catabolism, clearly emphasizing the role of IlvE in aroma formation. In contrast to previous reports, we found that IlvE was the only enzyme that catalyzed the deamination of BCAAs in S...

  18. Insulinotropic and Muscle Protein Synthetic Effects of Branched-Chain Amino Acids: Potential Therapy for Type 2 Diabetes and Sarcopenia

    Directory of Open Access Journals (Sweden)

    Darren G. Candow

    2012-11-01

    Full Text Available The loss of muscle mass and strength with aging (i.e., sarcopenia has a negative effect on functional independence and overall quality of life. One main contributing factor to sarcopenia is the reduced ability to increase skeletal muscle protein synthesis in response to habitual feeding, possibly due to a reduction in postprandial insulin release and an increase in insulin resistance. Branched-chain amino acids (BCAA, primarily leucine, increases the activation of pathways involved in muscle protein synthesis through insulin-dependent and independent mechanisms, which may help counteract the “anabolic resistance” to feeding in older adults. Leucine exhibits strong insulinotropic characteristics, which may increase amino acid availability for muscle protein synthesis, reduce muscle protein breakdown, and enhance glucose disposal to help maintain blood glucose homeostasis.

  19. Ingestion of branched-chain amino acids and tryptophan during sustained exercise in man: failure to affect performance

    DEFF Research Database (Denmark)

    Van Hall, Gerrit; Raaymakers, J S; Saris, W H

    1995-01-01

    1. An increased uptake of tryptophan in the brain may increase serotoninergic activity and recently has been suggested to be a cause of fatigue during prolonged exercise. The present study, therefore, investigates whether ingestion of tryptophan or the competing branched-chain amino acids (BCAAs...... tryptophan ingestion caused a 7- to 20-fold increase. Exercise time to exhaustion was not different between treatments (122 +/- 3 min). 3. The data suggest that manipulation of tryptophan supply to the brain either has no additional effect upon serotoninergic activity during prolonged exhaustive exercise......) or (3) a high dose of BCAA (18 g l-1). 2. These treatments greatly increased the plasma concentration of the respective amino acids. Using the kinetic parameters of transport of human brain capillaries, BCAA supplements were estimated to reduce brain tryptophan uptake at exhaustion by 8-12%, while...

  20. Transamination of branched chain amino acids (BCAA) in rat adipose tissue

    Energy Technology Data Exchange (ETDEWEB)

    Frick, G.P.; Goodman, H.M.

    1986-03-05

    Like most extrahepatic tissues, adipose tissue can transaminate the BCAA faster than they are oxidized. Catabolism of the BCAA by adipose tissue appears to be limited by the activity of branched chain ..cap alpha..-keto acid dehydrogenase (BCDH). Conditions which stimulate the activity of this intramitochondrial enzyme in tissue extracts also increase the rate at which (1-/sup 14/C)leucine (L) and (1-/sup 14/C)valine (V) are oxidized by tissue segments. However, when maximum rates of oxidation were measured, 10 mM L was oxidized to /sup 14/CO/sub 2/ 5 times faster than 10 mM V (30 +/- 2 vs. 6 +/- 1 nmol min/sup -1/ g tis/sup -1/). In contrast, the ..cap alpha..-keto analogs of L and V were oxidized by tissue segments at nearly equal rates which slightly exceeded the rate of L oxidation. These results suggested that transamination might limit the catabolism of V, perhaps due to its inaccessibility to transaminase. The distribution of transaminase activity in tissue extracts was determined after centrifugation to obtain mitochondrial and cytosolic fractions. L and V were transaminated at similar rates by enzymes in both fractions. Transaminase activity in the mitochondrial fraction was greater than that of the cytosol and exceeded the capacity of the tissue to oxidize L. Catabolism of BCAA may depend upon intramitochondrial transamination and oxidation of V may be slower than that of L because uptake of V by mitochondria may be slower than that of L.

  1. Biosynthesis of branched-chain amino acids is essential for effective symbioses between betarhizobia and Mimosa pudica.

    Science.gov (United States)

    Chen, Wen-Ming; Prell, Jurgen; James, Euan K; Sheu, Der-Shyan; Sheu, Shih-Yi

    2012-07-01

    Burkholderia phymatum STM815 and Cupriavidus taiwanensis LMG19424 are betaproteobacterial strains that can effectively nodulate several species of the large legume genus Mimosa. A Tn5 mutant, derived from B. phymatum STM815 (KM60), and another derived from C. taiwanensis LMG19424 (KM184-55) induced Fix(-) nodules on Mimosa pudica. The Tn5-interrupted genes of the mutants showed strong homologies to ilvE, which encodes a branched-chain amino acid aminotransferase, and leuC, which encodes the large subunit of isopropylmalate isomerase. Both enzymes are known to be involved in the biosynthetic pathways for branched-chain amino acids (BCAAs) (leucine, valine and isoleucine). The B. phymatum ilvE mutant, KM60, was not auxotrophic for BCAAs and could grow well on minimal medium with pyruvate as a carbon source and ammonia as a nitrogen source. However, it grew less efficiently than the wild-type (WT) strain when ammonia was substituted with valine or isoleucine as a nitrogen source. The BCAA aminotransferase activity of KM60 was significantly reduced relative to the WT strain, especially with isoleucine and valine as amino group donors. The C. taiwanensis leuC mutant, KM184-55, could not grow on a minimal medium with pyruvate as a carbon source and ammonia as a nitrogen source, but its growth was restored when leucine was added to the medium. The isopropylmalate isomerase activity of KM184-55 was completely lost compared with the WT strain. Both mutants recovered their respective enzyme activities after complementation with the WT ilvE or leuC genes and were subsequently able to grow as well as their parental strains on minimal medium. They were also able to form nitrogen-fixing nodules on M. pudica. We conclude that the biosynthesis of BCAAs is essential for the free-living growth of betarhizobia, as well as for their ability to form effective symbioses with their host plant.

  2. Potential therapeutic effects of branched-chain amino acids supplementation on resistance exercise-based muscle damage in humans

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    da Luz Claudia R

    2011-12-01

    Full Text Available Abstract Branched-chain amino acids (BCAA supplementation has been considered an interesting nutritional strategy to improve skeletal muscle protein turnover in several conditions. In this context, there is evidence that resistance exercise (RE-derived biochemical markers of muscle soreness (creatine kinase (CK, aldolase, myoglobin, soreness, and functional strength may be modulated by BCAA supplementation in order to favor of muscle adaptation. However, few studies have investigated such effects in well-controlled conditions in humans. Therefore, the aim of this short report is to describe the potential therapeutic effects of BCAA supplementation on RE-based muscle damage in humans. The main point is that BCAA supplementation may decrease some biochemical markers related with muscle soreness but this does not necessarily reflect on muscle functionality.

  3. Effects of glucose, glucose plus branched-chain amino acids, or placebo on bike performance over 100 km

    DEFF Research Database (Denmark)

    Madsen, Klavs; MacLean, David A; Kiens, Bente

    1996-01-01

    This study was undertaken to determine the effects of ingesting either glucose (trial G) or glucose plus branched-chain amino acids (BCAA: trial B), compared with placebo (trial P), during prolonged exercise. Nine well-trained cyclists with a maximal oxygen uptake of 63.1 +/- 1.5 ml O2. min-1.kg-1...... between the three trials (160.1 +/- 4.1, 157.2 +/- 4.5, and 159.8 +/- 3.7 min, respectively). In trial B, plasma BCAA levels increased from 339 +/- 28 microM at rest to 1,026 +/- 62 microM after exercise (P ... glucose plus BCAA ingestion during 100 km of cycling enhance performance in well-trained cyclists....

  4. Does Branched-Chain Amino Acids Supplementation Modulate Skeletal Muscle Remodeling through Inflammation Modulation? Possible Mechanisms of Action

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    Humberto Nicastro

    2012-01-01

    Full Text Available Skeletal muscle protein turnover is modulated by intracellular signaling pathways involved in protein synthesis, degradation, and inflammation. The proinflammatory status of muscle cells, observed in pathological conditions such as cancer, aging, and sepsis, can directly modulate protein translation initiation and muscle proteolysis, contributing to negative protein turnover. In this context, branched-chain amino acids (BCAAs, especially leucine, have been described as a strong nutritional stimulus able to enhance protein translation initiation and attenuate proteolysis. Furthermore, under inflammatory conditions, BCAA can be transaminated to glutamate in order to increase glutamine synthesis, which is a substrate highly consumed by inflammatory cells such as macrophages. The present paper describes the role of inflammation on muscle remodeling and the possible metabolic and cellular effects of BCAA supplementation in the modulation of inflammatory status of skeletal muscle and the consequences on protein synthesis and degradation.

  5. Color identification and fuzzy reasoning based monitoring and controlling of fermentation process of branched chain amino acid

    Science.gov (United States)

    Ma, Lei; Wang, Yizhong; Xu, Qingyang; Huang, Huafang; Zhang, Rui; Chen, Ning

    2009-11-01

    The main production method of branched chain amino acid (BCAA) is microbial fermentation. In this paper, to monitor and to control the fermentation process of BCAA, especially its logarithmic phase, parameters such as the color of fermentation broth, culture temperature, pH, revolution, dissolved oxygen, airflow rate, pressure, optical density, and residual glucose, are measured and/or controlled and/or adjusted. The color of fermentation broth is measured using the HIS color model and a BP neural network. The network's input is the histograms of hue H and saturation S, and output is the color description. Fermentation process parameters are adjusted using fuzzy reasoning, which is performed by inference rules. According to the practical situation of BCAA fermentation process, all parameters are divided into four grades, and different fuzzy rules are established.

  6. Ingestion of branched-chain amino acids and tryptophan during sustained exercise in man: failure to affect performance

    DEFF Research Database (Denmark)

    Van Hall, Gerrit; Raaymakers, J S; Saris, W H

    1995-01-01

    tryptophan ingestion caused a 7- to 20-fold increase. Exercise time to exhaustion was not different between treatments (122 +/- 3 min). 3. The data suggest that manipulation of tryptophan supply to the brain either has no additional effect upon serotoninergic activity during prolonged exhaustive exercise......1. An increased uptake of tryptophan in the brain may increase serotoninergic activity and recently has been suggested to be a cause of fatigue during prolonged exercise. The present study, therefore, investigates whether ingestion of tryptophan or the competing branched-chain amino acids (BCAAs......) affect performance. Ten endurance-trained male athletes were studied during cycle exercise at 70-75% maximal power output, while ingesting, ad random and double-blind, drinks that contained 6% sucrose (control) or 6% sucrose supplemented with (1) tryptophan (3 g l-1), (2) a low dose of BCAA (6 g l-1...

  7. Elevation of branched-chain amino acid levels in diabetes and NAFL and changes with antidiabetic drug treatment.

    Science.gov (United States)

    Iwasa, Motoh; Ishihara, Tomoaki; Mifuji-Moroka, Rumi; Fujita, Naoki; Kobayashi, Yoshinao; Hasegawa, Hiroshi; Iwata, Kazuko; Kaito, Masahiko; Takei, Yoshiyuki

    2015-01-01

    Diabetes mellitus (DM), non-alcoholic fatty liver (NAFL), and obesity are associated with elevated branched-chain amino acid (BCAA) levels, but the mechanism and significance of this has not been elucidated. Eighty-four subjects were enrolled including 43 with DM. Serum BCAA levels were positively correlated with waist-hip ratio and ALT. Serum BCAA levels in subjects with DM were higher than non-DM and those in subjects with NAFL were also higher than non-NAFL. Treatment with pioglitazone and alogliptin (19 of 43 DM subjects) improved serum haemoglobin A1c and decreased BCAA levels. The decrease in BCAAs with improved glucose metabolism suggests that abnormal glucose metabolism is also a factor in elevated BCAA levels.

  8. Branched-chain and aromatic amino acid catabolism into aroma volatiles in Cucumis melo L. fruit

    Science.gov (United States)

    The unique aroma of melons (Cucumis melo L., Cucurbitaceae) is composed of many volatile compounds biosynthetically derived from fatty-acids, carotenoids, amino-acids as well as terpenes. Incubation of melon fruit cubes with amino- and a-keto acids led to the enhanced formation of aroma compounds be...

  9. Defects in muscle branched-chain amino acid oxidation contribute to impaired lipid metabolism

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    Carles Lerin

    2016-10-01

    Conclusions: Our data indicate that impaired muscle BCAA catabolism may contribute to the development of insulin resistance by perturbing both amino acid and fatty acid metabolism and suggest that targeting BCAA metabolism may hold promise for prevention or treatment of T2D.

  10. L-citrulline-malate influence over branched chain amino acid utilization during exercise.

    Science.gov (United States)

    Sureda, Antoni; Córdova, Alfredo; Ferrer, Miguel D; Pérez, Gerardo; Tur, Josep A; Pons, Antoni

    2010-09-01

    Exhaustive exercise induces disturbances in metabolic homeostasis which can result in amino acid catabolism and limited L-arginine availability. Oral L-citrulline supplementation raises plasma L-arginine concentration and augments NO-dependent signalling. Our aim was to evaluate the effects of diet supplementation with L-citrulline-malate prior to intense exercise on the metabolic handle of plasma amino acids and on the products of metabolism of arginine as creatinine, urea and nitrite and the possible effects on the hormonal levels. Seventeen voluntary male pre-professional cyclists were randomly assigned to one of two groups: control or supplemented (6 g L-citrulline-malate 2 h prior exercise) and participated in a 137-km cycling stage. Blood samples were taken in basal conditions, 15 min after the race and 3 h post race (recovery). Most essential amino acids significantly decreased their plasma concentration as a result of exercise; however, most non-essential amino acids tended to significantly increase their concentration. Citrulline-malate ingestion significantly increased the plasma concentration of citrulline, arginine, ornithine, urea, creatinine and nitrite (p urea.

  11. Oral branched-chain amino acid supplements that reduce brain serotonin during exercise in rats also lower brain catecholamines.

    Science.gov (United States)

    Choi, Sujean; Disilvio, Briana; Fernstrom, Madelyn H; Fernstrom, John D

    2013-11-01

    Exercise raises brain serotonin release and is postulated to cause fatigue in athletes; ingestion of branched-chain amino acids (BCAA), by competitively inhibiting tryptophan transport into brain, lowers brain tryptophan uptake and serotonin synthesis and release in rats, and reputedly in humans prevents exercise-induced increases in serotonin and fatigue. This latter effect in humans is disputed. But BCAA also competitively inhibit tyrosine uptake into brain, and thus catecholamine synthesis and release. Since increasing brain catecholamines enhances physical performance, BCAA ingestion could lower catecholamines, reduce performance and thus negate any serotonin-linked benefit. We therefore examined in rats whether BCAA would reduce both brain tryptophan and tyrosine concentrations and serotonin and catecholamine synthesis. Sedentary and exercising rats received BCAA or vehicle orally; tryptophan and tyrosine concentrations and serotonin and catecholamine synthesis rates were measured 1 h later in brain. BCAA reduced brain tryptophan and tyrosine concentrations, and serotonin and catecholamine synthesis. These reductions in tyrosine concentrations and catecholamine synthesis, but not tryptophan or serotonin synthesis, could be prevented by co-administering tyrosine with BCAA. Complete essential amino acid mixtures, used to maintain or build muscle mass, were also studied, and produced different effects on brain tryptophan and tyrosine concentrations and serotonin and catecholamine synthesis. Since pharmacologically increasing brain catecholamine function improves physical performance, the finding that BCAA reduce catecholamine synthesis may explain why this treatment does not enhance physical performance in humans, despite reducing serotonin synthesis. If so, adding tyrosine to BCAA supplements might allow a positive action on performance to emerge.

  12. The influence of alternative pathways of respiration that utilize branched-chain amino acids following water shortage in Arabidopsis.

    Science.gov (United States)

    Pires, Marcel V; Pereira Júnior, Adilson A; Medeiros, David B; Daloso, Danilo M; Pham, Phuong Anh; Barros, Kallyne A; Engqvist, Martin K M; Florian, Alexandra; Krahnert, Ina; Maurino, Veronica G; Araújo, Wagner L; Fernie, Alisdair R

    2016-06-01

    During dark-induced senescence isovaleryl-CoA dehydrogenase (IVDH) and D-2-hydroxyglutarate dehydrogenase (D-2HGDH) act as alternate electron donors to the ubiquinol pool via the electron-transfer flavoprotein/electron-transfer flavoprotein:ubiquinone oxidoreductase (ETF/ETFQO) pathway. However, the role of this pathway in response to other stresses still remains unclear. Here, we demonstrated that this alternative pathway is associated with tolerance to drought in Arabidopsis. In comparison with wild type (WT) and lines overexpressing D-2GHDH, loss-of-function etfqo-1, d2hgdh-2 and ivdh-1 mutants displayed compromised respiration rates and were more sensitive to drought. Our results demonstrated that an operational ETF/ETFQO pathway is associated with plants' ability to withstand drought and to recover growth once water becomes replete. Drought-induced metabolic reprogramming resulted in an increase in tricarboxylic acid (TCA) cycle intermediates and total amino acid levels, as well as decreases in protein, starch and nitrate contents. The enhanced levels of the branched-chain amino acids in loss-of-function mutants appear to be related to their increased utilization as substrates for the TCA cycle under water stress. Our results thus show that mitochondrial metabolism is highly active during drought stress responses and provide support for a role of alternative respiratory pathways within this response.

  13. Branched Chain Amino Acids Are Associated with Insulin Resistance Independent of Leptin and Adiponectin in Subjects with Varying Degrees of Glucose Tolerance

    NARCIS (Netherlands)

    Connelly, Margery A.; Wolak-Dinsmore, Justyna; Dullaart, Robin P. F.

    Background: Branched chain amino acids (BCAA) may be involved in the pathogenesis of insulin resistance and are associated with type 2 diabetes mellitus (T2DM) development. Adipokines such as leptin and adiponectin influence insulin resistance and reflect adipocyte dysfunction. We examined the

  14. Branched Chain Amino Acids Are Associated with Insulin Resistance Independent of Leptin and Adiponectin in Subjects with Varying Degrees of Glucose Tolerance

    NARCIS (Netherlands)

    Connelly, Margery A.; Wolak-Dinsmore, Justyna; Dullaart, Robin P. F.

    2017-01-01

    Background: Branched chain amino acids (BCAA) may be involved in the pathogenesis of insulin resistance and are associated with type 2 diabetes mellitus (T2DM) development. Adipokines such as leptin and adiponectin influence insulin resistance and reflect adipocyte dysfunction. We examined the exten

  15. Circulating branched-chain amino acid concentrations are associated with obesity and future insulin resistance in children and adolescents.

    Science.gov (United States)

    McCormack, S E; Shaham, O; McCarthy, M A; Deik, A A; Wang, T J; Gerszten, R E; Clish, C B; Mootha, V K; Grinspoon, S K; Fleischman, A

    2013-02-01

    What is already known about this subject Circulating concentrations of branched-chain amino acids (BCAAs) can affect carbohydrate metabolism in skeletal muscle, and therefore may alter insulin sensitivity. BCAAs are elevated in adults with diet-induced obesity, and are associated with their future risk of type 2 diabetes even after accounting for baseline clinical risk factors. What this study adds Increased concentrations of BCAAs are already present in young obese children and their metabolomic profiles are consistent with increased BCAA catabolism. Elevations in BCAAs in children are positively associated with insulin resistance measured 18 months later, independent of their initial body mass index. Branched-chain amino acid (BCAA) concentrations are elevated in response to overnutrition, and can affect both insulin sensitivity and secretion. Alterations in their metabolism may therefore play a role in the early pathogenesis of type 2 diabetes in overweight children. To determine whether paediatric obesity is associated with elevations in fasting circulating concentrations of BCAAs (isoleucine, leucine and valine), and whether these elevations predict future insulin resistance. Sixty-nine healthy subjects, ages 8-18 years, were enrolled as a cross-sectional cohort. A subset of subjects who were pre- or early-pubertal, ages 8-13 years, were enrolled in a prospective longitudinal cohort for 18 months (n = 17 with complete data). Elevations in the concentrations of BCAAs were significantly associated with body mass index (BMI) Z-score (Spearman's Rho 0.27, P = 0.03) in the cross-sectional cohort. In the subset of subjects that followed longitudinally, baseline BCAA concentrations were positively associated with homeostasis model assessment for insulin resistance measured 18 months later after controlling for baseline clinical factors including BMI Z-score, sex and pubertal stage (P = 0.046). Elevations in the concentrations of circulating BCAAs are significantly

  16. The Supplementation of Branched-Chain Amino Acids, Arginine, and Citrulline Improves Endurance Exercise Performance in Two Consecutive Days

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    I-Shiung Cheng, Yi-Wen Wang, I-Fan Chen, Gi-Sheng Hsu, Chun-Fang Hsueh, Chen-Kang Chang

    2016-09-01

    Full Text Available The central nervous system plays a crucial role in fatigue during endurance exercise. Branched-chain amino acids (BCAA could reduce cerebral serotonin synthesis by competing with its precursor tryptophan for crossing the blood brain barrier. Arginine and citrulline could prevent excess hyperammonemia accompanied by BCAA supplementation. This study investigated the combination of BCAA, arginine, and citrulline on endurance performance in two consecutive days. Seven male and three female endurance runners ingested 0.17 g·kg-1 BCAA, 0.05 g·kg-1 arginine and 0.05 g·kg-1 citrulline (AA trial or placebo (PL trial in a randomized cross-over design. Each trial contained a 5000 m time trial on the first day, and a 10000 m time trial on the second day. The AA trial had significantly better performance in 5000 m (AA: 1065.7 ± 33.9 s; PL: 1100.5 ± 40.4 s and 10000 m (AA: 2292.0 ± 211.3 s; PL: 2375.6 ± 244.2 s. The two trials reported similar ratings of perceived exertion. After exercise, the AA trial had significantly lower tryptophan/BCAA ratio, similar NH3, and significantly higher urea concentrations. In conclusion, the supplementation could enhance time-trial performance in two consecutive days in endurance runners, possibly through the inhibition of cerebral serotonin synthesis by BCAA and the prevention of excess hyperammonemia by increased urea genesis.

  17. NUTRITIONAL INTERACTIONS BETWEEN ZINC AND BRANCHED CHAIN AMINO ACID (BCAA SUPPLEMENT IN RATS: A MULTICOMPARTMENT MODELING APPROACH

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    JAIR RODRIGUES GARCIA-JÊNIOR

    2009-07-01

    Full Text Available

    The influence of supplementary-branched chain amino acids (BCAA on 65Zn metabolism in rats was investigated in this study. Nutritional indicators of Zn, as absorption, body retention and secretion, were estimated using a multicompartment model. Two groups of eight male rats were force-fed a zinc-adequate diet (control group and a zinc-adequate diet plus 0.52 9 BCAA/kg diet during 15 days. There was no significant difference for intake of Zn, absorption (34%, intestinal transit (tso and the leveI of Zn in the intravascular compartment (plasma. On the other hand the extravascular compartment (organs and specific concentration of Zn per 9 of tissue decreased after experimental period (p < 0.05 The rats supplememted with BCAA secreted Zn by urine twice faster than controls, but the secrotion of zinc by endogen feces were not decreased in this group. Thus, BCAA supplement changed the kinetic of Zn, increasing the urinary secretion and the loss of Zn from the body.

  18. Branched-chain amino acid supplementation reduces oxidative stress and prolongs survival in rats with advanced liver cirrhosis.

    Directory of Open Access Journals (Sweden)

    Motoh Iwasa

    Full Text Available Long-term supplementation with branched-chain amino acids (BCAA is associated with prolonged survival and decreased frequency of development of hepatocellular carcinoma (HCC in patients with liver cirrhosis. However, the pharmaceutical mechanism underlying this association is still unclear. We investigated whether continuous BCAA supplementation increases survival rate of rats exposed to a fibrogenic agent and influences the iron accumulation, oxidative stress, fibrosis, and gluconeogenesis in the liver. Further, the effects of BCAA on gluconeogenesis in cultured cells were also investigated. A significant improvement in cumulative survival was observed in BCAA-supplemented rats with advanced cirrhosis compared to untreated rats with cirrhosis (P<0.05. The prolonged survival due to BCAA supplementation was associated with reduction of iron contents, reactive oxygen species production and attenuated fibrosis in the liver. In addition, BCAA ameliorated glucose metabolism by forkhead box protein O1 pathway in the liver. BCAA prolongs survival in cirrhotic rats and this was likely the consequences of reduced iron accumulation, oxidative stress and fibrosis and improved glucose metabolism in the liver.

  19. Is administrating branched-chain amino acid-enriched nutrition achieved symptom-free in malnourished cirrhotic patients?

    Science.gov (United States)

    Tsuda, Yasuhiro; Fukui, Hideo; Sujishi, Tetsuya; Ohama, Hideko; Tsuchimoto, Yusuke; Asai, Akira; Fukunisi, Shinya; Higuchi, Kazuhide

    2014-01-01

    Administration of branched-chain amino acids (BCAA) has been reported to improve liver function, quality of life (QOL). However, in some malnourished patients, serum albumin levels do not improve in response to BCAA granules. In this study, we examined the effects of BCAA-enriched enteral nutrition in patients unresponsive to BCAA granules. Thirty-two decompensated cirrhotic patients at Osaka Medical College were enrolled in this study. Since all patients showed no improvement in serum albumin levels despite 3 months of BCAA granule administration, they were administered 50 g of a flavored BCAA-enriched enteral nutrient twice daily, i.e., during the daytime and late evening. Serum albumin levels and major cirrhotic symptoms were examined 1, 3, and 5 months after treatment initiation. Serum albumin levels improved significantly 3 months after treatment initiation (3.14 ± 0.32 g/dl vs 3.5 ± 0.31 g/dl, pBCAA-enriched nutrients improves QOL of cirrhotic patients unresponsive to BCAA granules.

  20. Cross-talk between branched-chain amino acids and hepatic mitochondria is compromised in nonalcoholic fatty liver disease.

    Science.gov (United States)

    Sunny, Nishanth E; Kalavalapalli, Srilaxmi; Bril, Fernando; Garrett, Timothy J; Nautiyal, Manisha; Mathew, Justin T; Williams, Caroline M; Cusi, Kenneth

    2015-08-15

    Elevated plasma branched-chain amino acids (BCAA) in the setting of insulin resistance have been relevant in predicting type 2 diabetes mellitus (T2DM) onset, but their role in the etiology of hepatic insulin resistance remains uncertain. We determined the link between BCAA and dysfunctional hepatic tricarboxylic acid (TCA) cycle, which is a central feature of hepatic insulin resistance and nonalcoholic fatty liver disease (NAFLD). Plasma metabolites under basal fasting and euglycemic hyperinsulinemic clamps (insulin stimulation) were measured in 94 human subjects with varying degrees of insulin sensitivity to identify their relationships with insulin resistance. Furthermore, the impact of elevated BCAA on hepatic TCA cycle was determined in a diet-induced mouse model of NAFLD, utilizing targeted metabolomics and nuclear magnetic resonance (NMR)-based metabolic flux analysis. Insulin stimulation revealed robust relationships between human plasma BCAA and indices of insulin resistance, indicating chronic metabolic overload from BCAA. Human plasma BCAA and long-chain acylcarnitines also showed a positive correlation, suggesting modulation of mitochondrial metabolism by BCAA. Concurrently, mice with NAFLD failed to optimally induce hepatic mTORC1, plasma ketones, and hepatic long-chain acylcarnitines, following acute elevation of plasma BCAA. Furthermore, elevated BCAA failed to induce multiple fluxes through hepatic TCA cycle in mice with NAFLD. Our data suggest that BCAA are essential to mediate efficient channeling of carbon substrates for oxidation through mitochondrial TCA cycle. Impairment of BCAA-mediated upregulation of the TCA cycle could be a significant contributor to mitochondrial dysfunction in NAFLD.

  1. Branched-chain amino acid-rich diet improves skeletal muscle wasting caused by cigarette smoke in rats.

    Science.gov (United States)

    Tomoda, Koichi; Kubo, Kaoru; Hino, Kazuo; Kondoh, Yasunori; Nishii, Yasue; Koyama, Noriko; Yamamoto, Yoshifumi; Yoshikawa, Masanori; Kimura, Hiroshi

    2014-04-01

    Cigarette smoke induces skeletal muscle wasting by a mechanism not yet fully elucidated. Branched-chain amino acids (BCAA) in the skeletal muscles are useful energy sources during exercise or systemic stresses. We investigated the relationship between skeletal muscle wasting caused by cigarette smoke and changes in BCAA levels in the plasma and skeletal muscles of rats. Furthermore, the effects of BCAA-rich diet on muscle wasting caused by cigarette smoke were also investigated. Wistar Kyoto (WKY) rats that were fed with a control or a BCAA-rich diet were exposed to cigarette smoke for four weeks. After the exposure, the skeletal muscle weight and BCAA levels in plasma and the skeletal muscles were measured. Cigarette smoke significantly decreased the skeletal muscle weight and BCAA levels in both plasma and skeletal muscles, while a BCAA-rich diet increased the skeletal muscle weight and BCAA levels in both plasma and skeletal muscles that had decreased by cigarette smoke exposure. In conclusion, skeletal muscle wasting caused by cigarette smoke was related to the decrease of BCAA levels in the skeletal muscles, while a BCAA-rich diet may improve cases of cigarette smoke-induced skeletal muscle wasting.

  2. Effects of eccentric exercise on branched-chain amino acid profiles in rat serum and skeletal muscle.

    Science.gov (United States)

    Qun, Z; Xinkai, Y; Jing, W

    2014-04-01

    Supplementation of branched-chain amino acid (BCAA) is often used to attenuate exercise-induced skeletal muscle damage and promote adaptation, but no definitive conclusion on the benefits of BCAA on muscle recovery after injurious exercise can be drawn. Exploration of the systematic BCAA alteration in muscular injury-repair stage per se without any BCAA supplement should provide some useful information in favour of BCAA application in muscle regeneration after injury. One bout of 90-min downhill-running exercise was performed to cause rat skeletal muscle injury. After exercise, myofibrillar BCAA concentrations showed minor changes compared with exercise before, while serum concentrations of BCAA were lower after exercise. Especially, serum leucine, isoleucine and total BCAA concentrations 2 weeks post-run were significantly lower than normal values of exercise before (p = 0.008, p = 0.041, p = 0.015). The data demonstrate that a single eccentric exercise can significantly decrease the serum BCAA concentrations, which mean high utilization of BCAA for myogenesis after injurious exercise.

  3. Branched-chain amino acids to tyrosine ratio value as a potential prognostic factor for hepatocellular carcinoma

    Institute of Scientific and Technical Information of China (English)

    Toru Ishikawa

    2012-01-01

    The prognosis of hepatocellular carcinoma (HCC) depends on tumor extension as well as hepatic function.Hepatic functional reserve is recognized as a factor affecting survival in the treatment of HCC; the ChildPugh classification system is the most extensively used method for assessing hepatic functional reserve in patients with chronic liver disease,using serum albumin level to achieve accurate assessment of the status of protein metabolism.However,insufficient attention has been given to the status of amino acid (AA) metabolism in chronic liver disease and HCC.Fischer's ratio is the molar ratio of branched-chain AAs (BCAAs:leucine,valine,isoleucine) to aromatic AAs (phenylalanine,tyrosine) and is important for assessing liver metabolism,hepatic functional reserve and the severity of liver dysfunction.Although this ratio is difficult to determine in clinical situations,BCAAs/tyrosine molar concentration ratio (BTR) has been proposed as a simpler substitute.BTR correlates with various liver function examinations,including markers of hepatic fibrosis,hepatic blood flow and hepatocyte function,and can thus be considered as reflecting the degree of hepatic impairment.This manuscript examines the literature to clarify whether BTR can serve as a prognostic factor for treatment of HCC.

  4. Branched-chain amino acids to tyrosine ratio value as a potential prognostic factor for hepatocellular carcinoma.

    Science.gov (United States)

    Ishikawa, Toru

    2012-05-07

    The prognosis of hepatocellular carcinoma (HCC) depends on tumor extension as well as hepatic function. Hepatic functional reserve is recognized as a factor affecting survival in the treatment of HCC; the Child-Pugh classification system is the most extensively used method for assessing hepatic functional reserve in patients with chronic liver disease, using serum albumin level to achieve accurate assessment of the status of protein metabolism. However, insufficient attention has been given to the status of amino acid (AA) metabolism in chronic liver disease and HCC. Fischer's ratio is the molar ratio of branched-chain AAs (BCAAs: leucine, valine, isoleucine) to aromatic AAs (phenylalanine, tyrosine) and is important for assessing liver metabolism, hepatic functional reserve and the severity of liver dysfunction. Although this ratio is difficult to determine in clinical situations, BCAAs/tyrosine molar concentration ratio (BTR) has been proposed as a simpler substitute. BTR correlates with various liver function examinations, including markers of hepatic fibrosis, hepatic blood flow and hepatocyte function, and can thus be considered as reflecting the degree of hepatic impairment. This manuscript examines the literature to clarify whether BTR can serve as a prognostic factor for treatment of HCC.

  5. Influence of lysolecithin and Tween 80 on the colloidal stability of branched chain amino acids in a nanosuspension system.

    Science.gov (United States)

    Hong, Chi Rac; Lee, Gyu Whan; Paik, Hyun-Dong; Chang, Pahn-Shick; Choi, Seung Jun

    2017-04-15

    This study examined the influence of stabilizers on the solubility and colloidal stability of branched chain amino acids (BCAAs) nanosuspended through high pressure homogenization at 70°C. Although homogenization increased the initial BCAA solubility, irrespective of pH (pH 3 or 6), homogenization alone was not sufficient to increase their long-term solubility. The incorporation of stabilizers into nanosuspensions increased the saturation concentration of BCAAs but the effect of stabilizers on the increase in the saturation concentration of BCAAs was more pronounced at pH 6.0. At pH 6, Tween 80 dramatically increased the colloidal stability of the BCAA nanosuspensions, independent of the BCAA:stabilizer ratio but not at pH 3. However, the effect of lysolecithin on the colloidal stability of nanosuspended BCAAs varied depending on pH and BCAA:lysolecithin ratio. In lysolecithin-related nanosuspensions, there was no clear relationship between the colloidal stability and nanosuspension conditions including pH and BCAA:lysolecithin ratio. This study could provide a useful information on stabilizer selection for the development of liquid or colloidal products with improved solubility and colloidal stability of nanosuspended BCAAs.

  6. Branched-Chain Amino Acids as Predictors for Individual Differences of Cisplatin Nephrotoxicity in Rats: A Pharmacometabonomics Study.

    Science.gov (United States)

    Zhang, Pei; Li, Wei; Chen, Jiaqing; Li, Ruiting; Zhang, Zunjian; Huang, Yin; Xu, Fengguo

    2017-03-17

    Nephrotoxicity is the dose-limiting adverse effect of cisplatin with large individual differences. Up to now, little has been done on how to recognize and predict the individual differences in either preclinical or clinical research. In the present study, important postdose indicators were screened out first and integrated into a grouping factor, according to which rats were recognized as lowly or highly sensitive individuals. Then, mass-spectrometry-based untargeted metabolomics approach was performed to dissect the metabolic differences in predose serum of the two groups. Eventually, branched-chain amino acids (BCAAs) were found to be most significant with the lowest p value of Mann-Whitney U test and the highest area under receiving operating characteristic curve (AUC-ROC). The findings were further confirmed by absolute quantitation of BCAAs using liquid chromatography-tandem mass spectrometry. Binary logistic regression showed that in the discovery set absolute BCAA contents in rat predose serum could predict cisplatin nephrotoxicity with accuracy of 85%. This result was validated by another two independent external validation sets with accuracy of 81.8 and 78.8%, respectively. This study could provide new insight into cisplatin nephrotoxicity and may help expedite personalized medicine of cisplatin or other antitumor drugs in future clinical studies.

  7. Branched Chain Amino Acids Cause Liver Injury in Obese/Diabetic Mice by Promoting Adipocyte Lipolysis and Inhibiting Hepatic Autophagy

    Directory of Open Access Journals (Sweden)

    Fuyang Zhang

    2016-11-01

    Full Text Available The Western meat-rich diet is both high in protein and fat. Although the hazardous effect of a high fat diet (HFD upon liver structure and function is well recognized, whether the co-presence of high protein intake contributes to, or protects against, HF-induced hepatic injury remains unclear. Increased intake of branched chain amino acids (BCAA, essential amino acids compromising 20% of total protein intake reduces body weight. However, elevated circulating BCAA is associated with non-alcoholic fatty liver disease and injury. The mechanisms responsible for this quandary remain unknown; the role of BCAA in HF-induced liver injury is unclear. Utilizing HFD or HFD + BCAA models, we demonstrated BCAA supplementation attenuated HFD-induced weight gain, decreased fat mass, activated mammalian target of rapamycin (mTOR, inhibited hepatic lipogenic enzymes, and reduced hepatic triglyceride content. However, BCAA caused significant hepatic damage in HFD mice, evidenced by exacerbated hepatic oxidative stress, increased hepatic apoptosis, and elevated circulation hepatic enzymes. Compared to solely HFD-fed animals, plasma levels of free fatty acids (FFA in the HFD + BCAA group are significantly further increased, due largely to AMPKα2-mediated adipocyte lipolysis. Lipolysis inhibition normalized plasma FFA levels, and improved insulin sensitivity. Surprisingly, blocking lipolysis failed to abolish BCAA-induced liver injury. Mechanistically, hepatic mTOR activation by BCAA inhibited lipid-induced hepatic autophagy, increased hepatic apoptosis, blocked hepatic FFA/triglyceride conversion, and increased hepatocyte susceptibility to FFA-mediated lipotoxicity. These data demonstrated that BCAA reduces HFD-induced body weight, at the expense of abnormal lipolysis and hyperlipidemia, causing hepatic lipotoxicity. Furthermore, BCAA directly exacerbate hepatic lipotoxicity by reducing lipogenesis and inhibiting autophagy in the hepatocyte.

  8. Branched Chain Amino Acids Cause Liver Injury in Obese/Diabetic Mice by Promoting Adipocyte Lipolysis and Inhibiting Hepatic Autophagy.

    Science.gov (United States)

    Zhang, Fuyang; Zhao, Shihao; Yan, Wenjun; Xia, Yunlong; Chen, Xiyao; Wang, Wei; Zhang, Jinglong; Gao, Chao; Peng, Cheng; Yan, Feng; Zhao, Huishou; Lian, Kun; Lee, Yan; Zhang, Ling; Lau, Wayne Bond; Ma, Xinliang; Tao, Ling

    2016-11-01

    The Western meat-rich diet is both high in protein and fat. Although the hazardous effect of a high fat diet (HFD) upon liver structure and function is well recognized, whether the co-presence of high protein intake contributes to, or protects against, HF-induced hepatic injury remains unclear. Increased intake of branched chain amino acids (BCAA, essential amino acids compromising 20% of total protein intake) reduces body weight. However, elevated circulating BCAA is associated with non-alcoholic fatty liver disease and injury. The mechanisms responsible for this quandary remain unknown; the role of BCAA in HF-induced liver injury is unclear. Utilizing HFD or HFD+BCAA models, we demonstrated BCAA supplementation attenuated HFD-induced weight gain, decreased fat mass, activated mammalian target of rapamycin (mTOR), inhibited hepatic lipogenic enzymes, and reduced hepatic triglyceride content. However, BCAA caused significant hepatic damage in HFD mice, evidenced by exacerbated hepatic oxidative stress, increased hepatic apoptosis, and elevated circulation hepatic enzymes. Compared to solely HFD-fed animals, plasma levels of free fatty acids (FFA) in the HFD+BCAA group are significantly further increased, due largely to AMPKα2-mediated adipocyte lipolysis. Lipolysis inhibition normalized plasma FFA levels, and improved insulin sensitivity. Surprisingly, blocking lipolysis failed to abolish BCAA-induced liver injury. Mechanistically, hepatic mTOR activation by BCAA inhibited lipid-induced hepatic autophagy, increased hepatic apoptosis, blocked hepatic FFA/triglyceride conversion, and increased hepatocyte susceptibility to FFA-mediated lipotoxicity. These data demonstrated that BCAA reduces HFD-induced body weight, at the expense of abnormal lipolysis and hyperlipidemia, causing hepatic lipotoxicity. Furthermore, BCAA directly exacerbate hepatic lipotoxicity by reducing lipogenesis and inhibiting autophagy in the hepatocyte.

  9. Chronic dietary exposure to branched chain amino acids impairs glucose disposal in vegans but not in omnivores.

    Science.gov (United States)

    Gojda, J; Rossmeislová, L; Straková, R; Tůmová, J; Elkalaf, M; Jaček, M; Tůma, P; Potočková, J; Krauzová, E; Waldauf, P; Trnka, J; Štich, V; Anděl, M

    2017-05-01

    Branched chain amino acids (BCAA) are among nutrients strongly linked with insulin sensitivity (IS) measures. We investigated the effects of a chronic increase of BCAA intake on IS in two groups of healthy subjects differing in their basal consumption of BCAA, that is, vegans and omnivores. Eight vegans and eight matched omnivores (five men and three women in each group) received 15 g (women) or 20 g (men) of BCAA daily for 3 months. Anthropometry, blood analyses, glucose clamp, arginine test, subcutaneous abdominal adipose tissue (AT) and skeletal muscle (SM) biopsies (mRNA levels of selected metabolic markers, respiratory chain (RC) activity) were performed at baseline, after the intervention and after a 6 month wash-out period. Compared with omnivores, vegans had higher IS at baseline (GIR, glucose infusion rate: 9.6±2.4 vs 7.1±2.4 mg/kg/min, 95% CI for difference: 0.55 to 5.82) that declined after the intervention and returned to baseline values after the wash-out period (changes in GIR with 95% CI, 3-0 months: -1.64 [-2.5; -0.75] and 9-3 months: 1.65 [0.75; 2.54] mg/kg/min). No such change was observed in omnivores. In omnivores the intervention led to an increased expression of lipogenic genes (DGAT2, FASN, PPARγ, SCD1) in AT. SM RC activity increased in both groups. Negative impact of increased BCAA intake on IS was only detected in vegans, that is, subjects with low basal amino acids/BCAA intake, which appear to be unable to induce sufficient compensatory changes within AT and SM on a BCAA challenge.

  10. Aspectos atuais sobre aminoácidos de cadeia ramificada e exercício físico Current aspects of branched chain amino acid and exercise

    Directory of Open Access Journals (Sweden)

    Marcelo Macedo Rogero

    2008-12-01

    Full Text Available Em humanos saudáveis, nove aminoácidos são considerados essenciais, uma vez que não podem ser sintetizados endogenamente e, portanto, devem ser ingeridos por meio da dieta. Dentre os aminoácidos essenciais, se incluem os três aminoácidos de cadeia ramificada, ou seja, leucina, valina e isoleucina. Esses aminoácidos participam da regulação do balanço protéico corporal além de serem fonte de nitrogênio para a síntese de alanina e glutamina. No tocante à regulação da síntese protéica muscular, verifica-se que a leucina age estimulando a fase de iniciação da tradução do RNA-mensageiro em proteína, por mecanismos tanto dependentes quanto independentes de insulina. No que concerne ao exercício físico, supõe-se que esses aminoácidos estejam envolvidos na fadiga central, no balanço protéico muscular, na secreção de insulina, na modulação da imunocompetência, no aumento da performance de indivíduos que se exercitam em ambientes quentes e na diminuição do grau de lesão muscular. Nesse contexto, essa revisão aborda os aspectos atuais do metabolismo e da suplementação de aminoácidos de cadeia ramificada no exercício físico.In healthy humans, nine amino acids are considered to be essential once they cannot be endogenously synthesised and must therefore be ingested in the diet. Amongst the essential amino acids are the three branched chain amino acids, namely, leucine, valine and isoleucine. These amino acids participate in the regulation of protein balance in addition to being nitrogen sources for the synthesis of alanine and glutamine. As to the regulation of muscle protein synthesis, leucine acts in the stimulation of initiation of mRNA translation into protein, both through mechanisms that are dependent and independent of insulin. In the physiology of physical exercise, these branched amino acids play a role in central fatigue hypothesis, in muscle protein balance, in the secretion of insulin, in the

  11. A novel branched chain amino acids responsive transcriptional regulator, BCARR, negatively acts on the proteolytic system in Lactobacillus helveticus.

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    Taketo Wakai

    Full Text Available Transcriptional negative regulation of the proteolytic system of Lactobacillus helveticus CM4 in response to amino acids seems to be very important for the control of antihypertensive peptide production; however, it remains poorly understood. A 26-kDa protein with N-terminal cystathionine β-synthase domains (CBS domain protein, which seems to be involved in the regulatory system, was purified by using a DNA-sepharose bound 300-bp DNA fragment corresponding to the upstream regions of the six proteolytic genes that are down-regulated by amino acids. The CBS domain protein bound to a DNA fragment corresponding to the region upstream of the pepV gene in response to branched chain amino acids (BCAAs. The expression of the pepV gene in Escherichia coli grown in BCAA-enriched medium was repressed when the CBS domain protein was co-expressed. These results reveal that the CBS domain protein acts as a novel type of BCAA-responsive transcriptional regulator (BCARR in L. helveticus. From comparative analysis of the promoter regions of the six proteolysis genes, a palindromic AT-rich motif, 5'-AAAAANNCTWTTATT-3', was predicted as the consensus DNA motif for the BCARR protein binding. Footprint analysis using the pepV promotor region and gel shift analyses with the corresponding short DNA fragments strongly suggested that the BCARR protein binds adjacent to the pepV promoter region and affects the transcription level of the pepV gene in the presence of BCAAs. Homology search analysis of the C-terminal region of the BCARR protein suggested the existence of a unique βαββαβ fold structure that has been reported in a variety of ACT (aspartate kinase-chorismate mutase-tyrA domain proteins for sensing amino acids. These results also suggest that the sensing of BCAAs by the ACT domain might promote the binding of the BCARR to DNA sequences upstream of proteolysis genes, which affects the gene expression of the proteolytic system in L. helveticus.

  12. Branched chain and aromatic amino acids change acutely following two medical therapies for type 2 diabetes mellitus

    Science.gov (United States)

    Walford, Geoffrey A.; Davis, Jaclyn; Warner, A. Sofia; Ackerman, Rachel J.; Billings, Liana K.; Chamarthi, Bindu; Fanelli, Rebecca R.; Hernandez, Alicia M.; Huang, Chunmei; Khan, Sabina Q.; Littleton, Katherine R.; Lo, Janet; McCarthy, Rita M.; Rhee, Eugene P.; Deik, Amy; Stolerman, Elliot; Taylor, Andrew; Hudson, Margo S.; Wang, Thomas J.; Altshuler, David; Grant, Richard W.; Clish, Clary B.; Gerszten, Robert E.; Florez, Jose C.

    2013-01-01

    Objective Elevated circulating levels of branched chain and aromatic amino acids (BCAA/AAAs) are associated with insulin resistance and incident type 2 diabetes (T2D). BCAA/AAAs decrease acutely during an oral glucose tolerance test (OGTT), a diagnostic test for T2D. It is unknown whether changes in BCAA/AAAs also signal an early response to commonly used medical therapies for T2D. Materials and Methods A liquid chromatography-mass spectrometry approach was used to measure BCAA/AAAs in 30 insulin sensitive (IS) and 30 insulin resistant (IR) subjects before and after: 1) one dose of a sulfonylurea medication, glipizide, 5 mg orally; 2) two days of twice daily metformin 500 mg orally; and 3) a 75-gram OGTT. Percent change in BCAA/AAAs was determined after each intervention. Results Following glipizide, which increased insulin and decreased glucose in both subject groups, BCAA/AAAs decreased in the IS subjects only (all P<0.05). Following metformin, which decreased glucose and insulin in only the IR subjects, 4 BCAA/AAAs increased in the IR subjects at or below P=0.05, and none changed in the IS subjects. Following OGTT, which increased glucose and insulin in all subjects, BCAA/AAAs decreased in all subjects (P<0.05). Conclusions BCAA/AAAs changed acutely during glipizide and metformin administration, and the magnitude and direction of change differed by the insulin resistance status of the individual and the intervention. These results indicate that BCAA/AAAs may be useful biomarkers for monitoring the early response to therapeutic interventions for T2D. PMID:23953891

  13. Branched chain amino acid suppresses hepatocellular cancer stem cells through the activation of mammalian target of rapamycin.

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    Shinobu Nishitani

    Full Text Available Differentiation of cancer stem cells (CSCs into cancer cells causes increased sensitivity to chemotherapeutic agents. Although inhibition of mammalian target of rapamycin (mTOR leads to CSC survival, the effect of branched chain amino acids (BCAAs, an mTOR complex 1 (mTORC1 activator remains unknown. In this study, we examined the effects of BCAA on hepatocellular carcinoma (HCC cells expressing a hepatic CSC marker, EpCAM. We examined the effects of BCAA and/or 5-fluorouracil (FU on expression of EpCAM and other CSC-related markers, as well as cell proliferation in HCC cells and in a xenograft mouse model. We also characterized CSC-related and mTOR signal-related molecule expression and tumorigenicity in HCC cells with knockdown of Rictor or Raptor, or overexpression of constitutively active rheb (caRheb. mTOR signal-related molecule expression was also examined in BCAA-treated HCC cells. In-vitro BCAA reduced the frequency of EpCAM-positive cells and improved sensitivity to the anti-proliferative effect of 5-FU. Combined 5-FU and BCAA provided better antitumor efficacy than 5-FU alone in the xenograft model. Stimulation with high doses of BCAA activated mTORC1. Knockdown and overexpression experiments revealed that inhibition of mTOR complex 2 (mTORC2 or activation of mTORC1 led to decreased EpCAM expression and little or no tumorigenicity. BCAA may enhance the sensitivity to chemotherapy by reducing the population of cscs via the mTOR pathway. This result suggests the utility of BCAA in liver cancer therapy.

  14. Effects of carbohydrate, branched-chain amino acids, and arginine in recovery period on the subsequent performance in wrestlers

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    Jang Tsong-Rong

    2011-11-01

    Full Text Available Abstract Many athletes need to participate in multiple events in a single day. The efficient post-exercise glycogen recovery may be critical for the performance in subsequent exercise. This study examined whether post-exercise carbohydrate supplementation could restore the performance in the subsequent simulated wrestling match. The effect of branched-chain amino acids and arginine on glucose disposal and performance was also investigated. Nine well-trained male wrestlers participated in 3 trials in a random order. Each trial contained 3 matches with a 1-hr rest between match 1 and 2, and a 2-hr rest between match 2 and 3. Each match contained 3 exercise periods interspersed with 1-min rests. The subjects alternated 10-s all-out sprints and 20-s rests in each exercise period. At the end of match 2, 3 different supplementations were consumed: 1.2 g/kg glucose (CHO trial, 1 g/kg glucose + 0.1 g/kg Arg + 0.1 g/kg BCAA (CHO+AA trial, or water (placebo trial. The peak and average power in the 3 matches was similar in the 3 trials. After the supplementation, CHO and CHO+AA trial showed significantly higher glucose and insulin, and lower glycerol and non-esterified fatty acid concentrations than the placebo trial. There was no significant difference in these biochemical parameters between the CHO and CHO+AA trials. Supplementation of carbohydrate with or without BCAA and arginine during the post-match period had no effect on the performance in the following simulated match in wrestlers. In addition, BCAA and arginine did not provide additional insulinemic effect.

  15. Branched-Chain Amino Acid Levels Are Related with Surrogates of Disturbed Lipid Metabolism among Older Men

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    Urho M Kujala

    2016-11-01

    Full Text Available Aims/hypothesis Existing studies suggest that decreased branched-chain amino acid (BCAA catabolism and thus elevated levels in blood are associated with metabolic disturbances. Based on such information we have developed a hypothesis how BCAA degradation mechanistically connects to tricarboxylic acid (TCA cycle, intramyocellular lipid storage and oxidation thus allowing more efficient mitochondrial energy production from lipids as well as providing better metabolic health. We analyzed whether data from aged Finnish men are in line with our mechanistic hypothesis linking BCAA catabolism and metabolic disturbances. Methods Older Finnish men enriched with individuals having been athletes in young adulthood (n=593; mean age 72.6 ± 5.9 years responded to questionnaires, participated in a clinical examination including assessment of body composition with bioimpedance and gave fasting blood samples for various analytes as well as participated in a 2 hour 75 g oral glucose tolerance test. Metabolomics measurements from serum included BCAAs (isoleucine, leucine and valine.Results Out of the 593 participants 59 had previously known type 2 diabetes, further 67 had screen-detected type 2 diabetes, 127 IGT and 125 IFG while 214 had normal glucose regulation. There were group differences in all of the BCAA concentrations (p≤0.005 for all BCAAs, such that those with normal glucose tolerance had the lowest and those with diabetes mellitus had the highest BCAA concentrations. All BCAA levels correlated positively with body fat percentage (r=.29 - .34, p<.0001 for all. Expected associations with high BCAA concentrations and unfavorable metabolic profile indicators from metabolomics analysis were found. Except for glucose concentrations, the associations were stronger with isoleucine and leucine than with valine. Conclusions/interpretation The findings provided further support for our hypothesis by strengthening the idea that the efficiency of BCAA catabolism

  16. Structure of the Branched-chain Amino Acid and GTP-sensing Global Regulator, CodY, from Bacillus subtilis*

    Science.gov (United States)

    Levdikov, Vladimir M.; Blagova, Elena; Young, Vicki L.; Belitsky, Boris R.; Lebedev, Andrey; Sonenshein, Abraham L.

    2017-01-01

    CodY is a branched-chain amino acid (BCAA) and GTP sensor and a global regulator of transcription in low G + C Gram-positive bacteria. It controls the expression of over 100 genes and operons, principally by repressing during growth genes whose products are required for adaptations to nutrient limitation. However, the mechanism by which BCAA binding regulates transcriptional changes is not clear. It is known that CodY consists of a GAF (cGMP-stimulated phosphodiesterases, adenylate cyclases, FhlA) domain that binds BCAAs and a winged helix-turn-helix (wHTH) domain that binds to DNA, but the way in which these domains interact and the structural basis of the BCAA dependence of this interaction are unknown. To gain new insights, we determined the crystal structure of unliganded CodY from Bacillus subtilis revealing a 10-turn α-helix linking otherwise discrete GAF and wHTH domains. The structure of CodY in complex with isoleucine revealed a reorganized GAF domain. In both complexes CodY was tetrameric. Size exclusion chromatography with multiangle laser light scattering (SEC-MALLS) experiments showed that CodY is a dimer at concentrations found in bacterial cells. Comparison of structures of dimers of unliganded CodY and CodY-Ile derived from the tetramers showed a splaying of the wHTH domains when Ile was bound; splaying is likely to account for the increased affinity of Ile-bound CodY for DNA. Electrophoretic mobility shift and SEC-MALLS analyses of CodY binding to 19–36-bp operator fragments are consistent with isoleucine-dependent binding of two CodY dimers per duplex. The implications of these observations for effector control of CodY activity are discussed. PMID:28011634

  17. Effects of oral branched-chain amino acids on hepatic encephalopathy and outcome in patients with liver cirrhosis.

    Science.gov (United States)

    Kawaguchi, Takumi; Taniguchi, Eitaro; Sata, Michio

    2013-10-01

    Branched-chain amino acids (BCAAs) constituting of valine, leucine, and isoleucine act as both substrates of proteins and as key regulators for various nutrient metabolisms. Patients with liver cirrhosis frequently lack sufficient BCAAs and therefore suffer from various metabolic disorders. Hepatic encephalopathy (HE) is a severe metabolic disorder with neurologic manifestations such as flapping tremors and coma in patients with liver cirrhosis. In addition, a mild form of HE known as minimal HE (MHE) is an important social issue because it occurs in up to 80% of patients with chronic liver disease and affects prognosis and activities of daily living, possibly resulting in falls and motor vehicle accidents. Although HE/MHE can be caused by various pathological conditions, including in an accumulation of mercaptans, short-chain fatty acids, and alterations in the gut flora, hyperammonemia has also been implicated in an important pathogenesis of HE/MHE. Besides urea cycle of liver, ammonia can be detoxified in the skeletal muscles by the amidation process for glutamine synthesis using BCAAs. Thus, BCAA supplementation may enhance detoxification of ammonia in skeletal muscle and may be a possible therapeutic strategy for HE/MHE. In this review, we summarize the clinical impacts of BCAA supplementation on HE/MHE and discuss possible mechanisms for a BCAA-induced improvement of HE/MHE. Furthermore, we present some modifications of oral BCAA therapy for improvement of efficacy in HE treatment. We also briefly describe pleiotropic benefits of BCAAs on life-threatening events and overall prognosis in patients with liver cirrhosis.

  18. Branched-Chain Amino Acids as New Biomarkers of Major Depression - A Novel Neurobiology of Mood Disorder

    Science.gov (United States)

    Baranyi, Andreas; Amouzadeh-Ghadikolai, Omid; von Lewinski, Dirk; Rothenhäusler, Hans-Bernd; Theokas, Simon; Robier, Christoph; Mangge, Harald; Reicht, Gerhard; Hlade, Peter; Meinitzer, Andreas

    2016-01-01

    Background The proteinogenic branched-chain amino acids (BCAAs) valine, leucine and isoleucine might play an unrecognised crucial role in the development of depression through their activation of the mammalian target of rapamycin (mTor) pathway. The aim of this research project is to evaluate whether BCAAs are altered in patients with major depression and might thus be appropriate biomarkers for major depression. Methods The concentrations of valine, leucine and isoleucine were determined in 71 in-patients with major depression and 48 healthy controls by high-pressure liquid chromatography. Psychiatric and laboratory assessments were obtained at the time of in-patient admittance. Results The BCAAs are significantly decreased in patients with major depression in comparison with healthy subjects (valine: Mann-Whitney-U: 968.0; p <0.0001, leucine: Mann-Whitney-U: 1246.5; p = 0.013, isoleucine: Mann-Whitney-U: 1252.5; p = 0.014). Furthermore, as shown by Spearman's rank correlation coefficients, there is a significant negative correlation between valine, leucine and isoleucine concentrations and the Hamilton Depression Rating Scale (HAMD-17) as well as Beck Depression Inventory (BDI-II) scores. Conclusions Our study results are strong evidence that in patients with major depression, BCAAs might be appropriate biomarkers for depression. Reduced activation of the mammalian target of rapamycin (mTor) due to a reduction of BCAAs might play a crucial unrecognised factor in the etiology of depression and may evoke depressive symptomatology and lower energy metabolism in patients with major depression. In the future, mTor and its up- and downstream signalling partners might be important targets for the development of novel antidepressants. PMID:27490818

  19. Inactivation of the ilvB1 gene in Mycobacterium tuberculosis leads to branched-chain amino acid auxotrophy and attenuation of virulence in mice.

    Science.gov (United States)

    Awasthy, Disha; Gaonkar, Sheshagiri; Shandil, R K; Yadav, Reena; Bharath, Sowmya; Marcel, Nimi; Subbulakshmi, Venkita; Sharma, Umender

    2009-09-01

    Acetohydroxyacid synthase (AHAS) is the first enzyme in the branched-chain amino acid biosynthesis pathway in bacteria. Bioinformatics analysis revealed that the Mycobacterium tuberculosis genome contains four genes (ilvB1, ilvB2, ilvG and ilvX) coding for the large catalytic subunit of AHAS, whereas only one gene (ilvN or ilvH) coding for the smaller regulatory subunit of this enzyme was found. In order to understand the physiological role of AHAS in survival of the organism in vitro and in vivo, we inactivated the ilvB1 gene of M. tuberculosis. The mutant strain was found to be auxotrophic for all of the three branched-chain amino acids (isoleucine, leucine and valine), when grown with either C(6) or C(2) carbon sources, suggesting that the ilvB1 gene product is the major AHAS in M. tuberculosis. Depletion of these branched chain amino acids in the medium led to loss of viability of the DeltailvB1 strain in vitro, resulting in a 4-log reduction in colony-forming units after 10 days. Survival kinetics of the mutant strain cultured in macrophages maintained with sub-optimal concentrations of the branched-chain amino acids did not show any loss of viability, indicating either that the intracellular environment was rich in these amino acids or that the other AHAS catalytic subunits were functional under these conditions. Furthermore, the growth kinetics of the DeltailvB1 strain in mice indicated that although this mutant strain showed defective growth in vivo, it could persist in the infected mice for a long time, and therefore could be a potential vaccine candidate.

  20. The 1.9 A Structure of the Branched-Chain Amino-Acid Transaminase (IlvE) from Mycobacterium tuberculosis

    Energy Technology Data Exchange (ETDEWEB)

    Tremblay, L.; Blanchard, J

    2009-01-01

    Unlike mammals, bacteria encode enzymes that synthesize branched-chain amino acids. The pyridoxal 5'-phosphate-dependent transaminase performs the final biosynthetic step in these pathways, converting keto acid precursors into {alpha}-amino acids. The branched-chain amino-acid transaminase from Mycobacterium tuberculosis (MtIlvE) has been crystallized and its structure has been solved at 1.9 {angstrom} resolution. The MtIlvE monomer is composed of two domains that interact to form the active site. The biologically active form of IlvE is a homodimer in which each monomer contributes a substrate-specificity loop to the partner molecule. Additional substrate selectivity may be imparted by a conserved N-terminal Phe30 residue, which has previously been observed to shield the active site in the type IV fold homodimer. The active site of MtIlvE contains density corresponding to bound PMP, which is likely to be a consequence of the presence of tryptone in the crystallization medium. Additionally, two cysteine residues are positioned at the dimer interface for disulfide-bond formation under oxidative conditions. It is unknown whether they are involved in any regulatory activities analogous to those of the human mitochondrial branched-chain amino-acid transaminase.

  1. High affinity, bioavailable 3-amino-1,4-benzodiazepine-based gamma-secretase inhibitors.

    Science.gov (United States)

    Owens, Andrew P; Nadin, Alan; Talbot, Adam C; Clarke, Earl E; Harrison, Timothy; Lewis, Huw D; Reilly, Michael; Wrigley, Jonathan D J; Castro, José L

    2003-11-17

    In this paper, we describe the development of a novel series of high affinity, orally bioavailable 3-amino-1,4 benzodiazepine-based gamma-secretase inhibitors for the potential treatment of Alzheimer's disease. We disclose structure-activity relationships based around the 1, 3 and 5 positions of the benzodiazepine core structure.

  2. Effects of Acute Exposure to Increased Plasma Branched-Chain Amino Acid Concentrations on Insulin-Mediated Plasma Glucose Turnover in Healthy Young Subjects

    OpenAIRE

    Sarah Everman; Mandarino, Lawrence J.; Carroll, Chad C.; Katsanos, Christos S.

    2015-01-01

    Background Plasma branched-chain amino acids (BCAA) are inversely related to insulin sensitivity of glucose metabolism in humans. However, currently, it is not known whether there is a cause-and-effect relationship between increased plasma BCAA concentrations and decreased insulin sensitivity. Objective To determine the effects of acute exposure to increased plasma BCAA concentrations on insulin-mediated plasma glucose turnover in humans. Methods Ten healthy subjects were randomly assigned to...

  3. High Leucine Diets Stimulate Cerebral Branched-Chain Amino Acid Degradation and Modify Serotonin and Ketone Body Concentrations in a Pig Model

    OpenAIRE

    Wessels, Anna G.; Holger Kluge; Frank Hirche; Andreas Kiowski; Alexandra Schutkowski; Etienne Corrent; Jörg Bartelt; Bettina König; Stangl, Gabriele I.

    2016-01-01

    In addition to its role as an essential protein component, leucine (Leu) displays several other metabolic functions such as activation of protein synthesis. This property makes it an interesting amino acid for the therapy of human muscle atrophy and for livestock production. However, Leu can stimulate its own degradation via the branched-chain keto acid dehydrogenase complex (BCKDH). To examine the response of several tissues to excessive Leu, pigs were fed diets containing two- (L2) and four...

  4. Association of branched-chain amino acids with carotid intima-media thickness and coronary artery disease risk factors.

    Directory of Open Access Journals (Sweden)

    Ruiyue Yang

    Full Text Available BACKGROUND: Recent studies have determined that branched-chain (BCAAs and aromatic (AAAs amino acids are strongly correlated with obesity and atherogenic dyslipidemia and are strong predictors of diabetes. However, it is not clear if these amino acids are capable of identifying subjects with coronary artery disease (CAD, particularly with subclinical atherosclerosis who are at risk of developing CAD. METHODS: Four hundred and seventy two Chinese subjects (272 males and 200 females, 42-97 y of age undergoing physical exams were recruited at random for participation in the cross-sectional study. Serum BCAAs and AAAs were measured using our previously reported isotope dilution liquid chromatography tandem mass spectrometry method. Bilateral B-mode carotid artery images for carotid intima-media thickness (cIMT were acquired at end diastole and cIMT values more than 0.9 mm were categorized as increased. Correlations of BCAAs with cIMT and other CAD risk factors were analyzed. RESULTS: BCAAs and AAAs were significantly and positively associated with risk factors of CAD, e.g., cIMT, BMI, waist circumference, blood pressure, fasting blood glucose, TG, apoB, apoB/apoAI ratio, apoCII, apoCIII and hsCRP, and were significantly and negatively associated with HDL-C and apoAI. Stepwise multiple linear regression analysis revealed that age (β = 0.175, P<0.001, log BCAA (β = 0.147, P<0.001 and systolic blood pressure (β = 0.141, P = 0.012 were positively and independently associated with cIMT. In the logistic regression model, the most and only powerful laboratory factor correlated with increased cIMT was BCAA (the odds ratio of the fourth quartile compared to the first quartile was 2.679; P = 0.009. CONCLUSION: BCAAs are independently correlated with increased cIMT. This correlation would open a new field of research in the mechanistic understanding and risk assessment of CAD.

  5. Branched chain enriched amino acid versus glucose treatment of hepatic encephalopathy. A double-blind study of 65 patients with cirrhosis

    DEFF Research Database (Denmark)

    Vilstrup, Hendrik; Gluud, C; Hardt, F;

    1990-01-01

    . In the glucose group ten died, three developed renal and two respiratory failure, and one remained encephalopathic. The coma score worsened in three of the patients who died in the amino acid group, but in all patients who died in the glucose group. The negative nitrogen balance on entry reversed in the amino...... acid group, but not in the glucose group. Thus, the branched chain enriched amino acid supplement did not change the prognosis for wake-up, but had other effects on the cerebral state and on nitrogen homeostasis....

  6. Supplemental branched-chain amino acids improve performance and immune response of newly-received feedlot calves

    Science.gov (United States)

    Supplemental branched-chain AA (BCAA) improved N balance of steers during a simulated pathogen challenge. The objective of this study was to determine the effect of supplemental BCAA on growth and health of newly-received feedlot steers. Steers (n = 120; initial BW = 376 ± 5 kg) were blocked by BW a...

  7. Activation of mTORC1 by leucine is potentiated by branched-chain amino acids and even more so by essential amino acids following resistance exercise

    DEFF Research Database (Denmark)

    Moberg, Marcus; Apró, William; Ekblom, Björn

    2016-01-01

    of the branched-chain amino acids (BCAA) to this effect is yet unknown. Here we compare the stimulatory role of leucine, BCAA, and EAA ingestion on anabolic signaling following exercise. Accordingly, eight trained volunteers completed four sessions of resistance exercise during which they ingested either placebo......, leucine, BCAA, or EAA (including the BCAA) in random order. Muscle biopsies were taken at rest, immediately after exercise, and following 90 and 180 min of recovery. Following 90 min of recovery the activity of S6 kinase 1 (S6K1) was greater than at rest in all four trials (PlaceboBCAA...6K1, being 18% higher with EAA than BCAA. However, after 180 min of recovery this difference between EAA and BCAA had disappeared, although with both these supplements the increases were still higher than with leucine (40%, P

  8. Regulation of adipose branched-chain amino acid catabolism enzyme expression and cross-adipose amino acid flux in human obesity.

    Science.gov (United States)

    Lackey, Denise E; Lynch, Christopher J; Olson, Kristine C; Mostaedi, Rouzbeh; Ali, Mohamed; Smith, William H; Karpe, Fredrik; Humphreys, Sandy; Bedinger, Daniel H; Dunn, Tamara N; Thomas, Anthony P; Oort, Pieter J; Kieffer, Dorothy A; Amin, Rajesh; Bettaieb, Ahmed; Haj, Fawaz G; Permana, Paska; Anthony, Tracy G; Adams, Sean H

    2013-06-01

    Elevated blood branched-chain amino acids (BCAA) are often associated with insulin resistance and type 2 diabetes, which might result from a reduced cellular utilization and/or incomplete BCAA oxidation. White adipose tissue (WAT) has become appreciated as a potential player in whole body BCAA metabolism. We tested if expression of the mitochondrial BCAA oxidation checkpoint, branched-chain α-ketoacid dehydrogenase (BCKD) complex, is reduced in obese WAT and regulated by metabolic signals. WAT BCKD protein (E1α subunit) was significantly reduced by 35-50% in various obesity models (fa/fa rats, db/db mice, diet-induced obese mice), and BCKD component transcripts significantly lower in subcutaneous (SC) adipocytes from obese vs. lean Pima Indians. Treatment of 3T3-L1 adipocytes or mice with peroxisome proliferator-activated receptor-γ agonists increased WAT BCAA catabolism enzyme mRNAs, whereas the nonmetabolizable glucose analog 2-deoxy-d-glucose had the opposite effect. The results support the hypothesis that suboptimal insulin action and/or perturbed metabolic signals in WAT, as would be seen with insulin resistance/type 2 diabetes, could impair WAT BCAA utilization. However, cross-tissue flux studies comparing lean vs. insulin-sensitive or insulin-resistant obese subjects revealed an unexpected negligible uptake of BCAA from human abdominal SC WAT. This suggests that SC WAT may not be an important contributor to blood BCAA phenotypes associated with insulin resistance in the overnight-fasted state. mRNA abundances for BCAA catabolic enzymes were markedly reduced in omental (but not SC) WAT of obese persons with metabolic syndrome compared with weight-matched healthy obese subjects, raising the possibility that visceral WAT contributes to the BCAA metabolic phenotype of metabolically compromised individuals.

  9. Activation of mTORC1 by leucine is potentiated by branched-chain amino acids and even more so by essential amino acids following resistance exercise.

    Science.gov (United States)

    Moberg, Marcus; Apró, William; Ekblom, Björn; van Hall, Gerrit; Holmberg, Hans-Christer; Blomstrand, Eva

    2016-06-01

    Protein synthesis is stimulated by resistance exercise and intake of amino acids, in particular leucine. Moreover, activation of mammalian target of rapamycin complex 1 (mTORC1) signaling by leucine is potentiated by the presence of other essential amino acids (EAA). However, the contribution of the branched-chain amino acids (BCAA) to this effect is yet unknown. Here we compare the stimulatory role of leucine, BCAA, and EAA ingestion on anabolic signaling following exercise. Accordingly, eight trained volunteers completed four sessions of resistance exercise during which they ingested either placebo, leucine, BCAA, or EAA (including the BCAA) in random order. Muscle biopsies were taken at rest, immediately after exercise, and following 90 and 180 min of recovery. Following 90 min of recovery the activity of S6 kinase 1 (S6K1) was greater than at rest in all four trials (PlaceboBCAABCAA. However, after 180 min of recovery this difference between EAA and BCAA had disappeared, although with both these supplements the increases were still higher than with leucine (40%, P BCAA.

  10. Arabidopsis and Maize RidA Proteins Preempt Reactive Enamine/Imine Damage to Branched-Chain Amino Acid Biosynthesis in Plastids[C][W][OPEN

    Science.gov (United States)

    Niehaus, Thomas D.; Nguyen, Thuy N.D.; Gidda, Satinder K.; ElBadawi-Sidhu, Mona; Lambrecht, Jennifer A.; McCarty, Donald R.; Downs, Diana M.; Cooper, Arthur J.L.; Fiehn, Oliver; Mullen, Robert T.; Hanson, Andrew D.

    2014-01-01

    RidA (for Reactive Intermediate Deaminase A) proteins are ubiquitous, yet their function in eukaryotes is unclear. It is known that deleting Salmonella enterica ridA causes Ser sensitivity and that S. enterica RidA and its homologs from other organisms hydrolyze the enamine/imine intermediates that Thr dehydratase forms from Ser or Thr. In S. enterica, the Ser-derived enamine/imine inactivates a branched-chain aminotransferase; RidA prevents this damage. Arabidopsis thaliana and maize (Zea mays) have a RidA homolog that is predicted to be plastidial. Expression of either homolog complemented the Ser sensitivity of the S. enterica ridA mutant. The purified proteins hydrolyzed the enamines/imines formed by Thr dehydratase from Ser or Thr and protected the Arabidopsis plastidial branched-chain aminotransferase BCAT3 from inactivation by the Ser-derived enamine/imine. In vitro chloroplast import assays and in vivo localization of green fluorescent protein fusions showed that Arabidopsis RidA and Thr dehydratase are chloroplast targeted. Disrupting Arabidopsis RidA reduced root growth and raised the root and shoot levels of the branched-chain amino acid biosynthesis intermediate 2-oxobutanoate; Ser treatment exacerbated these effects in roots. Supplying Ile reversed the root growth defect. These results indicate that plastidial RidA proteins can preempt damage to BCAT3 and Ile biosynthesis by hydrolyzing the Ser-derived enamine/imine product of Thr dehydratase. PMID:25070638

  11. Lrp of Corynebacterium glutamicum controls expression of the brnFE operon encoding the export system for L-methionine and branched-chain amino acids.

    Science.gov (United States)

    Lange, Christian; Mustafi, Nurije; Frunzke, Julia; Kennerknecht, Nicole; Wessel, Mirja; Bott, Michael; Wendisch, Volker F

    2012-04-30

    Corynebacterium glutamicum possesses export systems for various amino acids including BrnFE, a two-component export system for L-methionine and the branched-chain amino acids L-valine, L-isoleucine and L-leucine. A gene for a putative transcriptional regulator of the Lrp family is transcribed divergently to the brnFE operon and is required for L-isoleucine export. By comparing global gene expression changes due to L-isoleucine addition we revealed increased brnFE expression in response to L-isoleucine in C. glutamicum wild type but not in an lrp deletion mutant. ChIP-to-chip analysis, band shift experiments and DNAse footprint analysis demonstrated that Lrp binds to the intergenic region between lrp and brnF. Expression analysis of transcriptional fusions with the lrp and brnFE promoters indicated that branched-chain amino acids and L-methionine when added to the growth medium stimulated brnFE expression in the order L-leucine > L-methionine > L-isoleucine > L-valine and that Lrp was required for activation of brnFE expression. Thus, regulation of brnFE by Lrp ensures that BrnFE is synthesized only if its substrate amino acids accumulate in cells which is commensurate with its role to counteract such situations of metabolic imbalance.

  12. Efficient synthesis of D-branched-chain amino acids and their labeled compounds with stable isotopes using D-amino acid dehydrogenase.

    Science.gov (United States)

    Akita, Hironaga; Suzuki, Hirokazu; Doi, Katsumi; Ohshima, Toshihisa

    2014-02-01

    D-Branched-chain amino acids (D-BCAAs) such as D-leucine, D-isoleucine, and D-valine are known to be peptide antibiotic intermediates and to exhibit a variety of bioactivities. Consequently, much effort is going into achieving simple stereospecific synthesis of D-BCAAs, especially analogs labeled with stable isotopes. Up to now, however, no effective method has been reported. Here, we report the establishment of an efficient system for enantioselective synthesis of D-BCAAs and production of D-BCAAs labeled with stable isotopes. This system is based on two thermostable enzymes: D-amino acid dehydrogenase, catalyzing NADPH-dependent enantioselective amination of 2-oxo acids to produce the corresponding D-amino acids, and glucose dehydrogenase, catalyzing NADPH regeneration from NADP(+) and D-glucose. After incubation with the enzymes for 2 h at 65°C and pH 10.5, 2-oxo-4-methylvaleric acid was converted to D-leucine with an excellent yield (>99 %) and optical purity (>99 %). Using this system, we produced five different D-BCAAs labeled with stable isotopes: D-[1-(13)C,(15)N]leucine, D-[1-(13)C]leucine, D-[(15)N]leucine, D-[(15)N]isoleucine, and D-[(15)N]valine. The structure of each labeled D-amino acid was confirmed using time-of-flight mass spectrometry and nuclear magnetic resonance analysis. These analyses confirmed that the developed system was highly useful for production of D-BCAAs labeled with stable isotopes, making this the first reported enzymatic production of D-BCAAs labeled with stable isotopes. Our findings facilitate tracer studies investigating D-BCAAs and their derivatives.

  13. Prevention of methionine and ammonia-induced coma by intravenous infusion of a branched chain amino acid solution to rats with liver injury.

    Directory of Open Access Journals (Sweden)

    Shiota,Tetsuya

    1984-10-01

    Full Text Available The prevention of hepatic encephalopathy by the intravenous infusion of a branched chain amino acid (BCAA-enriched solution was investigated in methionine and ammonium acetate-treated rats whose liver was already injured with carbon tetrachloride. A BCAA-enriched solution protected the rats from entering a coma. The brain BCAA contents became higher, and the brain methionine and tyrosine levels and the ratio of glutamine to glutamic acid in the brain diminished after administering the BCAA-enriched solution.

  14. A Branched-Chain Amino Acid-Related Metabolic Signature that Differentiates Obese and Lean Humans and Contributes to Insulin Resistance

    OpenAIRE

    Newgard, Christopher B.; An, Jie; Bain, James R.; Muehlbauer, Michael J.; Robert D. Stevens; Lien, Lillian F.; Haqq, Andrea M.; Shah, Svati H.; Arlotto, Michelle; Slentz, Cris A.; Rochon, James; Gallup, Dianne; Ilkayeva, Olga; Brett R. Wenner; Yancy, William E

    2009-01-01

    Metabolomic profiling of obese versus lean humans reveals a branched-chain amino acid (BCAA)-related metabolite signature that is suggestive of increased catabolism of BCAA and correlated with insulin resistance. To test its impact on metabolic homeostasis, we fed rats on high-fat (HF), HF with supplemented BCAA (HF/BCAA) or standard chow (SC) diets. Despite having reduced food intake and weight gain equivalent to the SC group, HF/BCAA rats were equally insulin resistant as HF rats. Pair-feed...

  15. Effects of Low-Protein Diets Supplemented with Branched-Chain Amino Acid on Lipid Metabolism in White Adipose Tissue of Piglets.

    Science.gov (United States)

    Li, Yinghui; Wei, Hongkui; Li, Fengna; Duan, Yehui; Guo, Qiuping; Yin, Yulong

    2017-04-05

    This study evaluated the effect of branched-chain amino acid (BCAA) supplementation in low-protein diets on lipid metabolism in dorsal subcutaneous adipose (DSA), abdominal subcutaneous adipose (ASA), and perirenal adipose (PRA) tissues. A total of 24 piglets were allotted to four treatments, and each group was fed the adequate protein (AP) diet, low-protein (LP) diet, LP diet supplemented with BCAA (LP + B), or LP diet supplemented with twice BCAA (LP + 2B). Serum concentrations of leptin in the BCAA-supplemented treatments were higher (P effect was likely modulated by the expression of lipid metabolic regulators in DSA, ASA, and PRA in a depot-specific manner.

  16. Effects of leucine supplementation and serum withdrawal on branched-chain amino acid pathway gene and protein expression in mouse adipocytes.

    Directory of Open Access Journals (Sweden)

    Abderrazak Kitsy

    Full Text Available The essential branched-chain amino acids (BCAA, leucine, valine and isoleucine, are traditionally associated with skeletal muscle growth and maintenance, energy production, and generation of neurotransmitter and gluconeogenic precursors. Recent evidence from human and animal model studies has established an additional link between BCAA levels and obesity. However, details of the mechanism of regulation of BCAA metabolism during adipogenesis are largely unknown. We interrogated whether the expression of genes and proteins involved in BCAA metabolism are sensitive to the adipocyte differentiation process, and responsive to nutrient stress from starvation or BCAA excess. Murine 3T3-L1 preadipocytes were differentiated to adipocytes under control conditions and under conditions of L-leucine supplementation or serum withdrawal. RNA and proteins were isolated at days 0, 4 and 10 of differentiation to represent pre-differentiation, early differentiation and late differentiation stages. Expression of 16 BCAA metabolism genes was quantified by quantitative real-time PCR. Expression of the protein levels of branched-chain amino acid transaminase 2 (Bcat2 and branched-chain alpha keto acid dehydrogenase (Bckdha was quantified by immunoblotting. Under control conditions, all genes displayed induction of gene expression during early adipogenesis (Day 4 compared to Day 0. Leucine supplementation resulted in an induction of Bcat2 and Bckdha genes during early and late differentiation. Western blot analysis demonstrated condition-specific concordance between gene and protein expression. Serum withdrawal resulted in undetectable Bcat2 and Bckdha protein levels at all timepoints. These results demonstrate that the expression of genes related to BCAA metabolism are regulated during adipocyte differentiation and influenced by nutrient levels. These results provide additional insights on how BCAA metabolism is associated with adipose tissue function and extends our

  17. Metabolic Fate of Branched-Chain Amino Acids During Adipogenesis, in Adipocytes From Obese Mice and C2C12 Myotubes.

    Science.gov (United States)

    Estrada-Alcalde, Isabela; Tenorio-Guzman, Miriam R; Tovar, Armando R; Salinas-Rubio, Daniela; Torre-Villalvazo, Ivan; Torres, Nimbe; Noriega, Lilia G

    2017-04-01

    Branched-chain amino acid (BCAA) catabolism is regulated by the branched-chain aminotransferase (BCAT2) and the branched-chain α-keto acid dehydrogenase complex (BCKDH). BCAT2 and BCKDH expression and activity are modified during adipogenesis and altered in adipose tissues of mice with genetic or diet-induced obesity. However, little is known about how these modifications and alterations affect the intracellular metabolic fate of BCAAs during adipogenesis, in adipocytes from mice fed a control or high-fat diet or in C2C12 myotubes. Here, we demonstrate that BCAAs are mainly incorporated into proteins during the early stages of adipocyte differentiation. However, they are oxidized and incorporated into lipids during the late days of differentiation. Conversely, 92% and 97% of BCAA were oxidized, 1.6% and 6% were used for protein synthesis and 1.2% and 1.5% were incorporated into lipids in adipocytes from epididymal and subcutaneous adipose tissue, respectively. All three pathways were decreased in adipocytes from mice fed a high-fat diet. In C2C12 myotubes, leucine is mainly used for protein synthesis and palmitate is incorporated into lipids. Interestingly, leucine decreased both palmitate oxidation and its incorporation to lipids and proteins; and palmitate increased leucine oxidation and decreased its incorporation to lipids and proteins in a dose-dependent manner. These results demonstrate that BCAA metabolic fate differs between the early and late stages of adipocyte differentiation and in adipocytes from mice fed a control or high-fat diet; and that leucine affects the metabolic fate of palmitate and vice versa in C2C12 myotubes. J. Cell. Biochem. 118: 808-818, 2017. © 2016 Wiley Periodicals, Inc.

  18. Arabidopsis and maize RidA proteins preempt reactive enamine/imine damage to branched-chain amino acid biosynthesis in plastids.

    Science.gov (United States)

    Niehaus, Thomas D; Nguyen, Thuy N D; Gidda, Satinder K; ElBadawi-Sidhu, Mona; Lambrecht, Jennifer A; McCarty, Donald R; Downs, Diana M; Cooper, Arthur J L; Fiehn, Oliver; Mullen, Robert T; Hanson, Andrew D

    2014-07-01

    RidA (for Reactive Intermediate Deaminase A) proteins are ubiquitous, yet their function in eukaryotes is unclear. It is known that deleting Salmonella enterica ridA causes Ser sensitivity and that S. enterica RidA and its homologs from other organisms hydrolyze the enamine/imine intermediates that Thr dehydratase forms from Ser or Thr. In S. enterica, the Ser-derived enamine/imine inactivates a branched-chain aminotransferase; RidA prevents this damage. Arabidopsis thaliana and maize (Zea mays) have a RidA homolog that is predicted to be plastidial. Expression of either homolog complemented the Ser sensitivity of the S. enterica ridA mutant. The purified proteins hydrolyzed the enamines/imines formed by Thr dehydratase from Ser or Thr and protected the Arabidopsis plastidial branched-chain aminotransferase BCAT3 from inactivation by the Ser-derived enamine/imine. In vitro chloroplast import assays and in vivo localization of green fluorescent protein fusions showed that Arabidopsis RidA and Thr dehydratase are chloroplast targeted. Disrupting Arabidopsis RidA reduced root growth and raised the root and shoot levels of the branched-chain amino acid biosynthesis intermediate 2-oxobutanoate; Ser treatment exacerbated these effects in roots. Supplying Ile reversed the root growth defect. These results indicate that plastidial RidA proteins can preempt damage to BCAT3 and Ile biosynthesis by hydrolyzing the Ser-derived enamine/imine product of Thr dehydratase. © 2014 American Society of Plant Biologists. All rights reserved.

  19. Suppression of Aflatoxin Biosynthesis in Aspergillus flavus by 2-Phenylethanol Is Associated with Stimulated Growth and Decreased Degradation of Branched-Chain Amino Acids

    Directory of Open Access Journals (Sweden)

    Perng-Kuang Chang

    2015-09-01

    Full Text Available The saprophytic soil fungus Aspergillus flavus infects crops and produces aflatoxin. Pichia anomala, which is a biocontrol yeast and produces the major volatile 2-phenylethanol (2-PE, is able to reduce growth of A. flavus and aflatoxin production when applied onto pistachio trees. High levels of 2-PE are lethal to A. flavus and other fungi. However, at low levels, the underlying mechanism of 2-PE to inhibit aflatoxin production remains unclear. In this study, we characterized the temporal transcriptome response of A. flavus to 2-PE at a subinhibitory level (1 μL/mL using RNA-Seq technology and bioinformatics tools. The treatment during the entire 72 h experimental period resulted in 131 of the total A. flavus 13,485 genes to be significantly impacted, of which 82 genes exhibited decreased expression. They included those encoding conidiation proteins and involved in cyclopiazonic acid biosynthesis. All genes in the aflatoxin gene cluster were also significantly decreased during the first 48 h treatment. Gene Ontology (GO analyses showed that biological processes with GO terms related to catabolism of propionate and branched-chain amino acids (valine, leucine and isoleucine were significantly enriched in the down-regulated gene group, while those associated with ribosome biogenesis, translation, and biosynthesis of α-amino acids were over-represented among the up-regulated genes. Kyoto Encyclopedia of Genes and Genomes (KEGG pathway analysis revealed that metabolic pathways negatively impacted among the down-regulated genes parallel to those active at 30 °C, a condition conducive to aflatoxin biosynthesis. In contrast, metabolic pathways positively related to the up-regulated gene group resembled those at 37 °C, which favors rapid fungal growth and is inhibitory to aflatoxin biosynthesis. The results showed that 2-PE at a low level stimulated active growth of A. flavus but concomitantly rendered decreased activities in branched-chain amino

  20. Suppression of Aflatoxin Biosynthesis in Aspergillus flavus by 2-Phenylethanol Is Associated with Stimulated Growth and Decreased Degradation of Branched-Chain Amino Acids.

    Science.gov (United States)

    Chang, Perng-Kuang; Hua, Sui Sheng T; Sarreal, Siov Bouy L; Li, Robert W

    2015-09-24

    The saprophytic soil fungus Aspergillus flavus infects crops and produces aflatoxin. Pichia anomala, which is a biocontrol yeast and produces the major volatile 2-phenylethanol (2-PE), is able to reduce growth of A. flavus and aflatoxin production when applied onto pistachio trees. High levels of 2-PE are lethal to A. flavus and other fungi. However, at low levels, the underlying mechanism of 2-PE to inhibit aflatoxin production remains unclear. In this study, we characterized the temporal transcriptome response of A. flavus to 2-PE at a subinhibitory level (1 μL/mL) using RNA-Seq technology and bioinformatics tools. The treatment during the entire 72 h experimental period resulted in 131 of the total A. flavus 13,485 genes to be significantly impacted, of which 82 genes exhibited decreased expression. They included those encoding conidiation proteins and involved in cyclopiazonic acid biosynthesis. All genes in the aflatoxin gene cluster were also significantly decreased during the first 48 h treatment. Gene Ontology (GO) analyses showed that biological processes with GO terms related to catabolism of propionate and branched-chain amino acids (valine, leucine and isoleucine) were significantly enriched in the down-regulated gene group, while those associated with ribosome biogenesis, translation, and biosynthesis of α-amino acids OPEN ACCESS Toxins 2015, 7 3888 were over-represented among the up-regulated genes. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed that metabolic pathways negatively impacted among the down-regulated genes parallel to those active at 30 °C, a condition conducive to aflatoxin biosynthesis. In contrast, metabolic pathways positively related to the up-regulated gene group resembled those at 37 °C, which favors rapid fungal growth and is inhibitory to aflatoxin biosynthesis. The results showed that 2-PE at a low level stimulated active growth of A. flavus but concomitantly rendered decreased activities in

  1. Supplementation with branched-chain amino acids to a low-protein diet regulates intestinal expression of amino acid and peptide transporters in weanling pigs.

    Science.gov (United States)

    Zhang, Shihai; Qiao, Shiyan; Ren, Man; Zeng, Xiangfang; Ma, Xi; Wu, Zhenlong; Thacker, Philip; Wu, Guoyao

    2013-11-01

    This study determined the effects of dietary branched-chain amino acids (AA) (BCAA) on growth performance, expression of jejunal AA and peptide transporters, and the colonic microflora of weanling piglets fed a low-protein (LP) diet. One hundred and eight Large White × Landrace × Duroc piglets (weaned at 28 days of age) were fed a normal protein diet (NP, 20.9 % crude protein), an LP diet (LP, 17.1 % crude protein), or an LP diet supplemented with BCAA (LP + BCAA, 17.9 % crude protein) for 14 days. Dietary protein restriction reduced piglet growth performance and small-intestinal villous height, which were restored by BCAA supplementation to the LP diet to values for the NP diet. Serum concentrations of BCAA were reduced in piglets fed the LP diet while those in piglets fed the LP + BCAA diet were similar to values for the NP group. mRNA levels for Na(+)-neutral AA exchanger-2, cationic AA transporter-1, b(0,+) AA transporter, and 4F2 heavy chain were more abundant in piglets fed the LP + BCAA diet than the LP diet. However, mRNA and protein levels for peptide transporter-1 were lower in piglets fed the LP + BCAA diet as compared to the LP diet. The colonic microflora did not differ among the three groups of pigs. In conclusion, growth performance, intestinal development, and intestinal expression of AA transporters in weanling piglets are enhanced by BCAA supplementation to LP diets. Our findings provide a new molecular basis for further understanding of BCAA as functional AA in animal nutrition.

  2. Rapid and precise measurement of serum branched-chain and aromatic amino acids by isotope dilution liquid chromatography tandem mass spectrometry.

    Directory of Open Access Journals (Sweden)

    Ruiyue Yang

    Full Text Available BACKGROUND: Serum branched-chain and aromatic amino acids (BCAAs and AAAs have emerged as predictors for the future development of diabetes and may aid in diabetes risk assessment. However, the current methods for the analysis of such amino acids in biological samples are time consuming. METHODS: An isotope dilution liquid chromatography tandem mass spectrometry (ID-LC/MS/MS method for serum BCAAs and AAAs was developed. The serum was mixed with isotope-labeled BCAA and AAA internal standards and the amino acids were extracted with acetonitrile, followed by analysis using LC/MS/MS. The LC separation was performed on a reversed-phase C18 column, and the MS/MS detection was performed via the positive electronic spray ionization in multiple reaction monitoring mode. RESULTS: Specific analysis of the amino acids was achieved within 2 min. Intra-run and total CVs for the amino acids were less than 2% and 4%, respectively, and the analytical recoveries ranged from 99.6 to 103.6%. CONCLUSION: A rapid and precise method for the measurement of serum BCAAs and AAAs was developed and may serve as a quick tool for screening serum BCAAs and AAAs in studies assessing diabetes risk.

  3. Nontargeted LC–MS Metabolomics Approach for Metabolic Profiling of Plasma and Urine from Pigs Fed Branched Chain Amino Acids for Maximum Growth Performance

    DEFF Research Database (Denmark)

    Assadi Soumeh, Elham; Hedemann, Mette Skou; Poulsen, Hanne Damgaard

    2016-01-01

    The metabolic response in plasma and urine of pigs when feeding an optimum level of branched chain amino acids (BCAAs) for best growth performance is unknown. The objective of the current study was to identify the metabolic phenotype associated with the BCAAs intake level that could be linked...... to the animal growth performance. Three dose–response studies were carried out to collect blood and urine samples from pigs fed increasing levels of Ile, Val, or Leu followed by a nontargeted LC–MS approach to characterize the metabolic profile of biofluids when dietary BCAAs are optimum for animal growth....... Results showed that concentrations of plasma hypoxanthine and tyrosine (Tyr) were higher while concentrations of glycocholic acid, tauroursodeoxycholic acid, and taurocholic acid were lower when the dietary Ile was optimum. Plasma 3-methyl-2-oxovaleric acid and creatine were lower when dietary Leu...

  4. Branched-chain amino acids increase arterial blood ammonia in spite of enhanced intrinsic muscle ammonia metabolism in patients with cirrhosis and healthy subjects

    DEFF Research Database (Denmark)

    Dam, Gitte; Keiding, Susanne; Munk, Ole Lajord

    2011-01-01

    Branched-chain amino acids (BCAA) are used in attempts to reduce blood ammonia in patients with cirrhosis and intermittent hepatic encephalopathy based on the hypothesis that BCAA stimulate muscle ammonia detoxification. We studied the effects of an oral dose of BCAA on the skeletal muscle...... the metabolism of blood-supplied ammonia and the A-V measurements were used to measure the total ammonia metabolism across the thigh muscle. After intake of BCAA, blood ammonia increased more than 30% in both groups of subjects (both P ....05). BCAA intake led to a massive glutamine release from the muscle (cirrhotic patients, P BCAA enhanced the intrinsic muscle metabolism of ammonia but not the metabolism of blood-supplied ammonia in both the patients with cirrhosis and in the healthy...

  5. [Usefulness of branched-chain amino acid (BCAA)-enriched nutrient mixture for nutritional treatment undergoing endoscopic treatment for esophageal varices].

    Science.gov (United States)

    Shibata, Naozumi; Matsui, Hidetaka; Takeshita, Eiji; Yokota, Tomoyuki; Higaki, Naoyuki; Murakami, Hidehiro; Ikeda, Yoshiou; Minami, Hisaka; Matsuura, Bunzo; Onji, Morikazu

    2005-07-01

    We investigated the alteration of nutritional status in 144 patients who were treated for the first time with endoscopic sclerotherapy or endoscopic variceal ligation during their therapies. The serum levels of albumin, cholinesterase and total cholesterol were compared before and after treatment. The serum level of cholinesterase declined significantly. To investigate the impact of aging on the changes of nutritional status we divided all patients into two groups: (1) under 65 years, and (2) over 65 years. The decline of serum albumin of elderly patients (n=65) was significantly greater than that of younger patients (n=79). A branched-chain amino acid (BCAA)-enriched nutrient mixture for nutritional treatment significantly suppressed the decline of serum albumin in elderly patients. Nutritional treatment with a BCAA-enriched nutrient mixture should be considered during endoscopic therapy for esophageal varices, especially in elderly patients.

  6. Insulin does not stimulate muscle protein synthesis during increased plasma branched-chain amino acids alone but still decreases whole body proteolysis in humans.

    Science.gov (United States)

    Everman, Sarah; Meyer, Christian; Tran, Lee; Hoffman, Nyssa; Carroll, Chad C; Dedmon, William L; Katsanos, Christos S

    2016-10-01

    Insulin stimulates muscle protein synthesis when the levels of total amino acids, or at least the essential amino acids, are at or above their postabsorptive concentrations. Among the essential amino acids, branched-chain amino acids (BCAA) have the primary role in stimulating muscle protein synthesis and are commonly sought alone to stimulate muscle protein synthesis in humans. Fourteen healthy young subjects were studied before and after insulin infusion to examine whether insulin stimulates muscle protein synthesis in relation to the availability of BCAA alone. One half of the subjects were studied in the presence of postabsorptive BCAA concentrations (control) and the other half in the presence of increased plasma BCAA (BCAA). Compared with that prior to the initiation of the insulin infusion, fractional synthesis rate of muscle protein (%/h) did not change (P > 0.05) during insulin in either the control (0.04 ± 0.01 vs 0.05 ± 0.01) or the BCAA (0.05 ± 0.02 vs. 0.05 ± 0.01) experiments. Insulin decreased (P BCAA (0.89 ± 0.07 vs 0.61 ± 0.03) experiments, but the change was not different between the two experiments (P > 0.05). In conclusion, insulin does not stimulate muscle protein synthesis in the presence of increased circulating levels of plasma BCAA alone. Insulin's suppressive effect on proteolysis is observed independently of the levels of circulating plasma BCAA.

  7. The development and application of a single-cell biosensor for the detection of l-methionine and branched-chain amino acids.

    Science.gov (United States)

    Mustafi, Nurije; Grünberger, Alexander; Kohlheyer, Dietrich; Bott, Michael; Frunzke, Julia

    2012-07-01

    The detection and quantification of specific metabolites in single bacterial cells is a major goal for industrial biotechnology. We have developed a biosensor based on the transcriptional regulator Lrp that detects intracellular l-methionine and branched-chain amino acids in Corynebacterium glutamicum. In assays, fluorescence output showed a linear relationship with cytoplasmic concentrations of the effector amino acids. In increasing order, the affinity of Lrp for the amino acids is l-valine, l-isoleucine, l-leucine and l-methionine. The sensor was applied for online monitoring and analysis of cell-to-cell variability of l-valine production by the pyruvate dehydrogenase-deficient C. glutamicum strain ΔaceE. Finally, the sensor system was successfully used in a high-throughput (HT) FACS screen for the isolation of amino acid-producing mutants after random mutagenesis of a non-producing wild type strain. These applications illustrate how one of nature's sensor devices - transcriptional regulators - can be used for the analysis, directed evolution and HT screening for microbial strain development.

  8. Targeted metabolomic analysis reveals the association between the postprandial change in palmitic acid, branched-chain amino acids and insulin resistance in young obese subjects.

    Science.gov (United States)

    Liu, Liyan; Feng, Rennan; Guo, Fuchuan; Li, Ying; Jiao, Jundong; Sun, Changhao

    2015-04-01

    Obesity is the result of a positive energy balance and often leads to difficulties in maintaining normal postprandial metabolism. The changes in postprandial metabolites after an oral glucose tolerance test (OGTT) in young obese Chinese men are unclear. In this work, the aim is to investigate the complex metabolic alterations in obesity provoked by an OGTT using targeted metabolomics. We used gas chromatography-mass spectrometry and ultra high performance liquid chromatography-triple quadrupole mass spectrometry to analyze serum fatty acids, amino acids and biogenic amines profiles from 15 control and 15 obese subjects at 0, 30, 60, 90 and 120 min during an OGTT. Metabolite profiles from 30 obese subjects as independent samples were detected in order to validate the change of metabolites. There were the decreased levels of fatty acid, amino acids and biogenic amines after OGTT in obesity. At 120 min, percent change of 20 metabolites in obesity has statistical significance when comparing with the controls. The obese parameters was positively associated with changes in arginine and histidine (Pchange in palmitic acid (PA), branched-chain amino acids (BCAAs) and phenylalanine between 1 and 120 min were positively associated with fasting insulin and HOMA-IR (all Presistance in obesity. Our findings offer new insights in the complex physiological regulation of the metabolism during an OGTT in obesity. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  9. Branched-chain amino acid requirements for enterally fed term neonates in the first month of life

    NARCIS (Netherlands)

    F. Maingay-de Groof (Femke); L. Huang; I. van Vliet (Ineke); G.J. Voortman (Gardi); H. Schierbeek (Henk); L.C.W. Roksnoer (Lodi); A. Vermes (Andras); C. Chen (Chao); Y. Huang (Ying); J.B. van Goudoever (Hans)

    2014-01-01

    textabstractBackground: Knowledge of essential amino acid requirements in infants is important because excessive intake of protein can lead to increased long-term morbidity such as obesity. A deficient intake may lead to suboptimal growth and impaired neurodevelopment. The current recommended branch

  10. Effect of a branched chain amino acid-enriched nutritional product on the pathophysiology of the liver and nutritional state of patients with liver cirrhosis.

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    Watanabe,Akiharu

    1983-08-01

    Full Text Available A new nutritional product (SF-1008C containing a high proportion of branched chain amino acids (BCAA and low proportion of aromatic amino acids (AAA and methionine was tested to see its effect on the impaired protein metabolism and abnormal nutritional state frequently observed in patients with advanced liver cirrhosis. A sharp increase in plasma BCAA levels and fall of AAA and methionine levels were found following the administration of an SF-1008C-supplemented diet to healthy controls and cirrhotic patients, which the BCAA levels increased only slightly following an isocaloric control diet. Blood ammonia levels increased within the normal range transiently following the diets. The SF-1008C-supplemented diet was given for 2 weeks to cirrhotic patients with histories of hepatic encephalopathy, who were taking a low-protein diet because of hyperammonemia. Serum prealbumin levels, nitrogen balance, molar ratio of plasma BCAA/phenylalanine and tyrosine, the number connection test and electroencephalograms improved during the period of the experimental diet. The results, therefore, indicate that a BCAA-supplemented diet is well tolerated by patients with advanced cirrhosis and useful for treatment of impaired protein metabolism. Furthermore, this product is beneficial in preventing hepatic encephalopathy in cirrhotics.

  11. Branched-chain Amino Acids are Beneficial to Maintain Growth Performance and Intestinal Immune-related Function in Weaned Piglets Fed Protein Restricted Diet.

    Science.gov (United States)

    Ren, M; Zhang, S H; Zeng, X F; Liu, H; Qiao, S Y

    2015-12-01

    As a novel approach for disease control and prevention, nutritional modulation of the intestinal health has been proved. However, It is still unknown whether branched-chain amino acid (BCAA) is needed to maintain intestinal immune-related function. The objective of this study was to determine whether BCAA supplementation in protein restricted diet affects growth performance, intestinal barrier function and modulates post-weaning gut disorders. One hundred and eight weaned piglets (7.96±0.26 kg) were randomly fed one of the three diets including a control diet (21% crude protein [CP], CON), a protein restricted diet (17% CP, PR) and a BCAA diet (BCAA supplementation in the PR diet) for 14 d. The growth performance, plasma amino acid concentrations, small intestinal morphology and intestinal immunoglobulins were tested. First, average daily gain (ADG) (pBCAA group improved ADG (pBCAA groups was not different (p>0.05). The PR and BCAA treatments had a higher (pBCAA supplementation significantly increased BCAA concentrations (pBCAA supplementation increased villous height in the duodenum (pBCAA supplementation increased levels of jejunal and ileal immunoglobulin mentioned above. In conclusion, BCAA supplementation to protein restricted diet improved intestinal immune defense function by protecting villous morphology and by increasing levels of intestinal immunoglobulins in weaned piglets. Our finding has the important implication that BCAA may be used to reduce the negative effects of a protein restricted diet on growth performance and intestinal immunity in weaned piglets.

  12. High Leucine Diets Stimulate Cerebral Branched-Chain Amino Acid Degradation and Modify Serotonin and Ketone Body Concentrations in a Pig Model.

    Science.gov (United States)

    Wessels, Anna G; Kluge, Holger; Hirche, Frank; Kiowski, Andreas; Schutkowski, Alexandra; Corrent, Etienne; Bartelt, Jörg; König, Bettina; Stangl, Gabriele I

    2016-01-01

    In addition to its role as an essential protein component, leucine (Leu) displays several other metabolic functions such as activation of protein synthesis. This property makes it an interesting amino acid for the therapy of human muscle atrophy and for livestock production. However, Leu can stimulate its own degradation via the branched-chain keto acid dehydrogenase complex (BCKDH). To examine the response of several tissues to excessive Leu, pigs were fed diets containing two- (L2) and four-fold (L4) higher Leu contents than the recommended amount (control). We found that the L4 diet led to a pronounced increase in BCKDH activity in the brain (2.5-fold, P serotonin synthesis. Compared to the controls, pigs treated with the high Leu diets consumed less food, showed increased plasma concentrations of 3-hydroxybutyrate and reduced levels of circulating serotonin. In conclusion, excessive Leu can stimulate BCKDH activity in several tissues, including the brain. Changes in cerebral tryptophan, along with the changes in amino acid-derived metabolites in the plasma may limit the use of high Leu diets to treat muscle atrophy or to increase muscle growth.

  13. Roux-en-Y Gastric Bypass Surgery, but Not Calorie Restriction, Reduces Plasma Branched-Chain Amino Acids in Obese Women Independent of Weight Loss or the Presence of Type 2 Diabetes

    NARCIS (Netherlands)

    Lips, M.A.; Klinken, J.B. van; Harmelen, V. van; Dharuri, H.K.; Hoen, P.A.C. 't; Laros, J.F.; Ommen, G.J.B. van; Janssen, I.M; Ramshorst, B. van; Wagensveld, B.A. van; Swank, D.J.; Dielen, F. Van; Dane, A.; Harms, A.; Vreeken, R.; Hankemeier, T.; Smit, J.W.A.; Pijl, H.; Dijk, K van

    2014-01-01

    OBJECTIVE: Obesity and type 2 diabetes mellitus (T2DM) have been associated with increased levels of circulating branched-chain amino acids (BCAAs) that may be involved in the pathogenesis of insulin resistance. However, weight loss has not been consistently associated with the reduction of BCAA

  14. Roux-en-Y Gastric Bypass Surgery, but Not Calorie Restriction, Reduces Plasma Branched-Chain Amino Acids in Obese Women Independent of Weight Loss or the Presence of Type 2 Diabetes

    NARCIS (Netherlands)

    Lips, M.A.; Klinken, J.B. van; Harmelen, V. van; Dharuri, H.K.; Hoen, P.A.C. 't; Laros, J.F.; Ommen, G.J.B. van; Janssen, I.M; Ramshorst, B. van; Wagensveld, B.A. van; Swank, D.J.; Dielen, F. Van; Dane, A.; Harms, A.; Vreeken, R.; Hankemeier, T.; Smit, J.W.A.; Pijl, H.; Dijk, K van

    2014-01-01

    OBJECTIVE: Obesity and type 2 diabetes mellitus (T2DM) have been associated with increased levels of circulating branched-chain amino acids (BCAAs) that may be involved in the pathogenesis of insulin resistance. However, weight loss has not been consistently associated with the reduction of BCAA lev

  15. Whey protein supplementation does not alter plasma branched-chained amino acid profiles but results in unique metabolomics patterns in obese women enrolled in an 8-week weight loss trial

    Science.gov (United States)

    Background: It has been suggested that perturbations in branched-chain amino acid (BCAA) catabolism are associated with insulin resistance and contribute to elevated systemic BCAAs. Evidence in rodents suggests dietary protein rich in BCAAs can increase BCAA catabolism, but there is limited evidence...

  16. Growth of the salt-tolerant yeast Zygosaccharomyces rouxii in microtiter plates : effects of NaCl, pH and temperature on growth and fusel alcohol production from branched-chain amino acids

    NARCIS (Netherlands)

    Jansen, Michael; Veurink, Janine H.; Euverink, Gert-Jan W.; Dijkhuizen, Lubbert

    2003-01-01

    Zygosaccharomyces rouxii, a salt-tolerant yeast isolated from the soy sauce process, produces fusel alcohols (isoamyl alcohol, active amyl alcohol and isobutyl alcohol) from branched-chain amino acids (leucine, isoleucine and valine, respectively) via the Ehrlich pathway. Using a high-throughput scr

  17. Eucalyptus ESTs associated with resistance to herbicide inhibitors of aromatic and branched-chain amino acid synthesis

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    Edivaldo Domingues Velini

    2005-01-01

    Full Text Available Herbicides inhibit enzymatic systems of plants. Acetolactate synthase (ALS, EC = 4.1.3.18 and 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS, EC 2.5.1.19 are key enzymes for herbicide action. Hundreds of compounds inhibit ALS. This enzyme is highly variable, enabling the selective control of weeds in a number of crops. Glyphosate, the only commercial herbicide inhibiting EPSPS is widely used for non-selective control of weeds in many crops. Recently, transgenic crops resistant to glyphosate were developed and have been used by farmers. The aim of this study was the data mining of eucalypt expressed sequence tags (ESTs in the FORESTs Genome Project database (https://forests.esalq.usp.br related to these enzymes. Representative amino acid sequences from the NCBI database associated with ALS and EPSPS were blasted with ESTs from the FORESTs database using the tBLASTx option of the blast tool. The best blasting reads and clusters from FORESTs, represented as nucleotide sequences, were blasted back with the NCBI database to evaluate the level of similarity with available sequences from different species. One and seven clusters were identified as showing high similarity with EPSPS and ALS sequences from the literature, respectively. The alignment of EPSPS sequences allowed the identification of conserved regions that can be used to design specific primers for additional sequencings.

  18. Improvement of regional cerebral blood flow after oral intake of branched-chain amino acids in patients with cirrhosis

    Institute of Scientific and Technical Information of China (English)

    Mika Yamamoto; Motoh Iwasa; Kaname Matsumura; Yuri Nakagawa; Naoki Fujita; Yoshinao Kobayashi; Masahiko Kaito; Kan Takeda; Yukihiko Adachi

    2005-01-01

    AIM: To evaluate the effect of oral intake of branchedchain amino acids (BCAA) on brain perfusion in patients with liver cirrhosis.METHODS: Single photon emission computed tomography scans were performed in 43 patients with cirrhosis and in 15 age-matched healthy subjects.Twenty-nine out of forty-three patients were randomly treated with either BCAA granules or placebo, and single photon emission computed tomography was performed before and after the treatment. We measured the regional cerebral blood flow values using a threedimensional stereotaxic region of interest template.RESULTS: Cirrhotic patients had regions of significant hypoperfusion in the bilateral central (right P=0.039,P<0.05; left P = 0.006 P<0.01), parietal (right P=0.018, P<0.05;left P=0.009, P<0.01), angular (right P=0.039, P<0.05;left P = 0.008, P<0.01), and left pericallosal segments (P= 0.038 P<0.05) as compared with healthy subjects. A significant increase in cerebral perfusion was observed 70 min after the oral intake of BCAA in the angular (right P=0.012,P<0.05;left P=0.049, P<0.05), temporal (right P=0.012, P<0.05; left P=0.038, P<0.05), pericallosal segments (right P = 0.025,P<0.05; left P = 0.049, P<0.05) and left precentral (P=0.044, P<0.05), parietal (P=0.040, P<0.05) and thalamus (P=0.033, P<0.05). No significant change in perfusion was observed in the placebo group.CONCLUSION: Administration of BCAA rapidly improves cerebral perfusion.

  19. Valine, a Branched-Chain Amino Acid, Reduced HCV Viral Load and Led to Eradication of HCV by Interferon Therapy in a Decompensated Cirrhotic Patient

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    Takumi Kawaguchi

    2012-10-01

    Full Text Available A decreased serum level of branched-chain amino acid (BCAA is a distinctive metabolic disorder in patients with liver cirrhosis. Recently, BCAA has been reported to exert various pharmacological activities, and valine, which is a BCAA, has been shown to affect lipid metabolism and the immune system in in vivo experiments. However, the clinical impact of valine supplementation on viral hepatitis C virus (HCV load has never been reported. Here, we first describe a case of HCV-related advanced liver cirrhosis that was treated by an oral valine agent. The administration of valine resulted in an improvement of fatigue and a reduction in hepatic fibrosis indexes as well as serum α-fetoprotein level. Furthermore, a marked reduction in HCV RNA levels was seen after valine treatment. The patient was then treated by interferon β, resulting in the successful eradication of chronic HCV infection. Thus, valine may be involved in the reduction of HCV viral load and could support a sustained virologic response to interferon therapy.

  20. Protective effects of branched-chain amino acids on hepatic ischemia-reperfusion-induced liver injury in rats: a direct attenuation of Kupffer cell activation.

    Science.gov (United States)

    Kitagawa, Tomomi; Yokoyama, Yukihiro; Kokuryo, Toshio; Nagino, Masato

    2013-02-15

    We determined whether there is a protective effect of branched-chain amino acid (BCAA) on hepatic ischemia-reperfusion (I/R)-induced acute liver injury. Wister rats were divided into the following four groups: simple laparotomy with vehicle; simple laparotomy with BCAA (1 g/kg body wt orally); I/R (30 min clamp) with vehicle; and I/R with BCAA. Serum liver function tests and the gene expression of adhesion molecules (intercellular adhesion molecule and vascular cell adhesion molecule) and vasoconstrictor-related genes (endothelin-1) in the liver were examined. In the in vivo study, portal venous pressure, leukocyte adhesion, and hepatic microcirculation were evaluated. Furthermore, Kupffer cells were isolated and cultured with various concentrations of BCAA in the presence or absence of lipopolysaccharide (LPS). Increased levels of liver function tests following I/R were significantly attenuated by BCAA treatment. The increased expression of adhesion molecules and endothelin-1 was also significantly attenuated by BCAA treatment. Moreover, increased portal venous pressure, enhanced leukocyte adhesion, and deteriorated hepatic microcirculation following I/R were all improved by BCAA treatment. In the experiment using isolated Kupffer cells, the expression of interleukin-6, interleukin-1β, and endothelin-1 in response to LPS stimulation was attenuated by BCAA in a dose-dependent fashion. These results indicate that perioperative oral administration of BCAA has excellent therapeutic potential to reduce I/R-induced liver injury. These beneficial effects may result from the direct attenuation of Kupffer cell activation under stressful conditions.

  1. High incidence of lipid deposition in the liver of rats fed a diet supplemented with branched-chain amino acids under vitamin B6 deficiency.

    Science.gov (United States)

    Kaimoto, Tae; Shibuya, Mayumi; Nishikawa, Kazutaka; Maeda, Hideo

    2013-01-01

    Male Wistar rats were fed four diets composed of purified 20% vitamin-free casein diet with (+) or without (-) vitamin B(6) (7.0 mg of pyridoxine HCl/kg of diet) and with (+) or without (-) branched-chain amino acids (BCAAs) of valine, leucine, and isoleucine (4.75%): B(6)(+)BCAA(-); B(6)(+)BCAA(+); B(6)(-)BCAA(-); and B(6)(-)BCAA(+) for 21 d. Among rats fed the B(6)(-)BCAA(+) diet, about a half showed lipid deposition in the liver. On the other hand, serum triacylglycerol levels in the B(6)(-)BCAA(+) group tended to be decreased. Hepatic triacylglycerol and cholesterol levels tended to increase in the B(6)(-)BCAA(+) group compared with the other three groups. Serum apolipoprotein B and apolipoprotein E (apo E) levels in the B(6)(-)BCAA(+) group were the lowest among the three groups. In contrast, hepatic apo E levels in the B(6)(-)BCAA(+) group were the highest among the three groups. High-performance liquid chromatography of pooled serum of rats with lipid deposits revealed that triacylglycerol and cholesterol levels in very low-density lipoprotein (VLDL) were decreased compared with other diet groups. These results strongly suggest that one of the mechanisms of lipid deposition in rats fed a B(6)(-)BCAA(+) diet is due to impaired secretion of VLDL.

  2. Branched chain amino acids maintain the molecular weight of poly(γ-glutamic acid) of Bacillus licheniformis ATCC 9945 during the fermentation.

    Science.gov (United States)

    Mitsunaga, Hitoshi; Meissner, Lena; Büchs, Jochen; Fukusaki, Eiichiro

    2016-10-01

    Poly(γ-glutamic acid) mainly produced by Bacillus spp. is an industrially important compound due to several useful features. Among them, molecular weight is an important characteristic affecting on the physical properties such as viscosities and negative charge densities. However, it is difficult to control the molecular size of PGA since it decreases during fermentation. Previous study reported that PGA produced in the media containing different carbon sources such as glucose and glycerol showed differences in molecular weight. Therefore in this study, the effect of carbon source on the PGA molecular weight was examined; with the aim of developing a strategy to maintain the high molecular weight of PGA during fermentation. Our result showed that the weight average molecular weight (Mw) of PGA of Bacillus licheniformis ATCC 9945 cultivated in the media containing PTS-sugars were higher than the medium containing glycerol (non-PTS). The result of metabolome analysis indicated the possibility of CodY (a global regulator protein) activation in the cells cultivated in the media containing PTS-sugars. To mimic this effect, branched-chain amino acids (BCAAs), which are activators of CodY, were added to a medium containing glycerol. As the result, the Mw of PGA in the BCAAs-supplemented media were maintained and high during the early production phase compared to the non BCAAs-supplemented medium. These results indicate that BCAAs can repress the PGA molecular weight reduction during fermentation in B. licheniformis ATCC 9945.

  3. Effects of Diets Supplemented with Branched-Chain Amino Acids on the Performance and Fatigue Mechanisms of Rats Submitted to Prolonged Physical Exercise

    Science.gov (United States)

    Falavigna, Gina; Junior, Jonas Alves de Araújo; Rogero, Marcelo Macedo; Pires, Ivanir Santana de Oliveira; Pedrosa, Rogério Graça; Junior, Eivor Martins; de Castro, Inar Alves; Tirapegui, Julio

    2012-01-01

    This study aimed to determine the effects of diets chronically supplemented with branched-chain amino acids (BCAA) on the fatigue mechanisms of trained rats. Thirty-six adult Wistar rats were trained for six weeks. The training protocol consisted of bouts of swimming exercise (one hour a day, five times a week, for six weeks). The animals received a control diet (C) (n = 12), a diet supplemented with 3.57% BCAA (S1) (n = 12), or a diet supplemented with 4.76% BCAA (S2) (n = 12). On the last day of the training protocol, half the animals in each group were sacrificed after one hour of swimming (1H), and the other half after a swimming exhaustion test (EX). Swimming time until exhaustion was increased by 37% in group S1 and reduced by 43% in group S2 compared to group C. Results indicate that the S1 diet had a beneficial effect on performance by sparing glycogen in the soleus muscle (p < 0.05) and by inducing a lower concentration of plasma ammonia, whereas the S2 diet had a negative effect on performance due to hyperammonemia (p < 0.05). The hypothalamic concentration of serotonin was not significantly different between the 1H and EX conditions. In conclusion, chronic BCAA supplementation led to increased performance in rats subjected to a swimming test to exhaustion. However, this is a dose-dependent effect, since chronic ingestion of elevated quantities of BCAA led to a reduction in performance. PMID:23201847

  4. Effects of intraduodenal infusion of the branched-chain amino acid leucine on ad libitum eating, gut motor and hormone functions, and glycemia in healthy men.

    Science.gov (United States)

    Steinert, Robert E; Landrock, Maria F; Ullrich, Sina S; Standfield, Scott; Otto, Bärbel; Horowitz, Michael; Feinle-Bisset, Christine

    2015-10-01

    Branched-chain amino acids (BCAAs), particularly leucine, act as nutrient signals regulating protein synthesis and degradation as well as glucose metabolism. In addition, leucine has been demonstrated in animal experiments to modulate eating and energy homeostasis. We aimed to characterize the effects of physiologic and supraphysiologic loads of intraduodenal leucine on eating, gut hormone and motor functions, and blood glucose in humans. Twelve lean men were studied on 3 occasions in a randomized, double-blind order. Antropyloroduodenal motility, plasma ghrelin, cholecystokinin, glucagon-like peptide 1, peptide YY, insulin, glucagon, blood glucose, appetite perceptions, and gastrointestinal symptoms were measured during 90-min intraduodenal infusions of leucine at 0.15 kcal/min (total 3.3 g, 13.5 kcal), 0.45 kcal/min (total 9.9 g, 40.5 kcal), or saline (control). Ad libitum eating from a buffet lunch was quantified immediately after the infusions. Leucine at 0.45 kcal/min inhibited eating (energy intake by ∼13%, P eating are probably not primarily mediated by changes in gut motor and hormone functions, with perhaps the exception of cholecystokinin. Instead, increased plasma leucine concentrations may be a potential signal mediating the eating-inhibitory effect of leucine. The study was registered as a clinical trial with the Australia and New Zealand Clinical Trial Registry (www.anzctr.org.au) as ACTRN12613000899741. © 2015 American Society for Nutrition.

  5. Branched-Chain Amino Acid Plus Glucose Supplement Reduces Exercise-Induced Delayed Onset Muscle Soreness in College-Age Females

    Science.gov (United States)

    Leahy, Danielle T.; Pintauro, Stephen J.

    2013-01-01

    Supplementation with branched-chain amino acids (BCAAs) has been used to stimulate muscle protein synthesis following exercise. The purpose of this study was to determine if supplementation with BCAAs in combination with glucose would reduce exercise-induced delayed onset muscle soreness (DOMS). Using a double-blind crossover design, 20 subjects (11 females, 9 males) were randomly assigned to either BCAA (n = 10) or placebo (n = 10) groups. Subjects performed a squatting exercise to elicit DOMS and rated their muscle soreness every 24 hours for four days following exercise while continuing to consume the BCAA or placebo. Following a three-week recovery period, subjects returned and received the alternate BCAA or placebo treatment, repeating the same exercise and DOMS rating protocol for the next four days. BCAA supplementation in female subjects resulted in a significant decrease in DOMS versus placebo at 24 hours following exercise (P = 0.018). No significant effect of BCAA supplementation versus placebo was noted in male subjects nor when male and female results were analyzed together. This gender difference may be related to dose per body weight differences between male and female subjects. PMID:24967261

  6. Branched-chain amino acids inhibit the TGF-beta-induced down-regulation of taurine biosynthetic enzyme cysteine dioxygenase in HepG2 cells.

    Science.gov (United States)

    Hagiwara, Asami; Ishizaki, Sonoko; Takehana, Kenji; Fujitani, Shoji; Sonaka, Ichiro; Satsu, Hideo; Shimizu, Makoto

    2014-05-01

    Taurine deficiency has been suggested to contribute to the pathogenesis and complications of advanced hepatic diseases. The molecular basis for a low level of taurine associated with hepatic failure is largely unknown. Using carbon tetrachloride (CCl4)-induced cirrhotic rat model, we found that the activity and expression of cysteine dioxygenase (CDO), a rate-limiting enzyme in taurine synthesis, were significantly decreased in the liver of these rats. To investigate the underlying mechanisms for the suppression, we examined the effects of pathological cytokines on CDO expression in human hepatoma HepG2 cells. Among the several cytokines, transforming growth factor-β (TGF-β), one of the key mediators of fibrogenesis, suppressed Cdo1 gene transcription through the MEK/ERK pathway. Finally, we further examined potential effects of branched-chain amino acids (BCAA) on CDO expression, as it has been reported that oral BCAA supplementation increased plasma taurine level in the patients with liver cirrhosis. BCAA, especially leucine, promoted Cdo1 gene transcription, and attenuated TGF-β-mediated suppression of Cdo1 gene expression. These results indicate that the low plasma level of taurine in advanced hepatic disease is due to decreased hepatic CDO expression, which can be partly attributed to suppressive effect of TGF-β on Cdo1 gene transcription. Furthermore, our observation that BCAA promotes Cdo1 expression suggests that BCAA may be therapeutically useful to improve hepatic taurine metabolism and further suppress dysfunctions associated with low level of taurine in hepatic diseases.

  7. Supplementation with branched-chain amino acids ameliorates hypoalbuminemia, prevents sarcopenia, and reduces fat accumulation in the skeletal muscles of patients with liver cirrhosis.

    Science.gov (United States)

    Kitajima, Yoichiro; Takahashi, Hirokazu; Akiyama, Takumi; Murayama, Kenichiro; Iwane, Shinji; Kuwashiro, Takuya; Tanaka, Kenichi; Kawazoe, Seiji; Ono, Naofumi; Eguchi, Takahisa; Anzai, Keizo; Eguchi, Yuichiro

    2017-07-24

    Liver cirrhosis induces marked metabolic disorders, protein-energy malnutrition, and sarcopenia. The objective of the study reported here was to investigate the effects of dietary branched-chain amino acids (BCAAs) on systemic glucose metabolism, skeletal muscle, and prognosis of patients with liver cirrhosis. Japanese patients with liver cirrhosis (n = 21) were enrolled into a longitudinal study in which their diets were supplemented with BCAAs. We evaluated glucose metabolism and analyzed the skeletal muscle area index (SAI) and intramuscular adipose tissue content (IMAC) using computed tomography. After 48 weeks of supplementation with BCAAs, there were no changes in glucose metabolism and skeletal muscle findings. In patients with ameliorated hypoalbuminemia, IMAC was significantly decreased and SAI was preserved concomitant with decreasing 90- and 120-min post-challenge plasma glucose levels (P increased albumin levels, IMAC was significantly increased and the SAI was significantly decreased (P decreased IMAC and 20.0% in patients with increased IMAC. Amelioration of hypoalbuminemia associated with BCAA supplementation correlated with decreased fat accumulation in skeletal muscle, maintenance of skeletal muscle mass, and improved glucose sensitivity, all factors which may contribute to improving the survival of patients with liver cirrhosis.

  8. High Leucine Diets Stimulate Cerebral Branched-Chain Amino Acid Degradation and Modify Serotonin and Ketone Body Concentrations in a Pig Model.

    Directory of Open Access Journals (Sweden)

    Anna G Wessels

    Full Text Available In addition to its role as an essential protein component, leucine (Leu displays several other metabolic functions such as activation of protein synthesis. This property makes it an interesting amino acid for the therapy of human muscle atrophy and for livestock production. However, Leu can stimulate its own degradation via the branched-chain keto acid dehydrogenase complex (BCKDH. To examine the response of several tissues to excessive Leu, pigs were fed diets containing two- (L2 and four-fold (L4 higher Leu contents than the recommended amount (control. We found that the L4 diet led to a pronounced increase in BCKDH activity in the brain (2.5-fold, P < 0.05, liver (1.8-fold, P < 0.05 and cardiac muscle (1.7-fold, P < 0.05, whereas we found no changes in enzyme activity in the pancreas, skeletal muscle, adipose tissue and intestinal mucosa. The L2 diet had only weak effects on BCKDH activity. Both high Leu diets reduced the concentrations of free valine and isoleucine in nearly all tissues. In the brain, high Leu diets modified the amount of tryptophan available: for serotonin synthesis. Compared to the controls, pigs treated with the high Leu diets consumed less food, showed increased plasma concentrations of 3-hydroxybutyrate and reduced levels of circulating serotonin. In conclusion, excessive Leu can stimulate BCKDH activity in several tissues, including the brain. Changes in cerebral tryptophan, along with the changes in amino acid-derived metabolites in the plasma may limit the use of high Leu diets to treat muscle atrophy or to increase muscle growth.

  9. Total branched-chain amino acids requirement in patients with maple syrup urine disease by use of indicator amino acid oxidation with L-[1-13C]phenylalanine.

    Science.gov (United States)

    Riazi, Roya; Rafii, Mahroukh; Clarke, Joe T R; Wykes, Linda J; Ball, Ronald O; Pencharz, Paul B

    2004-07-01

    Maple syrup urine disease (MSUD) is an autosomal recessive disorder caused by defects in the mitochondrial multienzyme complex branched-chain alpha-keto acid dehydrogenase (BCKD; EC 1.2.4.4), responsible for the oxidative decarboxylation of the branched-chain ketoacids (BCKA) derived from the branched-chain amino acids (BCAA) leucine, valine, and isoleucine. Deficiency of the enzyme results in increased concentrations of the BCAA and BCKA in body cells and fluids. The treatment of the disease is aimed at keeping the concentration of BCAA below the toxic concentrations, primarily by dietary restriction of BCAA intake. The objective of this study was to determine the total BCAA requirements of patients with classical MSUD caused by marked deficiency of BCKD by use of the indicator amino acid oxidation (IAAO) technique. Five MSUD patients from the MSUD clinic of The Hospital for Sick Children participated in the study. Each was randomly assigned to different intakes of BCAA mixture (0, 20, 30, 50, 60, 70, 90, 110, and 130 mg.kg(-1).day(-1)), in which the relative proportion of BCAA was the same as that in egg protein. Total BCAA requirement was determined by measuring the oxidation of l-[1-(13)C]phenylalanine to (13)CO(2). The mean total BCAA requirement was estimated using a two-phase linear regression crossover analysis, which showed that the mean total BCAA requirement was 45 mg.kg(-1).day(-1), with the safe level of intake (upper 95% confidence interval) at 62 mg.kg(-1).day(-1). This is the first time BCAA requirements in patients with MSUD have been determined directly.

  10. Comparative study of amino acid, ammonia and pancreatic hormone levels in the blood of cirrhotic patients following intragastric and intravenous administration of a branched-chain amino acid-enriched solution.

    Directory of Open Access Journals (Sweden)

    Watanabe,Akiharu

    1983-10-01

    Full Text Available The blood levels of amino acids, ammonia and pancreatic hormones following the intragastric and intravenous administration of a branched-chain amino acid (BCAA-enriched solution were comparatively investigated in control subjects and patients with liver cirrhosis. There was no essential difference in the time course of serum amino acid and blood ammonia levels between the intragastric and intravenous infusions. Elevation of serum insulin concentrations in cirrhotic patients was significant only immediately after the administration through the enteral route. However, plasma glucagon levels increased similarly when the BCAA-enriched solution was administered through either route. The results indicate that both enteral and intravenous infusions will have similar therapeutic effects on the impaired protein metabolism in cirrhotic patients with protein-calorie malnutrition.

  11. Branched-chain amino acid-containing dipeptides, identified from whey protein hydrolysates, stimulate glucose uptake rate in L6 myotubes and isolated skeletal muscles.

    Science.gov (United States)

    Morifuji, Masashi; Koga, Jinichiro; Kawanaka, Kentaro; Higuchi, Mitsuru

    2009-02-01

    In earlier studies we showed that dietary whey protein increased skeletal muscle and liver glycogen content in exercise-trained rats. However, little is known about whether ingredients of whey protein stimulate skeletal muscle glycogen accumulation. The aim of this study was to identify bioactive peptides in whey protein hydrolysates (WPH) which stimulated glucose uptake and glycogen synthesis rate in skeletal muscles. Branched-chain amino acid (BCAA)-containing dipeptides in WPH were identified using LC/MS/MS. L6 myotubes and isolated epitrochlearis muscles were used for the glucose uptake assays. The myotubes and muscles were incubated with or without 1 mM dipeptides, LY294002 a phosphoinositide 3-kinase (PI3-kinase) inhibitor, or GF102903X an atypical protein kinase C (aPKC) inhibitor, followed by measurement of 2-deoxyglucose uptake. Isolated muscles were incubated for 3 h with or without 1 mM Ile-Leu to determine glycogen synthesis rate. The BCAA-containing dipeptides, Ile-Val, Leu-Val, Val-Leu, Ile-Ile, Leu-Ile, Ile-Leu, and Leu-Leu were detected in the WPH by LC/MS/MS. These dipeptides caused significant stimulation in glucose uptake rate in the L6 myotubes. Ile-Leu, the main component in WPH, also stimulated glucose uptake in isolated skeletal muscles. Stimulation of glucose uptake by Ile-Leu was completely inhibited by treatment with either LY294002, or GF109203X in both L6 cells and isolated muscles. Ile-Leu increased glycogen contents in isolated muscles. These results suggest that BCAA-containing bioactive dipeptides in WPH stimulate glucose uptake in skeletal muscles via the PI3-kinase and aPKC pathways, resulting in increased skeletal muscle glycogen contents.

  12. Branched-chain amino acids ameliorate fibrosis and suppress tumor growth in a rat model of hepatocellular carcinoma with liver cirrhosis.

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    Jung Hoon Cha

    Full Text Available PURPOSE: Recent studies have revealed that branched-chain amino acids (BCAA reduce the development of hepatocellular carcinoma (HCC in patients with obesity and hepatitis C virus infection by improving insulin resistance (IR. The aim of this study was to examine the anti-cancer and anti-fibrotic effects of BCAA on the development of diethylnitrosamine (DEN-induced HCC and liver cirrhosis in a rat model. METHODS: Male SD rats received weekly intraperitoneal injections of DEN (50 mg/kg of body weight for 16 weeks to induce HCC. They were fed a diet containing 3% casein, 3% or 6% BCAA for 13 weeks beginning 6 weeks after DEN administration. DEN was used to induce HCC through stepwise development from cirrhosis to HCC. The effect of BCAA was evaluated in tumor tissues by histopathologic analyses, reverse transcription-polymerase chain reaction, and Western blotting. RESULTS: The mean area and number of dysplastic nodules (DNs and tumors in the casein group tended to be larger than those in the BCAA group 16 weeks after DEN administration. The mean fibrotic area in the BCAA group was smaller than that in the casein group. The BCAA group showed decreased mRNA levels for markers of fibrosis, angiogenesis, and apoptosis inhibition. Compared with the casein group, the BCAA group had lower levels of α-smooth muscle actin, vascular endothelial growth factor, p-β-catenin, p-p38 mitogen-activated protein kinase, proliferating cell nuclear antigen, and caspase-3 protein expression, as well as a higher level of cleaved caspase-3 protein expression. CONCLUSIONS: BCAA supplementation of the diet ameliorated liver fibrosis and HCC development in a DEN-induced rat model of HCC with liver cirrhosis, but not in the IR model. These results provide a rationale for anti-fibrosis and chemoprevention using BCAA treatment for HCC with liver cirrhosis, as well as decreasing the ammonia level.

  13. Branched-chain amino acids enhance premature senescence through mammalian target of rapamycin complex I-mediated upregulation of p21 protein.

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    Masayuki Nakano

    Full Text Available Branched-chain amino acids (BCAAs have been applied as an oral supplementation to patients with liver cirrhosis. BCAAs not only improve nutritional status of patients but also decrease the incidence of liver cancer. Mammalian target of rapamycin (mTOR links cellular metabolism with growth and proliferation in response to nutrients, energy, and growth factors. BCAAs, especially leucine, have been shown to regulate protein synthesis through mTOR activities. On the other hand, cellular senescence is suggested to function as tumor suppressor mechanisms, and induced by a variety of stimuli including DNA damage-inducing drugs. However, it is not clear how BCAA supplementation prevents the incidence of liver cancer in patients with cirrhosis. Here we showed that human cancer cells, HepG2 and U2OS, cultured in medium containing BCAAs with Fischer's ratio about 3, which was shown to have highest activities to synthesize and secrete of albumin, had higher activities to induce premature senescence and elevate mTORC1 activities. Furthermore, BCAAs themselves enhanced the execution of premature senescence induced by DNA damage-inducing drugs, which was effectively prevented by rapamycin. These results strongly suggested the contribution of the mTORC1 pathway to the regulation of premature senescence. Interestingly, the protein levels of p21, a p53 target and well-known gene essential for the execution of cellular senescence, were upregulated in the presence of BCAAs. These results suggested that BCAAs possibly contribute to tumor suppression by enhancing cellular senescence mediated through the mTOR signalling pathway.

  14. Effects of Diets Supplemented with Branched-Chain Amino Acids on the Performance and Fatigue Mechanisms of Rats Submitted to Prolonged Physical Exercise

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    Inar Alves de Castro

    2012-11-01

    Full Text Available This study aimed to determine the effects of diets chronically supplemented with branched-chain amino acids (BCAA on the fatigue mechanisms of trained rats. Thirty-six adult Wistar rats were trained for six weeks. The training protocol consisted of bouts of swimming exercise (one hour a day, five times a week, for six weeks. The animals received a control diet (C (n = 12, a diet supplemented with 3.57% BCAA (S1 (n = 12, or a diet supplemented with 4.76% BCAA (S2 (n = 12. On the last day of the training protocol, half the animals in each group were sacrificed after one hour of swimming (1H, and the other half after a swimming exhaustion test (EX. Swimming time until exhaustion was increased by 37% in group S1 and reduced by 43% in group S2 compared to group C. Results indicate that the S1 diet had a beneficial effect on performance by sparing glycogen in the soleus muscle (p < 0.05 and by inducing a lower concentration of plasma ammonia, whereas the S2 diet had a negative effect on performance due to hyperammonemia (p < 0.05. The hypothalamic concentration of serotonin was not significantly different between the 1H and EX conditions. In conclusion, chronic BCAA supplementation led to increased performance in rats subjected to a swimming test to exhaustion. However, this is a dose-dependent effect, since chronic ingestion of elevated quantities of BCAA led to a reduction in performance.

  15. Effects of acute exposure to increased plasma branched-chain amino acid concentrations on insulin-mediated plasma glucose turnover in healthy young subjects.

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    Sarah Everman

    Full Text Available Plasma branched-chain amino acids (BCAA are inversely related to insulin sensitivity of glucose metabolism in humans. However, currently, it is not known whether there is a cause-and-effect relationship between increased plasma BCAA concentrations and decreased insulin sensitivity.To determine the effects of acute exposure to increased plasma BCAA concentrations on insulin-mediated plasma glucose turnover in humans.Ten healthy subjects were randomly assigned to an experiment where insulin was infused at 40 mU/m2/min (40U during the second half of a 6-hour intravenous infusion of a BCAA mixture (i.e., BCAA; N = 5 to stimulate plasma glucose turnover or under the same conditions without BCAA infusion (Control; N = 5. In a separate experiment, seven healthy subjects were randomly assigned to receive insulin infusion at 80 mU/m2/min (80U in association with the above BCAA infusion (N = 4 or under the same conditions without BCAA infusion (N = 3. Plasma glucose turnover was measured prior to and during insulin infusion.Insulin infusion completely suppressed the endogenous glucose production (EGP across all groups. The percent suppression of EGP was not different between Control and BCAA in either the 40U or 80U experiments (P > 0.05. Insulin infusion stimulated whole-body glucose disposal rate (GDR across all groups. However, the increase (% in GDR was not different [median (1st quartile - 3rd quartile] between Control and BCAA in either the 40U ([199 (167-278 vs. 186 (94-308] or 80 U ([491 (414-548 vs. 478 (409-857] experiments (P > 0.05. Likewise, insulin stimulated the glucose metabolic clearance in all experiments (P 0.05.Short-term exposure of young healthy subjects to increased plasma BCAA concentrations does not alter the insulin sensitivity of glucose metabolism.

  16. Comparative metabolism of branched-chain amino acids to precursors of juvenile hormone biogenesis in corpora allata of lepidopterous versus nonlepidopterous insects

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    Brindle, P.A.; Schooley, D.A.; Tsai, L.W.; Baker, F.C.

    1988-08-05

    Comparative studies were performed on the role of branched-chain amino acids (BCAA) in juvenile hormone (JH) biosynthesis using several lepidopterous and nonlepidopterous insects. Corpora cardiaca-corpora allata complexes (CC-CA, the corpora allata being the organ of JH biogenesis) were maintained in culture medium containing a uniformly /sup 14/C-labeled BCAA, together with (methyl-/sup 3/H)methionine as mass marker for JH quantification. BCAA catabolism was quantified by directly analyzing the medium for the presence of /sup 14/C-labeled propionate and/or acetate, while JHs were extracted, purified by liquid chromatography, and subjected to double-label liquid scintillation counting. Our results indicate that active BCAA catabolism occurs within the CC-CA of lepidopterans, and this efficiently provides propionyl-CoA (from isoleucine or valine) for the biosynthesis of the ethyl branches of JH I and II. Acetyl-CoA, formed from isoleucine or leucine catabolism, is also utilized by lepidopteran CC-CA for biosynthesizing JH III and the acetate-derived portions of the ethyl-branched JHs. In contrast, CC-CA of nonlepidopterans fail to catabolize BCAA. Consequently, exogenous isoleucine or leucine does not serve as a carbon source for the biosynthesis of JH III by these glands, and no propionyl-CoA is produced for genesis of ethyl-branched JHs. This is the first observation of a tissue-specific metabolic difference which in part explains why these novel homosesquiterpenoids exist in lepidopterans, but not in nonlepidopterans.

  17. High concentration of branched-chain amino acids promotes oxidative stress, inflammation and migration of human peripheral blood mononuclear cells via mTORC1 activation.

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    Zhenyukh, Olha; Civantos, Esther; Ruiz-Ortega, Marta; Sánchez, Maria Soledad; Vázquez, Clotilde; Peiró, Concepción; Egido, Jesús; Mas, Sebastián

    2017-03-01

    Leucine, isoleucine and valine are essential aminoacids termed branched-chain amino acids (BCAA) due to its aliphatic side-chain. In several pathological and physiological conditions increased BCAA plasma concentrations have been described. Elevated BCAA levels predict insulin resistance development. Moreover, BCAA levels higher than 2mmol/L are neurotoxic by inducing microglial activation in maple syrup urine disease. However, there are no studies about the direct effects of BCAA in circulating cells. We have explored whether BCAA could promote oxidative stress and pro-inflammatory status in peripheral blood mononuclear cells (PBMCs) obtained from healthy donors. In cultured PBMCs, 10mmol/L BCAA increased the production of reactive oxygen species (ROS) via both NADPH oxidase and the mitochondria, and activated Akt-mTOR signalling. By using several inhibitors and activators of these molecular pathways we have described that mTOR activation by BCAA is linked to ROS production and mitochondrial dysfunction. BCAA stimulated the activation of the redox-sensitive transcription factor NF-κB, which resulted in the release of pro-inflammatory molecules, such as interleukin-6, tumor necrosis factor-α, intracellular adhesion molecule-1 or CD40L, and the migration of PBMCs. In conclusion, elevated BCAA blood levels can promote the activation of circulating PBMCs, by a mechanism that involving ROS production and NF-κB pathway activation. These data suggest that high concentrations of BCAA could exert deleterious effects on circulating blood cells and therefore contribute to the pro-inflammatory and oxidative status observed in several pathophysiological conditions.

  18. Preventive effects of branched-chain amino acid supplementation on the spontaneous development of hepatic preneoplastic lesions in C57BL/KsJ-db/db obese mice.

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    Terakura, Daishi; Shimizu, Masahito; Iwasa, Junpei; Baba, Atsushi; Kochi, Takahiro; Ohno, Tomohiko; Kubota, Masaya; Shirakami, Yohei; Shiraki, Makoto; Takai, Koji; Tsurumi, Hisashi; Tanaka, Takuji; Moriwaki, Hisataka

    2012-12-01

    Obesity and its associated disorders, such as non-alcoholic steatohepatitis, increase the risk of hepatocellular carcinoma. Branched-chain amino acids (BCAA), which improve protein malnutrition in patients with liver cirrhosis, reduce the risk of hepatocellular carcinoma in these patients with obesity. In the present study, the effects of BCAA supplementation on the spontaneous development of hepatic premalignant lesions, foci of cellular alteration, in db/db obese mice were examined. Male db/db mice were given a basal diet containing 3.0% of either BCAA or casein, a nitrogen-content-matched control of BCAA, for 36 weeks. On killing the mice, supplementation with BCAA significantly inhibited the development of foci of cellular alteration when compared with casein supplementation by inhibiting cell proliferation, but inducing apoptosis. BCAA supplementation increased the expression levels of peroxisome proliferator-activated receptor-γ, p21(CIP1) and p27(KIP1) messenger RNA and decreased the levels of c-fos and cyclin D1 mRNA in the liver. BCAA supplementation also reduced both the amount of hepatic triglyceride accumulation and the expression of interleukin (IL)-6, IL-1β, IL-18 and tumor necrosis factor-α mRNA in the liver. Increased macrophage infiltration was inhibited and the expression of IL-6, TNF-α, and monocyte chemoattractant protein-1 mRNA in the white adipose tissue were each decreased by BCAA supplementation. BCAA supplementation also reduced adipocyte size while increasing the expression of peroxisome proliferator-activated receptor-α, peroxisome proliferator-activated receptor-γ and adiponectin mRNA in the white adipose tissue compared with casein supplementation. These findings indicate that BCAA supplementation inhibits the early phase of obesity-related liver tumorigenesis by attenuating chronic inflammation in both the liver and white adipose tissue. BCAA supplementation may be useful in the chemoprevention of liver tumorigenesis in obese

  19. Branched-chain amino acids prevent hepatic fibrosis and development of hepatocellular carcinoma in a non-alcoholic steatohepatitis mouse model.

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    Takegoshi, Kai; Honda, Masao; Okada, Hikari; Takabatake, Riuta; Matsuzawa-Nagata, Naoto; Campbell, Jean S; Nishikawa, Masashi; Shimakami, Tetsuro; Shirasaki, Takayoshi; Sakai, Yoshio; Yamashita, Taro; Takamura, Toshinari; Tanaka, Takuji; Kaneko, Shuichi

    2017-02-13

    Oral supplementation with branched-chain amino acids (BCAA; leucine, isoleucine, and valine) in patients with liver cirrhosis potentially suppresses the incidence of hepatocellular carcinoma (HCC) and improves event-free survival. However, the detailed mechanisms of BCAA action have not been fully elucidated. BCAA were administered to atherogenic and high-fat (Ath+HF) diet-induced nonalcoholic steatohepatitis (NASH) model mice. Liver histology, tumor incidence, and gene expression profiles were evaluated. Ath+HF diet mice developed hepatic tumors at a high frequency at 68 weeks. BCAA supplementation significantly improved hepatic steatosis, inflammation, fibrosis, and tumors in Ath+HF mice at 68 weeks. GeneChip analysis demonstrated the significant resolution of pro-fibrotic gene expression by BCAA supplementation. The anti-fibrotic effect of BCAA was confirmed further using platelet-derived growth factor C transgenic mice, which develop hepatic fibrosis and tumors. In vitro, BCAA restored the transforming growth factor (TGF)-β1-stimulated expression of pro-fibrotic genes in hepatic stellate cells (HSC). In hepatocytes, BCAA restored TGF-β1-induced apoptosis, lipogenesis, and Wnt/β-Catenin signaling, and inhibited the transformation of WB-F344 rat liver epithelial stem-like cells. BCAA repressed the promoter activity of TGFβ1R1 by inhibiting the expression of the transcription factor NFY and histone acetyltransferase p300. Interestingly, the inhibitory effect of BCAA on TGF-β1 signaling was mTORC1 activity-dependent, suggesting the presence of negative feedback regulation from mTORC1 to TGF-β1 signaling. Thus, BCAA induce an anti-fibrotic effect in HSC, prevent apoptosis in hepatocytes, and decrease the incidence of HCC; therefore, BCAA supplementation would be beneficial for patients with advanced liver fibrosis with a high risk of HCC.

  20. Branched-chain amino acids, arginine, citrulline alleviate central fatigue after 3 simulated matches in taekwondo athletes: a randomized controlled trial.

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    Chen, I-Fan; Wu, Huey-June; Chen, Chung-Yu; Chou, Kuei-Ming; Chang, Chen-Kang

    2016-01-01

    The decline in cognitive performance has been shown after fatiguing exercise. Branched-chain amino acids (BCAA) have been suggested to alleviate exercise-induced central fatigue. Arginine and citrulline could remove the excess NH3 accumulation accompanied with BCAA supplementation by increasing nitric oxide biosynthesis and/or urea cycle. The purpose of this study is to investigate the effect of the combined supplementation of BCAA, arginine, and citrulline on central fatigue after three simulated matches in well-trained taekwondo athletes. In a double-blind randomized cross-over design, 12 male taekwondo athletes performed two trials containing three simulated matches each. Each match contained three 2-min rounds of high-intensity intermittent exercise. At the end of the second match, two different supplementations were consumed. In the AA trial, the subjects ingested 0.17 g/kg BCAA, 0.05 g/kg arginine and 0.05 g/kg citrulline, while placebo was consumed in the PL trial. A validated taekwondo-specific reaction test battery was used to measure the cognitive performance after each match. The premotor reaction time in the three single-task tests and the reaction time in the secondary task in the dual-task test were maintained in the AA trial after three matches, while they were impaired in the PL trial, resulting in significantly better performance in the AA trial. These improvements in the AA trial coincided with significantly lower plasma free tryptophan/BCAA ratio, increased NOx concentrations, and similar NH3 concentrations. This study suggested that the combined supplementation could alleviate the exercise-induced central fatigue in elite athletes.

  1. Exercise-induced muscle damage is reduced in resistance-trained males by branched chain amino acids: a randomized, double-blind, placebo controlled study

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    Howatson Glyn

    2012-07-01

    Full Text Available Abstract Background It is well documented that exercise-induced muscle damage (EIMD decreases muscle function and causes soreness and discomfort. Branched-chain amino acid (BCAA supplementation has been shown to increase protein synthesis and decrease muscle protein breakdown, however, the effects of BCAAs on recovery from damaging resistance training are unclear. Therefore, the aim of this study was to examine the effects of a BCAA supplementation on markers of muscle damage elicited via a sport specific bout of damaging exercise in trained volunteers. Methods Twelve males (mean ± SD age, 23 ± 2 y; stature, 178.3 ± 3.6 cm and body mass, 79.6 ± 8.4 kg were randomly assigned to a supplement (n = 6 or placebo (n = 6 group. The damaging exercise consisted of 100 consecutive drop-jumps. Creatine kinase (CK, maximal voluntary contraction (MVC, muscle soreness (DOMS, vertical jump (VJ, thigh circumference (TC and calf circumference (CC were measured as markers of muscle damage. All variables were measured immediately before the damaging exercise and at 24, 48, 72 and 96 h post-exercise. Results A significant time effect was seen for all variables. There were significant group effects showing a reduction in CK efflux and muscle soreness in the BCAA group compared to the placebo (P Conclusion The present study has shown that BCAA administered before and following damaging resistance exercise reduces indices of muscle damage and accelerates recovery in resistance-trained males. It seems likely that BCAA provided greater bioavailablity of substrate to improve protein synthesis and thereby the extent of secondary muscle damage associated with strenuous resistance exercise. Clinical Trial Registration Number: NCT01529281.

  2. Branched-chain amino acid supplements reduced ascites and increased the quality of life in a patient with liver cirrhosis: A case report.

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    Itou, Minoru; Kawaguchi, Takumi; Taniguchi, Eitaro; Oku, Yuichiro; Fukushima, Nobuyoshi; Ando, Eiji; Oriishi, Tetsuharu; Uchida, Yuki; Otsuka, Momoka; Tanaka, Suiko; Iwasaki, Shoko; Torii, Mari; Yoshida, Kiyomi; Adachi, Yuko; Suga, Mariko; Yoshiyama, Manami; Ibi, Ryoko; Akiyama, Yoshiko; Takakura, Machiko; Mitsuyama, Keiichi; Tsuruta, Osamu; Sata, Michio

    2009-01-01

    Liver cirrhosis is frequently accompanied by malnutrition and hypoalbuminemia, which in turn commonly induces ascites in patients with liver cirrhosis. Ascites leads to abdominal distention and appetite loss, resulting in a deteriorated quality of life (QOL). Administration of branched-chain amino acid (BCAA)-rich supplements reduces hepatic encephalopathy and malnutrition. In addition, BCAAs by themselves up-regulate albumin synthesis through an increase in Fisher's ratio. Thus, in patients with liver cirrhosis, BCAA-rich supplements seem to be effective at reducing ascites and improving the QOL. Here, we report the case of a 58-year-old Japanese man with liver cirrhosis with severe ascites and peripheral edema. The hepatic function of the patient was classified as Child-Pugh grade C. To reduce protein-energy malnutrition, BCAA-rich supplements were administered as a late evening snack as part of a regimen including 2000 kcal/day (32.5 kcal/kg/day) of total energy and 83.5 g/day (1.3 g/kg/day) of total protein intake. Eight weeks after admission, ascites and edema had decreased. Nutritional status also improved from the time of admission to discharge; the serum BCAA level increased from 365.4 to 450.2 µmol/l. Furthermore, the ratio of BCAAs to tyrosine (BTR) increased from 1.70 to 3.65. We also evaluated the effects of nutritional therapy on the patient's QOL using the Medical Outcomes Study 36-Item Short-Form Health Survey upon admission and at discharge. All subscores showed marked improvement and reached a level greater than the Japanese norm with nutritional treatment. In conclusion, BCAA supplementation not only reduced ascites, but also improved the QOL in a patient with liver cirrhosis.

  3. Suppressive effect of oral administration of branched-chain amino acid granules on oxidative stress and inflammation in HCV-positive patients with liver cirrhosis.

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    Ohno, Tomoyoshi; Tanaka, Yasuhito; Sugauchi, Fuminaka; Orito, Etsuro; Hasegawa, Izumi; Nukaya, Haruhiko; Kato, Atsunaga; Matunaga, Seijiro; Endo, Masayuki; Tanaka, Yoshito; Sakakibara, Kenji; Mizokami, Masashi

    2008-07-01

    In chronic hepatitis C virus (HCV) infection, it is thought that both chronic persistent inflammation and oxidative stress contribute to the development of hepatocellular carcinoma (HCC), and it has been reported that long-term oral supplementation with branched-chain amino acid (BCAA) granules could inhibit liver carcinogenesis. However, the extent of the involvement of these factors remains obscure. To clarify the involvement of inflammation and oxidative stress in the inhibition of liver carcinogenesis, we evaluated the effect of oral administration of BCAA granules on oxidative stress and inflammation in HCV-positive patients with liver cirrhosis. Twenty-seven patients were enrolled in the study: 18 of the patients were treated with BCAA granules (administered group) and nine were observed without BCAA granules (non-administered group). In the non-administered group, the production of oxidative stress, as indicated by urine 8-hydroxydeoxyguanosine (8-OHdG) and 15-F2t-Isoprostane (8-IsoPs), significantly increased with time, while in the administered group the levels of ferritin and 8-OHdG decreased significantly. Comparison of the two groups demonstrated that highly sensitive CRP, ferritin, 8-OHdG and 8-IsoPs were significantly reduced by taking BCAA granules. The time-course analysis showed that ferritin and highly sensitive CRP seemed to decrease first, followed by a decrease of 8-OHdG and 8-IsoPs. These findings indicated that the administration of BCAA granules influenced microinflammation and the metabolism of iron in HCV-positive patients with liver cirrhosis, and subsequently seemed to reduce the production of oxidative stress, possibly leading to a decrease in the occurrence of HCC.

  4. CypD(-/-) hearts have altered levels of proteins involved in Krebs cycle, branch chain amino acid degradation and pyruvate metabolism.

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    Menazza, Sara; Wong, Renee; Nguyen, Tiffany; Wang, Guanghui; Gucek, Marjan; Murphy, Elizabeth

    2013-03-01

    Cyclophilin D (CypD) is a mitochondrial chaperone that has been shown to regulate the mitochondrial permeability transition pore (MPTP). MPTP opening is a major determinant of mitochondrial dysfunction and cardiomyocyte death during ischemia/reperfusion (I/R) injury. Mice lacking CypD have been widely used to study regulation of the MPTP, and it has been shown recently that genetic depletion of CypD correlates with elevated levels of mitochondrial Ca(2+). The present study aimed to characterize the metabolic changes in CypD(-/-) hearts. Initially, we used a proteomics approach to examine protein changes in CypD(-/-) mice. Using pathway analysis, we found that CypD(-/-) hearts have alterations in branched chain amino acid metabolism, pyruvate metabolism and the Krebs cycle. We tested whether these metabolic changes were due to inhibition of electron transfer from these metabolic pathways into the electron transport chain. As we found decreased levels of succinate dehydrogenase and electron transfer flavoprotein in the proteomics analysis, we examined whether activities of these enzymes might be altered. However, we found no alterations in their activities. The proteomics study also showed a 23% decrease in carnitine-palmitoyltransferase 1 (CPT1), which prompted us to perform a metabolomics analysis. Consistent with the decrease in CPT1, we found a significant decrease in C4/Ci4, C5-OH/C3-DC, C12:1, C14:1, C16:1, and C20:3 acyl carnitines in hearts from CypD(-/-) mice. In summary, CypD(-/-) hearts exhibit changes in many metabolic pathways and caution should be used when interpreting results from these mice as due solely to inhibition of the MPTP.

  5. Effects of Combined Treatment with Branched-Chain Amino Acids, Citric Acid, L-Carnitine, Coenzyme Q10, Zinc, and Various Vitamins in Tumor-Bearing Mice.

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    Awa, Hiroko; Futamura, Akihiko; Higashiguchi, Takashi; Ito, Akihiro; Mori, Naoharu; Murai, Miyo; Ohara, Hiroshi; Chihara, Takeshi; Kaneko, Takaaki

    2017-03-01

    A functional dietary supplement (FDS) containing Coenzyme Q10, branched-chain amino acids and L-carnitine was administered to tumor-bearing mice, investigating its effects on tumor and muscle tissues. Experiment (A): B16 melanoma cells were implanted subcutaneously into the right side of the abdomen of 8- to 9-week-old C57BL/6J mice. The mice were divided into two groups: a FDS group that received oral administration of FDS (n=10), and a control group that received oral administration of glucose (n=10). The moribund condition was used as the endpoint, and median survival time was determined. Experiment (B): On day 21 after tumor implantation, tumors, soleus muscle, gastrocnemius muscle, and suprahyoid muscles were collected. Tumor and muscle weight and other aspects were evaluated in each group: FDS group (n=15) and control group (n=15). The median survival time was comparable (21 d in the FDS group vs. 18 d in the control group, p=0.30). However, cumulative food intake was significantly higher in the FDS group than the control group (p=0.011). Metastasis of melanoma to the lung was observed in the control group but not in the FDS group (p=0.043). The weight of the suprahyoid muscles was significantly higher in the FDS group than in the control group (p=0.0045). The weight of the tumor was significantly lower in the FDS group than in the control group (p=0.013). The results possibly suggest oral administration of FDS in tumor-bearing mice enhances the maintenance of suprahyoid muscles, resulting in an extended feeding period and suppression of tumor growth and metastasis.

  6. [Impact of glutamine, eicosapntemacnioc acid, branched-chain amino acid supplements on nutritional status and treatment compliance of esophageal cancer patients on concurrent chemoradiotherapy and gastric cancer patients on chemotherapy].

    Science.gov (United States)

    Cong, Minghua; Song, Chenxin; Zou, Baohua; Deng, Yingbing; Li, Shuluan; Liu, Xuehui; Liu, Weiwei; Liu, Jinying; Yu, Lei; Xu, Binghe

    2015-03-17

    To explore the effects of glutamine, eicosapntemacnioc acid (EPA) and branched-chain amino acids supplements in esophageal cancer patients on concurrent chemoradiotherapy and gastric cancer patients on chemotherapy. From April 2013 to April 2014, a total of 104 esophageal and gastric carcinoma patients on chemotherapy or concurrent chemoradiotherapy were recruited and randomly divided into experimental and control groups. Both groups received dietary counseling and routine nutritional supports while only experimental group received supplements of glutamine (20 g/d), EPA (3.3 g/d) and branched-chain amino acids (8 g/d). And body compositions, blood indicators, incidence of complications and completion rates of therapy were compared between two groups. After treatment, free fat mass and muscle weight increased significantly in experiment group while decreased in control group (P nutrition status, decrease the complications and improve compliance for esophageal cancer patients on concurrent chemo-radiotherapy and gastric cancer patients on postoperative adjuvant chemotherapy.

  7. Two randomized controlled studies comparing the nutritional benefits of branched-chain amino acid (BCAA) granules and a BCAA-enriched nutrient mixture for patients with esophageal varices after endoscopic treatment

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    Sakai, Yoshiyuki; Iwata, Yoshinori; Enomoto, Hirayuki; Saito, Masaki; Yoh, Kazunori; Ishii, Akio; Takashima, Tomoyuki; Aizawa, Nobuhiro; Ikeda, Naoto; Tanaka, Hironori; Iijima, Hiroko; Nishiguchi, Shuhei

    2014-01-01

    Background The usefulness of branched-chain amino acid (BCAA) granules and BCAA-enriched nutrient mixtures for patients with liver cirrhosis is often reported. However, no randomized controlled studies have investigated the usefulness of these supplements in the nutritional intervention of cirrhotic patients receiving endoscopic treatment for esophageal varices. Methods Patients without BCAA before endoscopic treatment were divided into study 1, and those who received BCAA were divided into s...

  8. Heating improves poor compliance with branched chain amino acid-rich supplementation in patients with liver cirrhosis: A before-after pilot study.

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    Itou, Minoru; Kawaguchi, Takumi; Taniguchi, Eitaro; Shiraishi, Satomi; Ibi, Ryoko; Mutou, Michiko; Okada, Teruyo; Uchida, Yuki; Otsuka, Momoka; Oriishi, Tetsuharu; Tanaka, Suiko; Takakura, Machiko; Mitsuyama, Keiichi; Tsuruta, Osamu; Sata, Michio

    2009-01-01

    Although branched chain amino acid (BCAA) supplementation improves malnutrition in cirrhotic patients, patient compliance with the administration of BCAA-rich supplements is poor due to their bitter taste. Since temperature is an important factor affecting taste, we examined the effect of heating on the stability of BCAAs and on the compliance of patients with liver cirrhosis with BCAA-rich supplement administration. A thermal denaturation test was first conducted, in which the BCAA-rich supplement Aminoleban® EN was heated to 37, 60, or 80°C for 30 or 60 min. The concentration of three amino acids, L-valine, L-leucine and L-isoleucine, was subsequently measured. The nutritional status of the cirrhotic patients was also evaluted. Patients presenting liver failure with a Child-Pugh class of A (n=2), B (n=2) or C (n=2) were hospitalized at Kurume University Hospital. Six patients with liver cirrhosis (HCV, n=3; HBV, n=1; alcohol, n=2) were enrolled. Venous blood samples were drawn in the morning after a 12-h overnight fast. The BCAA-rich supplement was administered to patients at room temperature (25°C) or heat loaded at 60°C for 10 min, with the temperature maintained above 45°C. Each patient was interviewed by a nationally registered dietitian regarding food consumption and intake of the BCAA-rich supplement immediately after each meal. Nutritional status was evaluated according to serum albumin levels, blood hemoglobin, prothrombin time and total lymphocyte count. No significant decrease was noted in valine, leucine or isoleucine levels following the heating of the BCAAs to 80°C. The caloric intake of the BCAA-rich supplement was significantly higher with administration after heating to 60°C, compared to caloric intake with administration at 25°C. In addition, heating of the BCAA-rich supplement significantly increased blood lymphocyte counts. In conclusion, heating did not affect the stability of the BCAAs, and may improve compliance with BCAA

  9. Regulation of adipose branched-chain amin acid catabolism enzyme expression and cross-adipose amino acid flux in human obesity

    Science.gov (United States)

    Elevated blood branched-chain amin acids (BCAA)are often assoicated with insulin resistance and type2 diabetes, which might result from a reduced cellular utilization and/or incomplete BCAA oxidation. White adipose tissue (WAT) has become appreciated as a potential player in whole body BCAA metaboli...

  10. Catabolism of Branched Chain Amino Acids Contributes Significantly to Synthesis of Odd-Chain and Even-Chain Fatty Acids in 3T3-L1 Adipocytes.

    Directory of Open Access Journals (Sweden)

    Scott B Crown

    Full Text Available The branched chain amino acids (BCAA valine, leucine and isoleucine have been implicated in a number of diseases including obesity, insulin resistance, and type 2 diabetes mellitus, although the mechanisms are still poorly understood. Adipose tissue plays an important role in BCAA homeostasis by actively metabolizing circulating BCAA. In this work, we have investigated the link between BCAA catabolism and fatty acid synthesis in 3T3-L1 adipocytes using parallel 13C-labeling experiments, mass spectrometry and model-based isotopomer data analysis. Specifically, we performed parallel labeling experiments with four fully 13C-labeled tracers, [U-13C]valine, [U-13C]leucine, [U-13C]isoleucine and [U-13C]glutamine. We measured mass isotopomer distributions of fatty acids and intracellular metabolites by GC-MS and analyzed the data using the isotopomer spectral analysis (ISA framework. We demonstrate that 3T3-L1 adipocytes accumulate significant amounts of even chain length (C14:0, C16:0 and C18:0 and odd chain length (C15:0 and C17:0 fatty acids under standard cell culture conditions. Using a novel GC-MS method, we demonstrate that propionyl-CoA acts as the primer on fatty acid synthase for the production of odd chain fatty acids. BCAA contributed significantly to the production of all fatty acids. Leucine and isoleucine contributed at least 25% to lipogenic acetyl-CoA pool, and valine and isoleucine contributed 100% to lipogenic propionyl-CoA pool. Our results further suggest that low activity of methylmalonyl-CoA mutase and mass action kinetics of propionyl-CoA on fatty acid synthase result in high rates of odd chain fatty acid synthesis in 3T3-L1 cells. Overall, this work provides important new insights into the connection between BCAA catabolism and fatty acid synthesis in adipocytes and underscores the high capacity of adipocytes for metabolizing BCAA.

  11. Intermediate Levels of Bacillus subtilis CodY Activity Are Required for Derepression of the Branched-Chain Amino Acid Permease, BraB.

    Directory of Open Access Journals (Sweden)

    Boris R Belitsky

    2015-10-01

    Full Text Available The global transcriptional regulator, CodY, binds strongly to the regulatory region of the braB gene, which encodes a Bacillus subtilis branched-chain amino acid (BCAA permease. However, under conditions that maximize CodY activity, braB expression was similar in wild-type and codY null mutant cells. Nonetheless, expression from the braB promoter was significantly elevated in cells containing partially active mutant versions of CodY or in wild-type cells under growth conditions leading to intermediate levels of CodY activity. This novel pattern of regulation was shown to be due to two opposing mechanisms, negative and positive, by which CodY affects braB expression. A strong CodY-binding site located downstream of the transcription start point conferred negative regulation by direct interaction with CodY. Additionally, sequences upstream and downstream of the promoter were required for repression by a second pleiotropic B. subtilis regulator, ScoC, whose own expression is repressed by CodY. ScoC-mediated repression of braB in codY null mutants cells was as efficient as direct, CodY-mediated repression in wild-type cells under conditions of high CodY activity. However, under conditions of reduced CodY activity, CodY-mediated repression was relieved to a greater extent than ScoC-mediated repression was increased, leading to elevated braB expression. We conclude that restricting increased expression of braB to conditions of moderate nutrient limitation is the raison d'être of the feed-forward regulatory loop formed by CodY and ScoC at the braB promoter. The increase in BraB expression only at intermediate activities of CodY may facilitate the uptake of BCAA when they are not in excess but prevent unneeded BraB synthesis when other BCAA transporters are active.

  12. Effect of branched-chain amino acid-enriched nutritional supplementation on interferon therapy in Japanese patients with chronic hepatitis C virus infection: a retrospective study

    Directory of Open Access Journals (Sweden)

    Nagao Yumiko

    2012-11-01

    Full Text Available Abstract Background The aims of this study were to evaluate the effects of nutritional supplementation with branched-chain amino acids (BCAA with zinc component (Aminofeel® on adherence to and outcome of therapy in patients treated with interferon (IFN for chronic hepatitis C and cirrhosis and to determine whether to recommend the supplement. Methods In this retrospective study, 51 patients who received IFN therapy were investigated among 203 consecutive patients who visited our hospital and were advised regarding the potential benefit of taking Aminofeel®. Each patient was free to choose whether to purchase and take Aminofeel®. Results Twenty four patients (group 1-A took Aminofeel® during standard IFN therapy and 13 (group 1-B did not. Low-dose, long-term IFN (maintenance therapy, mainly peglated (Peg-IFN alpha 2a, was administered to 14 patients who were difficult to treat, because of no effect or harmful side effects with standard IFN therapy, and who had advanced liver fibrosis. Among the 14, 11 patients (group 2-A took Aminofeel® and 3 (group 2-B did not. The prevalence of obesity was significantly higher (P=0.04 in group 1-A than in group 1-B. The rate of adherence to IFN therapy was higher in group 1-A (83.3% than in group 1-B (53.8%, P=0.05. There were no significant differences between the two groups in the rates of sustained virological response (SVR to IFN therapy. According to multivariate analysis, two factors, SVR and intake of Aminofeel®, were associated with successful adherence to IFN therapy. The adjusted odds ratios for these two factors were 13.25 and 12.59, respectively, and each was statistically significant. The SVR rate of maintenance IFN therapy was in 18.2% group 2-A and 0% in group 2-B. Conclusion Our data show that BCAA intake is useful for adherence to and effect of IFN therapy for patients with chronic hepatitis C. Nutritional supplementation with BCAA seems to be useful for HCV-infected patients receiving

  13. Catabolism of Branched Chain Amino Acids Contributes Significantly to Synthesis of Odd-Chain and Even-Chain Fatty Acids in 3T3-L1 Adipocytes.

    Science.gov (United States)

    Crown, Scott B; Marze, Nicholas; Antoniewicz, Maciek R

    2015-01-01

    The branched chain amino acids (BCAA) valine, leucine and isoleucine have been implicated in a number of diseases including obesity, insulin resistance, and type 2 diabetes mellitus, although the mechanisms are still poorly understood. Adipose tissue plays an important role in BCAA homeostasis by actively metabolizing circulating BCAA. In this work, we have investigated the link between BCAA catabolism and fatty acid synthesis in 3T3-L1 adipocytes using parallel 13C-labeling experiments, mass spectrometry and model-based isotopomer data analysis. Specifically, we performed parallel labeling experiments with four fully 13C-labeled tracers, [U-13C]valine, [U-13C]leucine, [U-13C]isoleucine and [U-13C]glutamine. We measured mass isotopomer distributions of fatty acids and intracellular metabolites by GC-MS and analyzed the data using the isotopomer spectral analysis (ISA) framework. We demonstrate that 3T3-L1 adipocytes accumulate significant amounts of even chain length (C14:0, C16:0 and C18:0) and odd chain length (C15:0 and C17:0) fatty acids under standard cell culture conditions. Using a novel GC-MS method, we demonstrate that propionyl-CoA acts as the primer on fatty acid synthase for the production of odd chain fatty acids. BCAA contributed significantly to the production of all fatty acids. Leucine and isoleucine contributed at least 25% to lipogenic acetyl-CoA pool, and valine and isoleucine contributed 100% to lipogenic propionyl-CoA pool. Our results further suggest that low activity of methylmalonyl-CoA mutase and mass action kinetics of propionyl-CoA on fatty acid synthase result in high rates of odd chain fatty acid synthesis in 3T3-L1 cells. Overall, this work provides important new insights into the connection between BCAA catabolism and fatty acid synthesis in adipocytes and underscores the high capacity of adipocytes for metabolizing BCAA.

  14. 运动中补充支链氨基酸对生理机能的有利效用%The Effects of Branched-Chain Amino Acid Supplements on Physiological Function during Exercises

    Institute of Scientific and Technical Information of China (English)

    俞璐; 丁树哲

    2009-01-01

    支链氨基酸(branched chain amino acid,BCAA)作为人体的必需氨基酸,与人体运动能力有着密切关系,能为长时间的耐力运动提供能量.从补充BCAA对抗中枢神经疲劳、对肌肉蛋白质代谢的作用及对线粒体功能的作用等几方面进行阐述诠释.

  15. Effect of dietary excess of branched-chain amino acids on performance and serum concentrations of amino acids in growing pigs.

    Science.gov (United States)

    Morales, A; Arce, N; Cota, M; Buenabad, L; Avelar, E; Htoo, J K; Cervantes, M

    2016-02-01

    Depressed performance and availability of some amino acids (AA) in pigs fed excess Leu diets appear to be related to lower feed intake. Surplus Ile and Val may help to overcome this effect. An experiment was conducted with 24 pigs (31.8 ± 1.2 kg initial BW) to evaluate the effect of dietary excess of either Leu alone or with surplus Ile and Val on performance and serum concentration (SC) of essential AA. Treatments were as follows: T1, basal diet; T2, basal plus 0.43% L-Leu (excess Leu); T3, basal added with 0.43% L-Leu, plus 0.20% L-Ile and 0.25% L-Val (excess LIV). The basal diet was formulated to contain 0.90% standardized ileal digestible Lys and added with crystalline L-Lys, L-Thr, DL-Met, L-Trp, L-Leu, L-Ile, L-His and L-Val to create essential AA:Lys ratios close to an ideal protein for growing pigs. All pigs were fed the same amount of feed twice a day (average, 3.42× the requirement of NEm). Blood samples were collected at 2.5 (absorptive) and 11.0 h (post-absorptive) post-prandial to analyse SC of AA. Excess of either Leu or LIV did not affect growth rate nor feed conversion. Excess Leu increased Leu SC and decreased Ile and Val SC (p pigs consuming excess Leu diets is attributed to a reduced absorption and increased cellular degradation rates of them.

  16. Novel consortium of Klebsiella variicola and Lactobacillus species enhances the functional potential of fermented dairy products by increasing the availability of branched-chain amino acids and the amount of distinctive volatiles.

    Science.gov (United States)

    Rosales-Bravo, H; Morales-Torres, H C; Vázquez-Martínez, J; Molina-Torres, J; Olalde-Portugal, V; Partida-Martínez, L P

    2017-08-17

    Identify novel bacterial taxa that could increase the availability of branched-chain amino acids and the amount of distinctive volatiles during skim milk fermentation. We recovered 344 bacterial isolates from stool samples of healthy and breastfed infants. Five were selected based on their ability to produce branched-chain amino acids. Three strains were identified as Escherichia coli, one as Klebsiella pneumoniae and other as Klebsiella variicola by molecular and biochemical methods. HPLC and solid-phase microextraction with GC-MS were used for the determination of free amino acids and volatile compounds respectively. The consortium formed by K. variicola and four Lactobacillus species showed the highest production of Leu and Ile in skim milk fermentation. In addition, the production of volatile compounds, such as acetoin, ethanol, 2-nonanone, and acetic, hexanoic and octanoic acids, increased in comparison to commercial yogurt, Emmental and Gouda cheese. Also, distinctive volatiles, such as 2,3-butanediol, 4-methyl-2- hexanone and octanol, were identified. The use of K. variicola in combination with probiotic Lactobacillus species enhances the availability of Leu and Ile and the amount of distinctive volatiles during skim milk fermentation. The identified consortium increases the functional potential of fermented dairy products. © 2017 The Society for Applied Microbiology.

  17. Research Progress of Branched Chain Amino Acids Applied in Hepatic Encephalopathy%支链氨基酸在肝性脑病中应用的研究进展

    Institute of Scientific and Technical Information of China (English)

    张丽丽; 蔡东联

    2011-01-01

    肝性脑病作为肝脏疾病终末期常见的并发症之一,严重降低病人的生活质量,影响疾病预后.不合理的营养摄人是肝性脑病的诱因之一.支链氨基酸的应用不仅可预防肝病病人发生肝性脑病,还可以降低肝性脑病病人的意识障碍.本文简述肝性脑病的发生机制,并从理论基础、临床研究叙述支链氨基酸的治疗作用机制,且对常见支链氨基酸药物及已报道的不良反应进行综述,目的在于规范支链氨基酸在肝性脑病中的应用,提高医师对其不良反应的重视.%As one of the common complications in the liver disease end - stage,hepatic encephalopathy seriously lowers the quality of patient lives and affects disease prognosis. Unreasonable nutritional intake is one of the reasons that lead to hepatic encephalopathy. The use of branched chain amino acids not only can prevent patients from hepatic encephalopathy , but also can lower the disorder of consciousness. This paper briefly narrates the happening mechanism of hepatic encephalopathy and lists the common medicine containing branched chain zmino acids.Besides , it summarizes the adverse reactions reported , aiming to set standards to the use of branched chain amino acids.

  18. Supplementation of branched-chain amino acids in protein-restricted diets modulates the expression levels of amino acid transporters and energy metabolism associated regulators in the adipose tissue of growing pigs

    Directory of Open Access Journals (Sweden)

    Yinghui Li

    2016-03-01

    Full Text Available This experiment was conducted to investigate the effects of branched-chain amino acids (BCAA supplemented in protein-restricted diets on the growth performance and the expression profile of amino acid transporters and energy metabolism related regulators in the white adipose tissue (WAT of different regional depots including dorsal subcutaneous adipose (DSA and abdominal subcutaneous adipose (ASA. A total of 24 crossbred barrows (7.40 ± 0.70 kg were randomly divided into 4 groups and were fed the following isocaloric diets for 33 days: 1 a recommended adequate protein diet (AP, 20% CP, as a positive control; 2 a low protein diet (LP, 17% CP; 3 the LP diet supplemented with BCAA (LP + B, 17% CP to reach the same level of the AP diet group; 4 the LP diet supplemented with 2 times the amount of BCAA (LP + 2B, 17% CP. The daily gain and daily feed intake of the LP diet group were the lowest among all the treatments (P  0.05. Moreover, BCAA supplementation down-regulated the expression levels of amino acid transporters including L-type amino acid transporter 1 and sodium-coupled neutral amino acid transporter 2 in DSA, but up-regulated the expression level of L-type amino acid transporter 4 in ASA (P < 0.05. Meanwhile, the energy sensor AMP-activated protein kinase α was activated in the DSA of pigs fed LP diet and in the ASA of the pigs fed AP or LP + 2B diets (P < 0.05. The mRNA expression profile of the selected mitochondrial component and mitochondrial biogenesis associated regulators in DSA and ASA also responded differently to dietary BCAA supplementation. These results suggested that the growth performance of growing pigs fed protein restricted diets supplemented with BCAA could catch up to that of the pigs fed AP diets. The results also partly demonstrated that the regulation mechanisms of BCAA are different in the adipose tissues of different depots.

  19. Intact Protein Analysis at 21 Tesla and X-Ray Crystallography Define Structural Differences in Single Amino Acid Variants of Human Mitochondrial Branched-Chain Amino Acid Aminotransferase 2 (BCAT2)

    Science.gov (United States)

    Anderson, Lissa C.; Håkansson, Maria; Walse, Björn; Nilsson, Carol L.

    2017-09-01

    Structural technologies are an essential component in the design of precision therapeutics. Precision medicine entails the development of therapeutics directed toward a designated target protein, with the goal to deliver the right drug to the right patient at the right time. In the field of oncology, protein structural variants are often associated with oncogenic potential. In a previous proteogenomic screen of patient-derived glioblastoma (GBM) tumor materials, we identified a sequence variant of human mitochondrial branched-chain amino acid aminotransferase 2 as a putative factor of resistance of GBM to standard-of-care-treatments. The enzyme generates glutamate, which is neurotoxic. To elucidate structural coordinates that may confer altered substrate binding or activity of the variant BCAT2 T186R, a 45 kDa protein, we applied combined ETD and CID top-down mass spectrometry in a LC-FT-ICR MS at 21 T, and X-Ray crystallography in the study of both the variant and non-variant intact proteins. The combined ETD/CID fragmentation pattern allowed for not only extensive sequence coverage but also confident localization of the amino acid variant to its position in the sequence. The crystallographic experiments confirmed the hypothesis generated by in silico structural homology modeling, that the Lys59 side-chain of BCAT2 may repulse the Arg186 in the variant protein (PDB code: 5MPR), leading to destabilization of the protein dimer and altered enzyme kinetics. Taken together, the MS and novel 3D structural data give us reason to further pursue BCAT2 T186R as a precision drug target in GBM. [Figure not available: see fulltext.

  20. Intact Protein Analysis at 21 Tesla and X-Ray Crystallography Define Structural Differences in Single Amino Acid Variants of Human Mitochondrial Branched-Chain Amino Acid Aminotransferase 2 (BCAT2)

    Science.gov (United States)

    Anderson, Lissa C.; Håkansson, Maria; Walse, Björn; Nilsson, Carol L.

    2017-07-01

    Structural technologies are an essential component in the design of precision therapeutics. Precision medicine entails the development of therapeutics directed toward a designated target protein, with the goal to deliver the right drug to the right patient at the right time. In the field of oncology, protein structural variants are often associated with oncogenic potential. In a previous proteogenomic screen of patient-derived glioblastoma (GBM) tumor materials, we identified a sequence variant of human mitochondrial branched-chain amino acid aminotransferase 2 as a putative factor of resistance of GBM to standard-of-care-treatments. The enzyme generates glutamate, which is neurotoxic. To elucidate structural coordinates that may confer altered substrate binding or activity of the variant BCAT2 T186R, a 45 kDa protein, we applied combined ETD and CID top-down mass spectrometry in a LC-FT-ICR MS at 21 T, and X-Ray crystallography in the study of both the variant and non-variant intact proteins. The combined ETD/CID fragmentation pattern allowed for not only extensive sequence coverage but also confident localization of the amino acid variant to its position in the sequence. The crystallographic experiments confirmed the hypothesis generated by in silico structural homology modeling, that the Lys59 side-chain of BCAT2 may repulse the Arg186 in the variant protein (PDB code: 5MPR), leading to destabilization of the protein dimer and altered enzyme kinetics. Taken together, the MS and novel 3D structural data give us reason to further pursue BCAT2 T186R as a precision drug target in GBM.

  1. Branched-chain amino acid ratios in low-protein diets regulate the free amino acid profile and the expression of hepatic fatty acid metabolism-related genes in growing pigs.

    Science.gov (United States)

    Duan, Y H; Li, F N; Wen, C Y; Wang, W L; Guo, Q P; Li, Y H; Yin, Y L

    2017-03-06

    Liver metabolism is affected by nutrients. The aim of this study was to explore the effects of low-protein diets (17% crude protein, CP) supplemented with branched-chain amino acids (BCAAs), including leucine (Leu), isoleucine (Ile) and valine (Val), on hepatic amino acid profile and lipid metabolism in growing pigs. The ratio of Leu : Ile : Val in all groups was 1 : 0.51 : 0.63 (20% crude protein, CP), 1 : 1 : 1 (17% CP), 1 : 0.75 : 0.75 (17% CP), 1 : 0.51 : 0.63 (17% CP) and 1 : 0.25 : 0.25 (17% CP) respectively. Results revealed that compared to the positive control group (1 : 0.51 : 0.63, 20% CP), the low-protein diets significantly augmented the concentrations of most essential amino acids and non-essential amino acids (p < .05), with the greatest values observed in the 1 : 0.25 : 0.25 group. Moreover, relative to the control, the low-protein diets with the Leu : Ile : Val ratio ranging from 1 : 0.75 : 0.75 to 1 : 0.25 : 0.25 markedly downregulated the mRNA abundance of acetyl-CoA carboxylase (ACC), lipoprotein lipase (LPL) and fatty acid-binding protein 4 (FABP-4) (p < .05), and upregulated the mRNA expression of hormone-sensitive lipase (HSL), peroxisome proliferator-activated receptor-g coactivator-1α (PGC-1α), uncoupling protein 3 (UCP3) and liver carnitine palmitoyltransferase 1 (L-CPT-1) (p < .05). Therefore, our data suggest that protein-restricted diets supplemented with optimal BCAA ratio, that is, 1 : 0.75 : 0.75-1 : 0.25 : 0.25, induce a shift from fatty acid synthesis to fatty acid oxidation in the liver of growing pigs. These effects may be associated with increased mitochondrial biogenesis.

  2. Branched-Chain Amino Acids as Pharmacological Nutrients in Chronic Liver Disease%支链氨基酸作为药理营养素在慢性肝病治疗中的应用

    Institute of Scientific and Technical Information of China (English)

    王芃; 谢青

    2011-01-01

    Branched-chain amino acids (BCAAs) are a group of essential amino acids including valine, leucine and isoleucine. A low ratio of plasma BCAAs to aromatic amino acids is a physiological hallmark of liver cirrhosis. Recent studies on BCAAs have revealed that, in addition to their role as protein constituents, they may have a role as pharmacological nutrients for patients with chronic liver disease. This review summarizes the possible effects of BCAAs on albumin synthesis and insulin resistance from clinical and basic viewpoints. Meanwhile, the newly discovered clinical impact of BCAAs on the prognosis and quality of life of patients with hepatocellular carcinoma and liver cirrhosis are reviewed.%支链氨基酸是一组包括缬氨酸、亮氨酸和异亮氨酸等的人体必需氨基酸.支链氨基酸除了其蛋白质成分的作用,可能对慢性肝病患者具有药理营养素作用.本文从基础研究和临床角度对支链氨基酸在合成白蛋白和胰岛素抵抗作用上可能产生的影响,以及对肝癌和肝硬化患者的生活质量和预后的临床作用作一综述.

  3. Consumo de aminoácidos de cadeia ramificada não afeta o desempenho de endurance Branched-chain amino acids ingestion does not affect endurance performance

    Directory of Open Access Journals (Sweden)

    Marco Carlos Uchida

    2008-02-01

    Full Text Available A suplementação com aminoácidos de cadeia ramificada (BCAA é uma das manipulações dietéticas mais populares entre atletas engajados em atividades de endurance. Entretanto, o papel ergogênico destes aminoácidos ainda não está totalmente estabelecido. Portanto, o objetivo do presente trabalho foi avaliar o efeito do consumo de BCAA sobre o exercício de endurance realizado até a exaustão. A fim de provocar redução do estoque de glicogênio muscular e, por conseguinte, maximizar a utilização dos BCAA, os sujeitos (n=17 foram submetidos a uma sessão prévia de exercício (corrida realizada a 75% do VO2max por 40 min seguida por 2 tiros a 90% do VO2max por 10 min cada um. Subseqüentemente, após o consumo aleatório de BCAA (77 mg.kg-1 ou placebo, seguindo modelo duplo cego cruzado, os participantes executaram um teste para determinação da capacidade de endurance (corrida a 90% do Limiar anaeróbio até a exaustão. Ambos os experimentos, BCAA e placebo, foram separados por uma semana. Com relação ao tempo até a exaustão e a distância percorrida, nenhuma diferença foi detectada entre as condições experimentais. (Placebo: 50,1±8,9 vs BCAA: 52,4±4,5 min, respectivamente (Placebo: 8,8±1,3 vs BCAA: 9,1±0,6 km, respectivamente. Além disto, também não foi evidenciada diferença na concentração plasmática de glicose, de lactato e de amônia entre ambas condições experimentais. Em conclusão, a suplementação de BCAA não afetou o desempenho de endurance em um teste de corrida até a exaustão.Branched-chain amino acids (BCAA supplementation is one of the most popular dietary manipulations used by endurance athletes. However, the ergogenic role of these amino acids in endurance exercise is not well established yet. Therefore, the aim of this study was to evaluate the effect of BCAA supplementation upon endurance exercise performed until exhaustion. In order to induce glycogen supply reduction, and thus maximize BCAA

  4. Identification and functional characterization of the Lactococcus lactis CodY-regulated branched-chain amino acid permease BcaP (CtrA)

    NARCIS (Netherlands)

    den Hengst, CD; Groeneveld, M; Kuipers, OP; Kok, J; Hengst, Chris D. den

    2006-01-01

    Transcriptome analyses have previously revealed that a gene encoding the putative amino acid transporter CtrA (YhdG) is one of the major targets of the pleiotropic regulator CodY in Lactococcus lactis and Bacillus subtilis. The role of ctrA in L. lactis was further investigated with respect to both

  5. Branched chain enriched amino acid versus glucose treatment of hepatic encephalopathy. A double-blind study of 65 patients with cirrhosis

    DEFF Research Database (Denmark)

    Vilstrup, Hendrik; Gluud, C; Hardt, F

    1990-01-01

    ), or isocaloric glucose (33 patients) for a maximum of 16 days. The regimens further included glucose infusion to a total of 26.5 kcal/kg per day and lactulose. The patients took part in the study for 5-6 days. In each group 17 patients woke up. In the amino acid group eleven died and four developed renal failure...

  6. Regulation of hepatic branched-chain alpha-keto acid dehydrogenase complex in rats fed a high-fat diet

    Science.gov (United States)

    Objective: Branched-chain alpha-keto acid dehydrogenase complex (BCKDC) regulates branched-chain amino acid (BCAA) metabolism at the level of branched chain alpha-ketoacid (BCKA) catabolism. It has been demonstrated that the activity of hepatic BCKDC is markedly decreased in type 2 diabetic animal...

  7. A tRNA body with high affinity for EF-Tu hastens ribosomal incorporation of unnatural amino acids.

    Science.gov (United States)

    Ieong, Ka-Weng; Pavlov, Michael Y; Kwiatkowski, Marek; Ehrenberg, Måns; Forster, Anthony C

    2014-05-01

    There is evidence that tRNA bodies have evolved to reduce differences between aminoacyl-tRNAs in their affinity to EF-Tu. Here, we study the kinetics of incorporation of L-amino acids (AAs) Phe, Ala allyl-glycine (aG), methyl-serine (mS), and biotinyl-lysine (bK) using a tRNA(Ala)-based body (tRNA(AlaB)) with a high affinity for EF-Tu. Results are compared with previous data on the kinetics of incorporation of the same AAs using a tRNA(PheB) body with a comparatively low affinity for EF-Tu. All incorporations exhibited fast and slow phases, reflecting the equilibrium fraction of AA-tRNA in active ternary complex with EF-Tu:GTP before the incorporation reaction. Increasing the concentration of EF-Tu increased the amplitude of the fast phase and left its rate unaltered. This allowed estimation of the affinity of each AA-tRNA to EF-Tu:GTP during translation, showing about a 10-fold higher EF-Tu affinity for AA-tRNAs formed from the tRNA(AlaB) body than from the tRNA(PheB) body. At ∼1 µM EF-Tu, tRNA(AlaB) conferred considerably faster incorporation kinetics than tRNA(PheB), especially in the case of the bulky bK. In contrast, the swap to the tRNA(AlaB) body did not increase the fast phase fraction of N-methyl-Phe incorporation, suggesting that the slow incorporation of N-methyl-Phe had a different cause than low EF-Tu:GTP affinity. The total time for AA-tRNA release from EF-Tu:GDP, accommodation, and peptidyl transfer on the ribosome was similar for the tRNA(AlaB) and tRNA(PheB) bodies. We conclude that a tRNA body with high EF-Tu affinity can greatly improve incorporation of unnatural AAs in a potentially generalizable manner.

  8. Effects of branched-chain amino acids and zinc-enriched nutrients on prognosticators in HCV-infected patients: a multicenter randomized controlled trial.

    Science.gov (United States)

    Kawaguchi, Takumi; Nagao, Yumiko; Abe, Kazumichi; Imazeki, Fumio; Honda, Koichi; Yamasaki, Kazumi; Miyanishi, Koji; Taniguchi, Eitaro; Kakuma, Tatsuyuki; Kato, Junji; Seike, Masataka; Yokosuka, Osamu; Ohira, Hiromasa; Sata, Michio

    2015-03-01

    Branched‑chain amino acids (BCAAs) and trace element deficiencies are associated with poor prognosis in hepatitis C virus (HCV)‑infected patients. The aim of this study was to investigate the effects of BCAA and zinc‑enriched supplementation on prognostic factors in HCV‑infected patients. Fifty‑three HCV‑infected patients were enrolled in this multicenter randomized controlled trial. The patients were assigned to either the placebo (n=27) or supplement group (n=26; 6,400 mg/day BCAAs and 10 mg/day zinc) and were followed up for 60 days. Primary outcomes were prognostic factors for chronic liver disease, including the serum BCAA‑to‑tyrosine ratio (BTR), zinc levels and α‑fetoprotein (AFP) levels. There were no significant differences in any of the prognostic factors between the placebo and supplement groups at baseline. In the supplement group, the BTR and zinc levels were significantly increased compared with the placebo group (BTR: 5.14 ± 1.59 vs. 4.23 ± 1.14, P=0.0290; zinc: 76 ± 11 vs. 68 ± 11 µg/dl, P=0.0497). No significant differences were observed in AFP levels between the groups in the whole analysis. However, a stratification analysis showed a significant reduction in ΔAFP levels in the supplement group, with elevated AFP levels compared with the other groups (‑2.72 ± 3.45 ng/ml, P=0.0079). It was demonstrated that BCAA and zinc‑enriched supplementation increased the BTR and zinc levels in the HCV‑infected patients. Furthermore, the supplementation reduced the serum AFP levels in patients who had elevated serum AFP levels at baseline. Thus, BCAA and zinc‑enriched supplementation may prolong the survival of HCV‑infected patients by improving amino acid imbalance and zinc deficiency, and by partly downregulating AFP.

  9. β-Alanine does not act through branched-chain amino acid catabolism in carp, a species with low muscular carnosine storage.

    Science.gov (United States)

    Geda, F; Declercq, A; Decostere, A; Lauwaerts, A; Wuyts, B; Derave, W; Janssens, G P J

    2015-02-01

    This study was executed to investigate the effect of dietary β-alanine (BA) on amino acid (AA) metabolism and voluntary feed intake in carp (Cyprinus carpio) at mildly elevated temperature to exert AA catabolism. Twenty-four fish in 12 aquaria were randomly assigned to either a control diet or the same diet with 500 mg BA/kg. A 14-day period at an ideal temperature (23 °C) was followed by 15 days at chronic mildly elevated temperature (27 °C). After the 15 days, all fish were euthanised for muscle analysis on histidine-containing dipeptides (HCD), whole blood on free AA and carnitine esters. The carnosine and anserine analysis indicated that all analyses were below the detection limit of 5 µmol/L, confirming that carp belongs to a species that does not store HCD. The increases in free AA concentrations due to BA supplementation failed to reach the level of significance. The effects of dietary BA on selected whole blood carnitine esters and their ratios were also not significant. The supplementation of BA tended to increase body weight gain (P = 0.081) and feed intake (P = 0.092). The lack of differences in the selected nutrient metabolites in combination with tendencies of improved growth performance warrants further investigation to unravel the mechanism of BA affecting feed intake. This first trial on the effect of BA supplementation on AA catabolism showed that its metabolic effect in carp at chronic mildly elevated temperature was very limited. Further studies need to evaluate which conditions are able to exert an effect of BA on AA metabolism.

  10. The Ratio of Dietary Branched-Chain Amino Acids is Associated with a Lower Prevalence of Obesity in Young Northern Chinese Adults: An Internet-Based Cross-Sectional Study.

    Science.gov (United States)

    Li, Yan-Chuan; Li, Ying; Liu, Li-Yan; Chen, Yang; Zi, Tian-Qi; Du, Shan-Shan; Jiang, Yong-Shuai; Feng, Ren-Nan; Sun, Chang-Hao

    2015-11-18

    This study aims to examine the association between the ratio of dietary branched chain amino acids (BCAA) and risk of obesity among young northern Chinese adults. A total of 948 randomly recruited participants were asked to finish our internet-based dietary questionnaire for the Chinese (IDQC). Associations between dietary BCAA ratio and prevalence of overweight/obesity and abdominal obesity were analyzed. Furthermore, 90 subjects were randomly selected to explore the possible mechanism. Dietary BCAA ratio in obese participants was significantly lower than non-obese participants. We found negative correlations between the ratio of dietary BCAA and body mass index (BMI) (r = -0.197, p BCAA ratio for overweight/obesity were 0.508 (0.265-0.972) and 0.389 (0.193-0.783), respectively (all p BCAA ratio were 0.351 (0.145-0.845) and 0.376 (0.161-0.876), respectively (all p BCAA ratio was inversely associated with 2-h postprandial glucose (2 h-PG) and status of inflammation. In conclusion, a higher ratio of dietary BCAA is inversely associated with prevalence of obesity, postprandial glucose and status of inflammation in young northern Chinese adults.

  11. Effects of Branched-Chain Amino Acid Supplements on Aerobic Exercise Stamina%补服支链氨基酸对有氧运动耐力的影响

    Institute of Scientific and Technical Information of China (English)

    邓杰

    2015-01-01

    为了研究补服支链氨基酸(branched-chain amino acid,BCAA)对有氧运动耐力的影响,给白鼠补服BCAA,检测其对白鼠的运动能力和血清游离氨基酸代谢的影响;另选择健康男性作为受试者,于测试前30 min分别口服氨基酸补剂及安慰剂,采取自身对照性分析法对比12 min跑成绩及最大吸氧量的变化.结果表明,白鼠补充BCAA后,有氧运动的衰竭推迟出现,血清中BCAA的含量显著提高;人体数据研究表明,补服BCAA后,12 min跑成绩和最大吸氧量均显著提高.表明补服BCAA能有效提高人体的有氧运动耐力.

  12. Effects and feasibility of exercise therapy combined with branched-chain amino acid supplementation on muscle strengthening in frail and pre-frail elderly people requiring long-term care: a crossover trial.

    Science.gov (United States)

    Ikeda, Takashi; Aizawa, Junya; Nagasawa, Hiroshi; Gomi, Ikuko; Kugota, Hiroyuki; Nanjo, Keigo; Jinno, Tetsuya; Masuda, Tadashi; Morita, Sadao

    2016-04-01

    This study examined the effects and feasibility of a twice-weekly combined therapy of branched-chain amino acids (BCAAs) and exercise on physical function improvement in frail and pre-frail elderly people requiring long-term care. We used a crossover design in which the combination of exercise and nutritional interventions was carried out twice a week during cycles A (3 months) and B (3 months) and the exercise intervention alone was performed during the washout period. The exercise intervention entailed the following 5 training sets: 3 sets of muscle training at 30% of maximum voluntary contraction, 1 set of aerobic exercise, and 1 set of balance training. For the nutritional intervention, 6 g of BCAAs or 6 g of maltodextrin was consumed 10 min before starting the exercise. We determined upper and lower limb isometric strength, performance on the Functional Reach Test (FRT) and the Timed Up and Go test, and activity level. In the comparison between the BCAA group and the control group after crossover, the improvement rates in gross lower limb muscle strength (leg press, knee extension) and FRT performance were significantly greater (by approximately 10%) in the BCAA group. In the comparison between different orders of BCAA administration, significant effects were shown for the leg press in both groups only when BCAAs were given. The combination of BCAA intake and exercise therapy yielded significant improvements in gross lower limb muscle strength and dynamic balance ability.

  13. Catabolism of leucine to branched-chain fatty acids in Staphylococcus xylosus

    DEFF Research Database (Denmark)

    Beck, Hans Christian; Hansen, A M; Lauritsen, F R

    2004-01-01

    Staphylococcus xylosus is an important starter culture in the production of flavours from the branched-chain amino acids leucine, valine and isoleucine in fermented meat products. The sensorially most important flavour compounds are the branched-chain aldehydes and acids derived from...

  14. 补充支链氨基酸对运动能力影响的研究进展%Research Development on Branched-Chain Amino Acid Supplement And Exercise Capacity

    Institute of Scientific and Technical Information of China (English)

    唐芳; 张蕴琨

    2012-01-01

    As necessary amino acids,Branched-chain amino acid(BCAA)is closely related to our exercise abilities.on the one hand,BCAA supplement can reduce protein catabolism,and promote gluconeogenesis,Therefore,the strength and anaerobic exercise capacity were improved.On the one hand,The ability of BCAA oxidation and decomposition in vivo to produce ATP is more efficiently than other amino acids.Branched-chain amino acid supplement can reduce the / ratio,and decrease the formation of 5-HT to provide energy reserving endurance exercise.However,some academics also suggested that the BCAA has no effect on exercise capacity and even have a negative impact,mainly because a large number of BCAA supplement may lead to blood ammonia elevated,thereby affecting exercise capacity,Therefore,scholars focus on the dose of BCAA during BCAA supplement.However,because of the complexity of the exercise test,the dose of BCAA supplement,the concentration of BCAA and the ratio of leucine,isoleucine and valine(Val) have no determine,how to supply BCAA reasonablly need further research Meanwhile,the mechanism of BCAA involved in glucose metabolism is not clear.In addition,during a long time,the effects of BCAA supplement on exercise capacity is still no conclusive.Following with application of micro-dialysis and other new technology,these problems will be solved.%支链氨基酸(BCAA)作为人体必需氨基酸,与人体的运动能力密切相关。一方面,补充BCAA能减少蛋白质的分解代谢,并促进糖异生,可以提高力量和无氧运动能力。另一方面,支链氨基酸在体内氧化分解产生ATP的效率高于其他氨基酸,补充BCAA可以降低[游离色氨酸(f-Trp)]/[BCAA]的比值,减少5-羟色胺(5-HT)形成,为机体长时间耐力运动提供能量。但是学术界也有人认为支链氨基酸对运动能力没有影响甚至会产生负面影响,主要原因是大量补充支链氨基酸会引起血氨升高,进而影响运动能力,故

  15. A Rapid and Sensitive UPLC-MS/MS-Method for the Separation and Quantification of Branched-Chain Amino Acids from Dried Blood Samples of Patients with Maple Syrup Urine Disease (MSUD

    Directory of Open Access Journals (Sweden)

    Ralph Fingerhut

    2016-06-01

    Full Text Available Newborn screening for MSUD is a special challenge since patients with MSUD can metabolically decompensate rapidly without adequate treatment within the first two weeks of life. However, the screening method does not detect the actual marker metabolite (alloisoleucine specifically, but only as part of the group of the other isobaric amino acids leucine, isoleucine and hydroxyproline. We describe a sensitive and rapid second-tier UPLC-MS/MS method to determine branched-chain amino acids from the initial extraction of the screening sample. Quantification is based on a seven-point calibration curve. Reference ranges (mean ± SD in µmol/L were determined from 179 normal, not pre-selected samples from the newborn screening: leucine: 72 ± 27; isoleucine: 37 ± 19; valine: 98 ± 46; hydroxyproline: 23 ± 13. The concentration of alloisoleucine was below the detection limit in about 55% of the cases, and the highest concentration was 1.9 µmol/L. In all 30 retrospectively studied screening samples from patients with confirmed MSUD the concentration of alloisoleucine was significantly increased. In 238 samples with false-positive newborn screening due to a significant increase in the combined concentration of leucine + isoleucine + alloisoleucine + hydroxyproline (400 to >4000 µmol/L, alloisoleucine was below 6.5 µmol/L (n = 57 or not detectable (n = 181. The application of this assay markedly reduces the false-positive rate and the associated anxiety and costs. It is also suitable for routinely monitoring blood spots of patients with MSUD.

  16. Effects of Supplementation of Branched-Chain Amino Acids to Reduced-Protein Diet on Skeletal Muscle Protein Synthesis and Degradation in the Fed and Fasted States in a Piglet Model

    Directory of Open Access Journals (Sweden)

    Liufeng Zheng

    2016-12-01

    Full Text Available Supplementation of branched-chain amino acids (BCAA has been demonstrated to promote skeletal muscle mass gain, but the mechanisms underlying this observation are still unknown. Since the regulation of muscle mass depends on a dynamic equilibrium (fasted losses–fed gains in protein turnover, the aim of this study was to investigate the effects of BCAA supplementation on muscle protein synthesis and degradation in fed/fasted states and the related mechanisms. Fourteen 26- (Experiment 1 and 28-day-old (Experiment 2 piglets were fed reduced-protein diets without or with supplemental BCAA. After a four-week acclimation period, skeletal muscle mass and components of anabolic and catabolic signaling in muscle samples after overnight fasting were determined in Experiment 1. Pigs in Experiment 2 were implanted with carotid arterial, jugular venous, femoral arterial and venous catheters, and fed once hourly along with the intravenous infusion of NaH13CO3 for 2 h, followed by a 6-h infusion of [1-13C]leucine. Muscle leucine kinetics were measured using arteriovenous difference technique. The mass of most muscles was increased by BCAA supplementation. During feeding, BCAA supplementation increased leucine uptake, protein synthesis, protein degradation and net transamination. The greater increase in protein synthesis than in protein degradation resulted in elevated protein deposition. Protein synthesis was strongly and positively correlated with the intramuscular net production of α-ketoisocaproate (KIC and protein degradation. Moreover, BCAA supplementation enhanced the fasted-state phosphorylation of protein translation initiation factors and inhibited the protein-degradation signaling of ubiquitin-proteasome and autophagy-lysosome systems. In conclusion, supplementation of BCAA to reduced-protein diet increases fed-state protein synthesis and inhibits fasted-state protein degradation, both of which could contribute to the elevation of skeletal muscle

  17. Effect on Nitrogen Balance, Thermogenesis, Body Composition, Satiety, and Circulating Branched Chain Amino Acid Levels up to One Year after Surgery: Protocol of a Randomized Controlled Trial on Dietary Protein During Surgical Weight Loss

    Science.gov (United States)

    Pi-Sunyer, Xavier; Vidal, Josep; Miner, Patricia; Boirie, Yves; Laferrère, Blandine

    2016-01-01

    Background Bariatric surgery (BS), the most effective treatment for severe obesity, typically results in 40-50 kg weight loss in the year following the surgery. Beyond its action on protein metabolism, dietary protein intake (PI) affects satiety, thermogenesis, energy efficiency, and body composition (BC). However, the required amount of PI after surgical weight loss is not known. The current daily PI recommendation for diet-induced weight loss is 0.8 g/kg ideal body weight (IBW) per day, but whether this amount is sufficient to preserve fat-free mass during active surgical weight loss is unknown. Objective To evaluate the effect of a 3-month dietary protein supplementation (PS) on nitrogen balance (NB), BC, energy expenditure, and satiety in women undergoing either gastric bypass or vertical sleeve gastrectomy. Methods In this randomized prospective study, participants will be randomized to a high protein supplementation group (1.2 g/kg IBW per day) or standard protein supplementation group (0.8 g/kg IBW per day) based on current guidelines. Outcome measures including NB, BC, circulating branched chain amino acids, and satiety, which will be assessed presurgery, and at 3-months and 12-months postsurgery. Results To date, no studies have examined the effect of dietary PS after BS. Current guidelines for PI after surgery are based on weak evidence. Conclusions The results of this study will contribute to the development of evidence-based data regarding the safe and optimal dietary PI and supplementation after BS. Trial Registration Clinicaltrials.gov NCT02269410; http://clinicaltrials.gov/ct2/show/NCT02269410 (Archived by WebCite at http://www.webcitation.org/6m2f2QLeg). PMID:27895003

  18. Effects of Supplementation of Branched-Chain Amino Acids to Reduced-Protein Diet on Skeletal Muscle Protein Synthesis and Degradation in the Fed and Fasted States in a Piglet Model.

    Science.gov (United States)

    Zheng, Liufeng; Wei, Hongkui; He, Pingli; Zhao, Shengjun; Xiang, Quanhang; Pang, Jiaman; Peng, Jian

    2016-12-28

    Supplementation of branched-chain amino acids (BCAA) has been demonstrated to promote skeletal muscle mass gain, but the mechanisms underlying this observation are still unknown. Since the regulation of muscle mass depends on a dynamic equilibrium (fasted losses-fed gains) in protein turnover, the aim of this study was to investigate the effects of BCAA supplementation on muscle protein synthesis and degradation in fed/fasted states and the related mechanisms. Fourteen 26- (Experiment 1) and 28-day-old (Experiment 2) piglets were fed reduced-protein diets without or with supplemental BCAA. After a four-week acclimation period, skeletal muscle mass and components of anabolic and catabolic signaling in muscle samples after overnight fasting were determined in Experiment 1. Pigs in Experiment 2 were implanted with carotid arterial, jugular venous, femoral arterial and venous catheters, and fed once hourly along with the intravenous infusion of NaH(13)CO₃ for 2 h, followed by a 6-h infusion of [1-(13)C]leucine. Muscle leucine kinetics were measured using arteriovenous difference technique. The mass of most muscles was increased by BCAA supplementation. During feeding, BCAA supplementation increased leucine uptake, protein synthesis, protein degradation and net transamination. The greater increase in protein synthesis than in protein degradation resulted in elevated protein deposition. Protein synthesis was strongly and positively correlated with the intramuscular net production of α-ketoisocaproate (KIC) and protein degradation. Moreover, BCAA supplementation enhanced the fasted-state phosphorylation of protein translation initiation factors and inhibited the protein-degradation signaling of ubiquitin-proteasome and autophagy-lysosome systems. In conclusion, supplementation of BCAA to reduced-protein diet increases fed-state protein synthesis and inhibits fasted-state protein degradation, both of which could contribute to the elevation of skeletal muscle mass in

  19. Consumption of Branched-Chain Amino Acids Is Inversely Associated with Central Obesity and Cardiometabolic Features in a Population of Brazilian Middle-Aged Men: Potential Role of Leucine Intake.

    Science.gov (United States)

    Cogate, P G; Natali, A J; de Oliveira, A; Alfenas, R C; Hermsdorff, H H M

    2015-08-01

    To assess the potential associations of branched-chain amino acids (BCAA) consumption with central obesity (CObesity) and cardiometabolic risk factors in a population of Brazilian middle-aged men. Special attention was given to leucine intake. Cross-sectional study. Universidade Federal de Viçosa, Viçosa, Brazil. Two-hundred ninety-six middle-aged men (Age: 50.5 ± 5.0 years; Body mass index: 25.8 ± 3.5 kg/m2). Anthropometry, lifestyle features, blood biochemical parameters were assessed and dietary intake was estimated by a food frequency questionnaire. Participants were classified by the occurrence of CObesity, hypertriglyceridemia (HTG), hypertriglyceridemic waist phenotype (HWP) and metabolic syndrome (MetS). Subjects in the BCAA highest tertile (≥0.17 g/kg/d) presented lower occurrence of CObesity (36.0% vs 72.4%, P<0.01, z=-5.10), HTG (17.0% vs 30.6% P<0.032, z=-2.32), HWP (23.0% vs 46.9%, P<0.01, z=-3.57) and MetS (19.0% vs 34.7%, P<0.01, z=-2.57). They also exhibited lower values for Castelli index (total cholesterol:HDL-c) and triglycerides: HDL-c ratio than those in the first tertile, regardless of interfering factors (i.e. habitual physical activity, work position, smoking habit, and energy intake). Interestingly, leucine consumption showed similar associations with cardiometabolic risk factors, as compared to BCAA consumption (P<0.05). BCAA consumption was cross-sectionally and negatively associated with relevant cardiometabolic risk factors in a population of Brazilian middle-aged men. The content of leucine in BCAA consumption may have contributed to such relationship.

  20. 支链氨基酸对大鼠TPH mRNA表达的影响%Effects of branched-chain amino acids spplementation on expression of TPH mRNA in rats

    Institute of Scientific and Technical Information of China (English)

    欧阳儒颖; 季红光; 蔡懿灵; 阮芳铭; 王海明

    2005-01-01

    目的探讨补充支链氨基酸(branched-chain amino acids,BCAA)对睡眠剥夺(sleep deprivation,SD)大鼠脑干色氨酸羟化酶(Tryptophan hydroxylase,TPH)mRNA表达的影响.方法采用小站台水环境(flower-pot)睡眠剥夺模型对大鼠进行睡眠剥夺.将56只雄性成年Sprague-Dawley大鼠按体重随机分为C(对照组,自由睡眠)、24 hSD(剥夺睡眠24 h)、24 h SDB(睡眠剥夺24h,进食添加3%BCAA的饲料)、48hSD、48h SDB、72 hSD和72 hSDB组,每组8只.睡眠剥夺结束后,断头取脑,冰上分离出脑干,半定量RT-PCR法检测TPH mBNA的表达.结果24和48h睡眠剥夺未明显改变大鼠脑干TPHmRNA的表达,与对照组比较差异无统计学意义(P>0.05);而72h睡眠剥夺后TPH mRNA表达量大幅度增加,与其它各组相比差异有统计学意义(P<0.01).补充BCAA与未补充BCAA组比较,大鼠脑干TPHmRNA的表达水平末见明显变化.结论一定时间的睡眠剥夺可以引起大鼠脑干TPHmRNA的表达上调,但补充一定剂量的支链氨基酸并不能改变睡眠剥夺大鼠脑干TPHmRNA的表达.

  1. Supplementation of branched-chain amino acids to a reduced-protein diet improves growth performance in piglets: involvement of increased feed intake and direct muscle growth-promoting effect.

    Science.gov (United States)

    Zheng, Liufeng; Wei, Hongkui; Cheng, Chuanshang; Xiang, Quanhang; Pang, Jiaman; Peng, Jian

    2016-06-01

    The aim of this study was to investigate whether supplementing branched-chain amino acids (AA) (BCAA) along with a reduced-protein diet increases piglet growth, and whether elevated feed intake and muscle growth-promoting effect contribute to this improvement. In Expt 1, twenty-eight weanling piglets were randomly fed one of the following four diets: a positive control (PC) diet, a reduced-protein negative control (NC) diet, an NC diet supplemented with BCAA to the same levels as in the PC diet (test 1 (T1)) and an NC diet supplemented with a 2-fold dose of BCAA in T1 diet (test 2 (T2)) for 28 d. In Expt 2, twenty-one weanling piglets were randomly assigned to NC, T1 and pair-fed T1 (P) groups. NC and T1 diets were the same as in Expt 1, whereas piglets in the P group were individually pair-fed with the NC group. In Expt 1, the NC group had reduced piglet growth and feed intake compared with the PC group, which were restored in T1 and T2 groups, but no differences were detected between T1 and T2 groups. In Expt 2, T1 and P groups showed increases in growth and mass of some muscles compared with the NC group. Increased feed intake after BCAA supplementation was associated with increased mRNA expressions of agouti-related peptide and co-express neuropeptide Y (NPY) and phosphorylation of mammalian target of rapamycin (mTOR) and ribosomal protein S6 kinase 1 (S6K1), as well as decreased mRNA expressions of melanocortin-4 receptor and cocaine- and amphetamine-regulated transcript and phosphorylation of eukaryotic initiation factor 2α in the hypothalamus. No differences were observed among PC, T1 and T2 groups except for higher NPY mRNA expression in the T2 group than in the PC group (Expt 1). Phosphorylation of mTOR and S6K1 in muscle was enhanced after BCAA supplementation, which was independent of change in feed intake (Expt 2). In conclusion, supplementing BCAA to reduced-protein diets increases feed intake and muscle mass, and contributes to better growth

  2. Rational design, synthesis and biological evaluations of amino-noscapine: a high affinity tubulin-binding noscapinoid

    Science.gov (United States)

    Naik, Pradeep K.; Chatterji, Biswa Prasun; Vangapandu, Surya N.; Aneja, Ritu; Chandra, Ramesh; Kanteveri, Srinivas; Joshi, Harish C.

    2011-05-01

    Noscapine and its derivatives are important microtubule-interfering agents shown to have potent anti-tumor activity. The binding free energies (Δ G bind) of noscapinoids computed using linear interaction energy (LIE) method with a surface generalized Born (SGB) continuum solvation model were in agreement with the experimental Δ G bind with average root mean square error of 0.082 kcal/mol. This LIE-SGB model guided us in designing a novel derivative of noscapine, amino-noscapine [(S)-3-((R)-9-amino-4-methoxy-6-methyl-5,6,7,8-tetrahydro [1, 3] dioxolo[4,5- g]isoquinolin-5-yl)-6,7-dimethoxy isobenzo-furan-1(3 H)-one] that has higher tubulin binding activity (predicted Δ G bind = -6.438 kcal/mol and experimental Δ G bind = -6.628 kcal/mol) than noscapine, but does not significantly change the total extent of the tubulin subunit/polymer ratio. The modes of interaction of amino-noscapine with the binding pocket of tubulin involved three hydrogen bonds and are distinct compared to noscapine which involved only one hydrogen bond. Also the patterns of non-bonded interactions are albeit different between both the lignads. The `blind docking' approach (docking of ligand with different binding sites of a protein and their evaluations) as well as the reasonable accuracy of calculating Δ G bind using LIE-SGB model constitutes the first evidence that this class of compounds binds to tubulin at a site overlapping with colchicine-binding site or close to it. Our results revealed that amino-noscapine has better anti-tumor activity than noscapine.

  3. Efeito dos aminoácidos de cadeia ramificada no músculo imobilizado e no músculo desnervado = Effect of branched - chain amino acids on immobilized skeletal muscle and denervated muscle

    Directory of Open Access Journals (Sweden)

    Pezolato, Vitor Alexandre

    2014-01-01

    Conclusão: Observa-se que a utilização dos aminoácidos de cadeia ramificada pode diminuir a atrofia provocada pelo desuso muscular, tanto na condição de desnervação quanto na condição de imobilização

  4. Association of increased serum branched-chain amino acid level with carotid plaque%血清支链氨基酸水平升高与颈动脉斑块的关系

    Institute of Scientific and Technical Information of China (English)

    隋小芳; 杨睿悦; 郭金发; 张磊艺; 黄佳滨; 费秀斌; 曾平; 朱玲

    2015-01-01

    目的:研究血清支链氨基酸(BCAA)水平与颈动脉斑块的关系。方法采用横断面调查方法,对收集的中老年体检人群472例[42~97岁,平均(70.1±6.6)岁,男性272例]进行问卷调查、体格检查、血液检查和颈动脉超声检测,同位素稀释液相色谱串联质谱法测定血清 BCAA 水平,分析其与颈动脉斑块的关系。结果血清 BCAA 浓度呈偏态和尖态分布,男性的 BCAA 水平[466.5(423.6~514.7)μmol/ L]显著高于女性[415.3(382.5~466.0)μmol/ L],P ﹤0.001。在校正年龄和性别后,血清 BCAA 水平与 BMI、SBP、DBP、FBG、TG 和 LDL-C 显著正相关(均为 P ﹤0.05),与 HDL-C 明显负相关(P ﹤0.001)。颈动脉斑块组的 BCAA 水平[450.0(405.9~492.1)]明显高于正常组[430.4(395.1~495.2)](P =0.039)。多因素 Logistic 回归分析发现,年龄、吸烟史、SBP 和 BCAA 水平升高是颈动脉斑块的独立危险因素,BCAA 次高四分位水平发生颈动脉斑块的风险是最低四分位水平的2.68倍(P =0.002,P趋势=0.018)。结论血清 BCAA 水平升高是颈动脉斑块的独立危险因素,可能影响动脉粥样硬化的发生发展。%Objective To investigate the association between serum branched chain amino acid (BCAA) and carotid plaque. Methods The cross-sectional study was undertaken on 472 middle-aged and elderly people [42-97 y, average (70. 1 ± 6. 6) y, 272 males] selected from the participants for health screen. The medical history of each participant was obtained by questionnaire. The samples of venous blood and the Bilateral B-mode carotid artery images were taken. Serum BCAA levels were determined by Isotope Dilution Liquid Chromatography Tandem Mass Spectrometry. The association of BCAA with carotid plaque was analyzed. Results The levels of BCAA showed skewed and leptokurtic distribution. Males [466. 5 (423. 6-514. 7 ) μmol/ L ] had significantly higher values of BCAA than females [ 415. 3 ( 382. 5-466. 0) μmol/ L] (P ﹤ 0. 001

  5. Effects of Branched-Chain Amino Acids on Growth Performance and Intestinal Development of Broilers%支链氨基酸对肉鸡生长性能及肠道发育的影响

    Institute of Scientific and Technical Information of China (English)

    常银莲; 刘国华; 常文环; 张姝; 郑爱娟; 蔡辉益

    2016-01-01

    This experiment was conducted to investigate the effects of branched-chain amino acids ( BCAAs) on the growth performance and intestinal development of broilers.Three hundred and eighty-four one-day-old Arbor Acres ( AA) healthy male broilers were randomly assigned into four groups with 6 replicates per group and 16 broilers per replicate.Each group was offered one of the following diets with different BCAAs levels:3.04%, 3.93%, 4.82%and 5.71%, respectively.The ratio of leucine (Leu), isoleucine (Ile) and valine ( Val) was 1.8∶1.0∶1.2.The experiment lasted for 21 days and all birds were free to feed and water.The re-sults showed that there was no significant difference in body weight or average daily gain among all groups ( P>0.05) , while the average daily feed intake in 5.71%group was significantly lower than that in 3.04%group ( P0.05) . At age of 10 and 21 days, the crypt depth of jejunum in 4.82%group was significantly higher than that in 3.93%group ( P0.05).The V/C of jejunum and ileum in 3.93%group was significantly higher than that in 5.71%group ( P<0.05) .In conclusion, under the situation of this experiment, with the in-crease of BCAAs level, the ratio of feed to gain can decrease;the addition of BCAAs with appropriate amount can promote intestinal development of broilers, while excessive addition can significantly decrease average daily feed intake and lead to the postponed development of the small intestine.%本试验旨在研究支链氨基酸( BCAAs)对肉鸡生长性能及肠道发育的影响。选取1日龄爱拔益加( AA)健康肉公鸡384只,随机分为4组,每组6个重复,每个重复16只。各组分别饲喂BCAAs水平为3.04%、3.93%、4.82%和5.71%的试验饲粮,各组饲粮中亮氨酸∶异亮氨酸∶缬氨酸均为1.8∶1.0∶1.2,自由采食和饮水,试验期21 d。结果表明:各组肉鸡体重及平均日增重均无显著差异(P>0.05),但5.71%组肉鸡

  6. Developmental Defects of Caenorhabditis elegans Lacking Branched-chain α-Ketoacid Dehydrogenase Are Mainly Caused by Monomethyl Branched-chain Fatty Acid Deficiency.

    Science.gov (United States)

    Jia, Fan; Cui, Mingxue; Than, Minh T; Han, Min

    2016-02-01

    Branched-chain α-ketoacid dehydrogenase (BCKDH) catalyzes the critical step in the branched-chain amino acid (BCAA) catabolic pathway and has been the focus of extensive studies. Mutations in the complex disrupt many fundamental metabolic pathways and cause multiple human diseases including maple syrup urine disease (MSUD), autism, and other related neurological disorders. BCKDH may also be required for the synthesis of monomethyl branched-chain fatty acids (mmBCFAs) from BCAAs. The pathology of MSUD has been attributed mainly to BCAA accumulation, but the role of mmBCFA has not been evaluated. Here we show that disrupting BCKDH in Caenorhabditis elegans causes mmBCFA deficiency, in addition to BCAA accumulation. Worms with deficiency in BCKDH function manifest larval arrest and embryonic lethal phenotypes, and mmBCFA supplementation suppressed both without correcting BCAA levels. The majority of developmental defects caused by BCKDH deficiency may thus be attributed to lacking mmBCFAs in worms. Tissue-specific analysis shows that restoration of BCKDH function in multiple tissues can rescue the defects, but is especially effective in neurons. Taken together, we conclude that mmBCFA deficiency is largely responsible for the developmental defects in the worm and conceivably might also be a critical contributor to the pathology of human MSUD.

  7. Periexercise coingestion of branched-chain amino acids and carbohydrate in men does not preferentially augment resistance exercise-induced increases in phosphatidylinositol 3 kinase/protein kinase B-mammalian target of rapamycin pathway markers indicative of muscle protein synthesis.

    Science.gov (United States)

    Ferreira, Maria Pontes; Li, Rui; Cooke, Matthew; Kreider, Richard B; Willoughby, Darryn S

    2014-03-01

    The effects of a single bout of resistance exercise (RE) in conjunction with periexercise branched-chain amino acid (BCAA) and carbohydrate (CHO) ingestion on skeletal muscle signaling markers indicative of muscle protein synthesis were determined. It was hypothesized that CHO + BCAA would elicit a more profound effect on these signaling markers compared with CHO. Twenty-seven males were randomly assigned to CHO, CHO + BCAA, or placebo (PLC) groups. Four sets of leg presses and leg extensions were performed at 80% 1 repetition maximum. Supplements were ingested 30 minutes and immediately before and after RE. Venous blood and muscle biopsy samples were obtained immediately before supplement ingestion and 0.5, 2, and 6 hours after RE. Serum insulin and glucose and phosphorylated levels of muscle insulin receptor substrate 1 (IRS-1), protein kinase B, mammalian target of rapamycin, phosphorylated 70S6 kinase, and 4E binding protein 1 were assessed. Data were analyzed by 2-way repeated-measures analysis of variance. Significant group × time interactions were observed for glucose and insulin (P protein kinase B (P = .031), mammalian target of rapamycin (P = .003), and phosphorylated 70S6 kinase (P = .001). Carbohydrate and CHO + BCAA supplementation significantly increased IRS-1 compared with PLC (P = .002). However, periexercise coingestion of CHO and BCAA did not augment RE-induced increases in skeletal muscle signaling markers indicative of muscle protein synthesis when compared with CHO.

  8. 补充支链氨基酸对睡眠剥夺大鼠血清支链氨基酸/色氨酸比值的影响%Effects of branched-chain amino acids supplementation on serum BCAA/Trp ratio in rats subjected to sleep deprivation

    Institute of Scientific and Technical Information of China (English)

    欧阳儒颖; 季红光; 蔡懿灵; 阮芳铭; 王海明

    2004-01-01

    目的探讨补充支链氨基酸(branched-chain amino acids, BCAA)对睡眠剥夺(sleep deprivation, SD)大鼠血清支链氨基酸/色氨酸(Tryptophan, Trp)比值的影响.方法采用小站台水环境(flower-pot)睡眠剥夺模型对大鼠进行睡眠剥夺.结果与对照组(26.58±2.49,33.75±2.98)相比,睡眠剥夺大鼠血清游离Trp浓度随睡眠剥夺时间的延长而逐渐升高(28.52±2.50~33.44±1.91)(P0.05),BCAA/Trp比值显著降低(31.89±3.53~27.23±1.56)(P<0.05);补充BCAA,可增加睡眠剥夺大鼠血清BCAA水平,提高BCAA/Trp比值(P<0.05).结论补充支链氨基酸可以提高睡眠剥夺大鼠血清BCAA/Trp比值.

  9. Effects of Branched Chain Amino Acids or Fatty Acid Supplementation on in Vitro Fermentation of Corn Straw and Bacterial Diversity%添加支链氨基酸或支链脂肪酸对玉米秸秆体外瘤胃发酵和细菌多样性的影响

    Institute of Scientific and Technical Information of China (English)

    张慧玲; 李林; 陈勇; 许晓莉; 杨玉霞

    2012-01-01

    In vitro culture technique was adopted to study the effects of branched-chain amino acid (BCAA) or branched-chain volatile fatty acid (BCVFA) on fermentation of corn straw. Results showed that the concentration of total volatile fatty acid,acetic acid,propanoic acid,and butyric acid,and the degradation of dry matter (DM) and neutral detergent fiber (NDF) were not affected by 2 mmol/L BCAA or BCVFA supplementation. Compared with the BCVFA,BCAA supplementation tended to increase the concentrations of total volatile fatty acid (TVFA). Supplementation of valine (Val) and isobutyric acid increased the concentrations of isobutyric acid in the ruminal fluid by 2. 85 and 3. 12 fold ( P <0. 05). Addition of leucine (Leu) ,isovaleric acid,isoleucine (He) and 2-methylbutyric acid increased the concentration of isovaleric acid in the ruminal fluid by 2. 20 - 2. 50 fold ( P <0. 05). Compared with the BCVFA,supplementation of Val,Leu and He increased the concentration of valeric acid in ruminal fluid by 27. 69% ( P < 0. 05) ,25. 68% ( P <0. 05) and 39. 34% ( P <0. 05) respectively. Final pH decreased and degradation of DM and NDF of corn straw increased and the concentrations of TVFA,acetic acid,propanoic acid and butyric acid tended to increase by 4 mmol/L BCAA or BCVFA supplementation. Addition of isovaleric acid, He and 2-methylbutyric acid increased the concentrations of propanoic acid by 29. 94% ( P < 0. 05), 26. 31% ( P <0. 05) and 25. 00% ( P'<0. 05) respectively. Supplementation of Val and isobutyric acid increased the concentrations of isobutyric acid in the ruminal fluid by 5. 30 and 5. 45 fold ( P <0. 05). Addition of 4 mmol/L Leu,isovaleric acid,He and 2-methylbutyric acid increased the concentration of isovaleric acid in the ruminal fluid by 3. 76 — 4. 20 fold ( P <0. 05). Compared with the BCVFA, supplementation of 4 mmol/L Val,Leu and He increased the concentration of valeric acid in ruminal fluid by 22. 92% ( P < 0.05), 33. 33% (P<0. 05) and 55. 00% ( P <0. 05

  10. Synthesis of a Hoechst 32258 analogue amino acid building block for direct incorporation of a fluorescent, high-affinity DNA binding motif into peptides

    DEFF Research Database (Denmark)

    Behrens, C; Harrit, N; Nielsen, P E

    2001-01-01

    The synthesis of a new versatile "Hoechst 33258-like" Boc-protected amino acid building block for peptide synthesis is described. It is demonstrated that this new ligand is an effective mimic of Hoechst 33258 in terms of DNA affinity and sequence specificity. Furthermore, this minor groove binder...

  11. Dietary branched-chain amino acid (BCAA) and tumor growth

    Energy Technology Data Exchange (ETDEWEB)

    Chan, W.; Baron, L.; Baron, P.; White, F.; Banks, W.L. Jr.

    1986-03-05

    The effects of high dietary BCAA on tumor growth was examined in adult male Fischer 344 rats inoculated with 10/sup 6/ viable MCA fibrosarcoma cells. Ten days after tumor inoculation, when tumors were of palpable size, rats were divided into two groups at random. The experimental(E) group was fed the AIN-76 diet supplemented with 4X the BCAA content of diet casein and the control(C) group was fed the AIN-76 made isonitrogenous with the E diet by glutamic acid supplementation. Five rats from each group were killed at days 0,3,6, and 9. Rats were injected with /sup 14/C-Tyrosine and /sup 3/H-Thymidine i.p. (2 and 4 uCi/100g BW, respectively) an hour before they were killed. The incorporation of /sup 14/C and /sup 3/H into the acid insoluble fraction of the tumor tissues samples were measured. Single cell suspension of tumor were prepared for cell cycle kinetics analysis using a Coulter EPICS IV flow microflorometer. The percentage of normal and hyperdiploid cells were analyzed. Results showed that both tumor size and weight were doubled at each time point the rats were killed. At day 0, the /sup 3/H and /sup 14/C incorporation were 32 +/- 10dpm and 27 +/- 4dpm/mg tumor, respectively. The /sup 3/H incorporation dropped in both diet groups at days 6 and 9 but the /sup 14/C incorporation showed a decrease only at day 9. These changes were statistically significant, P>0.05. No difference in the tumor growth parameters used in this study can be attributed to the high dietary BCAA.

  12. Mutation of zebrafish dihydrolipoamide branched-chain transacylase E2 results in motor dysfunction and models maple syrup urine disease

    Directory of Open Access Journals (Sweden)

    Timo Friedrich

    2012-03-01

    Analysis of zebrafish mutants that demonstrate abnormal locomotive behavior can elucidate the molecular requirements for neural network function and provide new models of human disease. Here, we show that zebrafish quetschkommode (que mutant larvae exhibit a progressive locomotor defect that culminates in unusual nose-to-tail compressions and an inability to swim. Correspondingly, extracellular peripheral nerve recordings show that que mutants demonstrate abnormal locomotor output to the axial muscles used for swimming. Using positional cloning and candidate gene analysis, we reveal that a point mutation disrupts the gene encoding dihydrolipoamide branched-chain transacylase E2 (Dbt, a component of a mitochondrial enzyme complex, to generate the que phenotype. In humans, mutation of the DBT gene causes maple syrup urine disease (MSUD, a disorder of branched-chain amino acid metabolism that can result in mental retardation, severe dystonia, profound neurological damage and death. que mutants harbor abnormal amino acid levels, similar to MSUD patients and consistent with an error in branched-chain amino acid metabolism. que mutants also contain markedly reduced levels of the neurotransmitter glutamate within the brain and spinal cord, which probably contributes to their abnormal spinal cord locomotor output and aberrant motility behavior, a trait that probably represents severe dystonia in larval zebrafish. Taken together, these data illustrate how defects in branched-chain amino acid metabolism can disrupt nervous system development and/or function, and establish zebrafish que mutants as a model to better understand MSUD.

  13. Lipase-mediated resolution of branched chain fatty acids

    NARCIS (Netherlands)

    Heinsman, N.W.J.T.; Franssen, M.C.R.; Padt, A. van der; Boom, R.M.; Riet, K. van 't; Groot, A.E. de

    2002-01-01

    Branched chain fatty acids (BCFAs) are fatty acids substituted with alkyl groups. Many of them are chiral and therefore occur in two enantiomeric forms. This review describes their occurrence in Nature, their biosynthesis, their properties as flavours, and their enzymatic kinetic resolution. Many li

  14. Lipase-mediated resolution of branched chain fatty acids

    NARCIS (Netherlands)

    Heinsman, N.W.J.T.; Franssen, M.C.R.; Padt, A. van der; Boom, R.M.; Riet, K. van 't; Groot, A.E. de

    2002-01-01

    Branched chain fatty acids (BCFAs) are fatty acids substituted with alkyl groups. Many of them are chiral and therefore occur in two enantiomeric forms. This review describes their occurrence in Nature, their biosynthesis, their properties as flavours, and their enzymatic kinetic resolution. Many li

  15. 高支链氨基酸饮食对乙型肝炎肝硬化患者营养状态及预后的影响%Influence of enriched branched-chain amino acid diet on nutrition condition and prognosis in patients with hepatitis B cirrhosis

    Institute of Scientific and Technical Information of China (English)

    袁伟; 张宇一; 张正国; 邹颖; 王介非; 钱志平

    2014-01-01

    influence of enriched branched-chain amino acid on nutrition condition,liver function and prognosis in patients with hepatitis B cirrhosis.Methods Ninety-two patients with liver cirrhosis were enrolled.The patients were randomly divided into two groups,including 47 cases treated with branched-chain amino acid in treatment group and 45 cases treated with regular dieteric treatment in control group.Evaluations of liver function and nutrition condition were followed up to the 96th week in both groups,as well as observations of incidence of cirrhosis related complications.Results Compared to pre-treatment,Child-Pugh score(7.70±1 .65 vs.7.38±1 .72,t=3.591 , P <0.05)intreatment group was obviously promoted at week 12;Compared to control group at week 48,arm muscle circumference[(24.76±1 .04)cm vs.(24.02±1 .64)cm,t=-2.297,P <0.05],total protein [(63.01 ±3.62 g/L vs. (59.39±4.05 )g/L,t = -4.004,P < 0.01 ],pre-albumin [(123.68 ±34.45 )mg/L vs.(97.03 ±28.60 )mg/L,t =-3.611 ,P <0.05],albumin[(34.05±3.13)g/L vs.(32.44±3.53)g/L,t=-2.057,P <0.05]and prothrombin time activity [(99.70±9.07)% vs.(87.75 ±20.95 )%,t = -3.109,P <0.01 ]were significantly increased in treatment group.During a follow-up 96 weeks,incidences of hepatic encephalopathy and infection in treatment group lower than those in control group (χ2 =3.921 ,P <0.05;χ2 =4.458 P <0.05).There was no significant difference in incidences of hepatorenal syndrome and hemorrhage of digestive tract between two groups.Furthermore,control group showed a poor prognosis (P <0.05).Conclusion Enriched branched-chain amino acid can improve nutrition condition,liver function and prognosis in patients with liver cirrhosis.

  16. BRANCHED-CHAIN AMINO ACIDS SUPPLEMENTATION ENHANCES SWIMMING PERFORMANCE OF TRAINED MICE: EVIDENCE FROM CHANGES OF PLASMA AMMONIA AND SEROTONIN%支链氨基酸增强小鼠的游泳能力对其血氨和5-羟色胺变化的研究

    Institute of Scientific and Technical Information of China (English)

    夏志; 汪清祥; 彭勇; 徐飞

    2012-01-01

    目的 探讨支链氨基酸(Branched-chain amino acids,BCAAs)对训练小鼠抗疲劳运动能力的影响.方法 50只小鼠随机分为实验组(B1、B2、B3、B4)和对照组(C).实验组分别灌胃补充BCAAs 0.45、1.25、3.75和10g/kg.d,对照组灌胃生理盐水.各组小鼠5周递增负荷游泳训练后进行力竭游泳并取材.观察BCAAs对小鼠脑组织5-羟色胺( 5-HT)、血氨(PA)和力竭游泳时间的影响.结果 实验组小鼠血氨含量升高,脑组织匀浆中5 -HT含量下降,但力竭运动游泳时间均显著延长.结果血氨浓度随剂量的增加而增大,C与B1组相比血氨水平无显著性差异,但明显低于B2、B3、B4组(P<0.01);C组小鼠5-羟色胺浓度显著高于实验组(P<0.01),实验组间随着补充剂量的增大5-羟色胺浓度减小;实验组小鼠力竭游泳能力明显强于对照组(P<0.01).实验组间,B2、B3组小鼠力竭时间明显长于B1组(分别为P<0.05,P<0.01),B4组力竭时间与B1、B2、B3组相比则下降.结论 BCAAs 能够对抗运动性疲劳的发生发展,增强运动能力,但具有量效关系.

  17. 支链氨基酸对划船运动员耐力运动后肌肉损伤的保护作用%Effects of branched- chain amino acid supplementation on training- induced muscle damage in rowing athletes

    Institute of Scientific and Technical Information of China (English)

    刘建红; 周志宏; 欧明毫; 王奎; 石幼琪

    2003-01-01

    目的:探讨支链氨基酸( branched- chain amino acid,BCAA)对运动后肌肉损伤的保护作用. 方法: 20名划船运动员用随机数字表法分为对照组( n=10)和实验组( n=10),实验组服用 BCAA,对照组不服用 BCAA,共 4周. 4周后运动员在划船测功仪上模拟 2 km和 5 km比赛训练,测定运动前、运动后即刻和运动后 30 min血肌酸激酶 (CK)、乳酸脱氢酶 (LDH)和谷丙转氨酶 (ALT)活性. 结果:对照组运动员血清 CK(μ mol/s)和 LDH(μ mol/s)活性在运动后即刻 (CK: 5.48± 2.27; LDH: 4.73± 0.93)和运动后 30 min(CK: 5.82± 1.87; LDH: 4.83± 2.38)均增加 , 差异有显著性意义( t=2.89,2.36;P均 < 0.05). BCAA组运动员血清 CK和 LDH活性在运动后即刻( CK: 4.42± 1.55; LDH: 4.00± 0.68)与运动前( CK: 3.85± 1.23; LDH: 3.77± 0.72)相比有增加,但没有显著性意义( t=2.24,2.31;P均 < 0.05),且明显低于对照组在运动后即刻的值,运动后 30 min( CK: 4.68± 2.05; LDH: 4.58± 1.10)与运动前比较则有十分显著性的增加,其中 CK活性也明显低于对照组在运动后 30 min的值. 结论:补充 BCAA能降低运动后血清中 CK和 LDH活性,有效保护肌肉组织,减轻肌肉组织的损伤.

  18. The Observation on Clinical Application of Sugar, Whey Protein, and Branched- Chain Amino Acids Supplement for Athletes%运动员补糖、支链氨基酸和乳清蛋白的临床应用观察

    Institute of Scientific and Technical Information of China (English)

    胡斌; 侯菊花; 张立新

    2012-01-01

    Objective: To observe the supplement of sugar drinks, whey protein, bran6hed - chain amino acids and other substances on the stage of the heavy load training to maintain or improve physical function and fatigue recovery. Methods : Arranged 10 athletes in the 30d Sanda physical training phase of the winter training stage, ac- cording to the nutritional supplement program for energy supplement, Selected the indexes of the output of the total body anaerobic power, maximum power, fatigue index, a ability to pull oxygen paddle time, power, paddle fre- quency, biochemical indicators of blood lactate (LA) blood urea nitrogen (BUN), glucose (GL), creatine ki- nase (CK) as the observation, Alternative 10 teammates as the control group, and observed the different before and after the training. Results: The total output power, maximum power after the training had significantly in- creased (P 〈 0.01 ) than before added of the observed team. Compared with the control group after added the ath- letes fatigue index and the paddle frequency was also improved significantly (P 〈 O. 05), and after -school train- ing blood glucose levels also had significantly increased (P 〈 0.05). The blood BUN and the recovery had signif- icant improvement (P 〈0. 001 ) more than the control groups after added whey protein, branched- chain amino acids. Conclusion: The training period added sugar, whey protein, branched -chain amino acids and substances such as energy, could significantly improve the athlete's aerobic capacity and anaerobic exercise performance, could significantly delayed the occurrence of exercise - induced fatigue and accelerate recovery of the exercise - induced fatigue.%目的:观察补充糖饮料、支链氨基酸和乳清蛋白等能量物质对阶段性大运动量训练维持或提高运动员身体机能和疲劳恢复的影响。方法:根据冬训阶段性训练计划安排,对10名散打运动员在30d的体能训

  19. 支链氨基酸强化的肠内肠外营养对肝硬化大鼠术后肝功能及血浆氨基酸谱的影响%Effects of branched-chain amino acids-enriched early parenteral and enteral nutrition on the liver function and serum aminograms in cirrhotic rats after partial hepatectomy

    Institute of Scientific and Technical Information of China (English)

    赖佳明; 胡文杰; 王恕同; 华赟鹏; 郝元涛; 罗时敏; 赖英荣; 梁力建

    2010-01-01

    amino acids-enriched early parenteral and enteral nutrition on the liver function and serum aminograms in cirrhotic rats after partial hepatectomy. Methods In this prospective randomized controlled study, 24 cirrhotic rats, induced by thioacetamide, were randomized into three groups: enteral nutrition (EN) group, EN + branched-chain amino acid (BCAA) group, and parenteral nutrition (PN) + BCAA group. After receiving partial hepatectomy, rats in all three groups were nutritionally supported with equal amount of calorie and nitrogen contents from the 1st postoperative day ( PO day 1 ) to PO day 5. On PO day 6, parameters including body weight, liver functions, prealbumin, transferring, and serum aminograms were measured or determined, and the level of liver albumin mRNA was detected by reversal transcription-polymerase chain reaction and morphological examinations such as HE staining and immunohistochemical staining, which were assessed by index of Ki67 protein index. Results Body weight was significantly decreased in all three groups on PO day 6 (P <0.05 ). Compared with EN + BCAA group, serum aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase after partial hepatectomy were significantly higher in PN + BCAA group (P <0.05 ). Serum alkaline phosphatase level was significantly higher in PN + BCAA group than in EN group ( P <0. 05). The level of prealbumin was significantly lower in PN + BCAA group when compared with EN group or EN +BCAA group ( both P < 0. 05 ), although no such significant difference was noted in terms of transferrin ( P >0. 05 ). The levels of leucine and isoleucine elevated while those of tyrosine, phenylalanine, arginine and tryptophan declined in PN + BCAA group or EN + BCAA group when compared with EN group ( P < 0. 05 ). Aminograms were not significantly different between EN + BCAA group and PN + BCAA group ( P > 0. 05 ). Levels of total amino acid and aromatic amino acid (AAA) were significantly lower while

  20. The efficacy and safety of a branched-chain amino acids granules in the treatment of cirrhotic hypoproteinemia--A multicenter, randomised, control study%复方支链氨基酸颗粒剂治疗肝硬化低蛋白血症多中心临床观察

    Institute of Scientific and Technical Information of China (English)

    袁耀宗

    2001-01-01

    Objective To investigate the efficacy and safety of a branched-chain oral amino acids granules (BCAA-G) on the cirrhotic hypoproteinemia. Methods 172 patients with cirrhotic hypoproteinemia from 6 hospitals in Shanghai area were studied. BCAA-G and control drug (amino acids capsule) were used respectively for 9 weeks. Symptoms、plasma protein、albumin and Fisher score were evaluated at 4 and 9 weeks to assess the efficacy and safety of BACC-G. Results After 4 and 9 weeks' therapy, symptoms of two groups improved (P< 0.05). Compared with the control group, fatigue was significantly improved (P = 0.03) in BCAA-G group. Plasma protein and albumin increased in BCAA-G group (P = 0.0001). The Fisher score in BCAA-G group improved after 4 weeks ( P = 0. 0005 ) and 9 weeks ( P = 0. 0001 ), while no marked change was found in the control group. The side effect rate was 7.50 % in the BCAA-G group and 3.85 % in the control group, respectively. Conclusion BCAA-G is effective and safe in the treatment of cirrhotic hypoproteinemia.%目的观察口服复方支链氨基酸颗粒剂(商品名力维特,Livact)对肝硬化低蛋白血症的疗效和安全性。方法上海地区6家医院共选择有低蛋白血症的肝硬化患者172例,其中治疗组120例,对照组52例,分别给予复方支链氨基酸颗粒剂和对照药物复合氨基酸胶囊,治疗4周和9周后观察其临床症状、血清总蛋白和白蛋白及Fisher比值的变化,评估其有效性和安全性。结果复方支链氨基酸颗粒剂治疗组和对照组各项症状在治疗后均有显著改善(P<0.05),其中复方支链氨基酸颗粒剂对疲劳感的改善情况优于对照组(P=0.03)。复方支链氨基酸颗粒剂组在治疗4周和9周后,血清总蛋白和白蛋白均显著升高(P均为0.0001)。Fisher比值复方支链氨基酸颗粒剂组治疗4周和9周后较前有显著改变(P分别为0.0005和0.0001),对照组则无明显变化(P=0.75)。治疗过程中,复

  1. 补充支链氨基酸对失眠老年大鼠行为和糖代谢的影响%Effects of branched-chain amino acids supplementation on behaviour and glucose metabolism in aged rats subjected to in- somnia(

    Institute of Scientific and Technical Information of China (English)

    欧阳儒颖; 张济生; 蔡懿灵

    2014-01-01

    Objectives To explore the effects of branched-chain amino acids( BCAA) supplementation on behaviour and glucose metabolism in aged rats subjected to isomnia.Methods Experimental model of isomnia was established by use of sleep deprivation method(flower pot),thirty aged male rats were divided into five groups randomly:C(control)group, 6h(6 hours sleep per day) group, 6hB(6 hours sleep per day and fed fodder enriched with BCAA)group,4h(4 hours sleep per day)group and 4hB group.Re-sults Time of resting on the center lattic,number of make-up and number of excrete were significantly increased in experimental groups compared with C group(PC group>4h group(P0.05).Conpared with C group,the level of FPG in experimential groups were increased apparently (P4h (P<0.05).Conclusions supplementation of BCAA may ameliorates behavior and glucose metabolism in aged rats subjected to isomnia.%目的:探讨补充支链氨基酸( branched-chain amino acids,BCAA)对失眠老年大鼠行为和糖代谢的影响。方法采用小平台水环境的睡眠剥夺方法建立动物失眠模型,30只雄性老年大鼠随机分为对照组( C组)、6h组(每天6小时睡眠组)、6hB组(每天睡眠6小时,进食添加BCAA的饲料),4h组(每天睡眠4小时),4hB组(每天睡眠4小时,进食添加BCAA的饲料)。结果实验组大鼠的修饰行为次数、中央格停留时间、粪便颗粒数均高于C组,差异具有显著性( P<0.05)。且修饰行为次数、中央格停留时间4hB组<4h组(P<0.05);旷场实验得分6h组>C组>4h组,差异具有显著性(P<0.05)。但旷场实验得分、粪便颗粒数6h组与6hB组,4h组与4hB组比较无差异(P>0.05)。与C组比较,实验组的FPG水平显著升高(P<0.05);但4hB组<4h组(P<0.05)。与C组比较4h组和4hB组的血清INS水平显著升高(P<0.05),且4hB组<4h组(P<0.05)。与C组比较,

  2. The Laplace Functional and Moments for Markov Branching Chains in Random Environments

    Institute of Scientific and Technical Information of China (English)

    HU Di-he; ZHANG Shu-lin

    2005-01-01

    The concepts of random Markov matrix, Markov branching chain in random environment (MBCRE) and Laplace functional of Markov branching chain in random environment (LFMBCRE) are introduced. The properties of LFMBCRE and the explicit formulas of moments of MBCRE are given.

  3. Purification, crystallization and preliminary X-ray crystallographic analysis of branched-chain aminotransferase from Deinococcus radiodurans

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Chung-Der; Huang, Tien-Feng [Department of Physics, National Tsing-Hua University, Hsinchu 30013,Taiwan (China); Lin, Chih-Hao [Institute of Biological Chemistry, National Taiwan University, Taipei 110,Taiwan (China); Guan, Hong-Hsiang; Hsieh, Yin-Cheng [Life Science Group, Research Division, National Synchrotron Radiation Research Center, Hsinchu 30076,Taiwan (China); Institute of Bioinformatics and Structural Biology, National Tsing-Hua University, Hsinchu 30013,Taiwan (China); Lin, Yi-Hung; Huang, Yen-Chieh; Liu, Ming-Yih [Life Science Group, Research Division, National Synchrotron Radiation Research Center, Hsinchu 30076,Taiwan (China); Chang, Wen-Chang, E-mail: wchang@ntu.edu.tw [Institute of Biological Chemistry, National Taiwan University, Taipei 110,Taiwan (China); Chen, Chun-Jung, E-mail: wchang@ntu.edu.tw [Department of Physics, National Tsing-Hua University, Hsinchu 30013,Taiwan (China); Life Science Group, Research Division, National Synchrotron Radiation Research Center, Hsinchu 30076,Taiwan (China)

    2007-06-01

    The crystallization of branched-chain aminotransferase from D. radiodurans is described. The branched-chain amino-acid aminotransferase (BCAT), which requires pyridoxal 5′-phosphate (PLP) as a cofactor, is a key enzyme in the biosynthetic pathway of the hydrophobic amino acids leucine, isoleucine and valine. DrBCAT from Deinococcus radiodurans, which has a molecular weight of 40.9 kDa, was crystallized using the hanging-drop vapour-diffusion method. According to X-ray diffraction data to 2.50 Å resolution from a DrBCAT crystal, the crystal belongs to space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 56.37, b = 90.70, c = 155.47 Å. Preliminary analysis indicates the presence of two DrBCAT molecules in the asymmetric unit, with a solvent content of 47.52%.

  4. 支链氨基酸和相关拮抗剂降低大鼠中枢5-羟色胺合成减轻术后疲劳综合征的研究%Synthesis reduction of central neurotransmitter 5-hydroxytryptamine by branched chain amino acid and associated antagonists improves postoperative fatigue syndrome

    Institute of Scientific and Technical Information of China (English)

    董千铜; 章晓东; 余震; 闫竞一; 陈笑雷

    2014-01-01

    ;纹状体中5-HT水平比较,假手术组最低,模型组与干预组差异无统计学意义(P>0.05);模型组下丘脑5-HIAA含量高于氟西汀组(P<0.05),但与BCH组和BCAA组比较,差异无统计学意义(P>0.05)。结论 BCAA以及相关拮抗剂BCH和氟西汀,能降低大鼠神经中枢对色氨酸的摄取,减少大脑5-HT合成,改善大鼠POFS。%Objective To observe the change of postoperative fatigue in rats after the effect of branched chain amino acid (BCAA) and associated antagonists on central neurotransmitter 5-HT metabolic pathway, and to investigate the role of 5-HT in the development of postoperative fatigue syndrome (POFS). Methods Fifty SD rats were randomly divided into sham operation group (C group, n=10), model group (M group, n=10), L-type amino acid transporter inhibitor group (L group, n=10), 5-HT uptake inhibitor group (F group, n=10) and branched chain amino acids (B group, n=10). The rats in the C group and the M group were injected with normal saline, while other three groups were respectively injected with BCH, fluoxetine, BCAA(val∶leucine∶isoleucine=5∶3∶2), on preoperative 1 h, postoperative day 1, 2, 3, 4. The rats, except for those in the C group, underwent resection of 70% of the middle small intestine with end-to-end anastomosis. General status of the rats was observed before and after surgery. Morris water maze test, including the hidden platform test and search space test (detecting the learning ability of rats) and tail suspension test (detecting physical endurance of rats) were used to evaluate the degree of POFS from postoperative day 1 to day 7. Concentration of tryptophan (TRP), 5-HT, 5-hydroxyindoleacetic acid (5-HIAA) in different position of brain (hippocampus, striatum, hypothalamus) of rats were measured by high performance liquid chromatography(HPLC) at postoperative day 8. Results As compared to the M group, other four groups showed better general condition and less fatigue

  5. THE EXISTENCE AND MOMENTS OF CANONICAL BRANCHING CHAIN IN RANDOM ENVIRONMENT

    Institute of Scientific and Technical Information of China (English)

    胡迪鹤

    2004-01-01

    The concepts of branching chain in random environmnet and canonical branching chain in random environment axe introduced. Moreover the existence of these chains is proved. Finally the exact formulas of mathematical expectation and variance of branching chain in random environment axe also given.

  6. 长期口服支链氨基酸改善肝硬化预后及其安全性的Meta分析%Prognosis and safety of long-term oral branched-chain amino acids for cirrhosis: a meta-analysis

    Institute of Scientific and Technical Information of China (English)

    吕彩霞; 刘春亮; 王琦

    2014-01-01

    Objective To evaluate the prognosis and safety of long-term oral Branched-Chain Amino Acids (BCAAs) for patients with cirrhosis.Methods Randomized controlled trials (RCTs) were identified from CBM (between January 1978 and September 2013),CNKI (between January 1979 and September 2013),PubMed (between January 1970 and September 2013),EMBASE (between January 1970 and September 2013),and Cochrane Library (issue 4,2013).Publications of the RCTs on the treatment of cirrhosis with oral BCAAS were included and analyzed according to the criteria of Cocbrane handbook.Results Six RCTs involving 1 047 patients were included.The results showed that oral BCAAs improved the event-free survival when compared with the control group [RR =1.13,95% CI =(1.05,1.23),P =0.001].BCAAs supplements had no effect on mortality or had no definite effect on the deteriorative rate of minimal hepatic encephalopathy (MHE) or quality of life (QOL).As reported in some trials,the main side effects of BCAAs were gastrointestinal symptoms.Conclusions Long-term oral BCAAs may improve event-free survival in cirrhotic patients.However,no definite conclusion can be made without evidences from larger,randomized,double-blind,placebo-controlled,and multicenter trials.%目的 评价长期口服支链氨基酸改善肝硬化预后及其安全性.方法 计算机检索PubMed、CNKI等多个中英文数据库,收集以口服支链氨基酸(BCAAs)为干预措施治疗肝硬化的随机对照试验(RCT),按照Cochrane系统评价方法,由两位研究者独立地对符合纳入标准的试验进行资料提取,并对纳入文献进行质量评价和对提取的有效数据进行Meta分析.结果 有6个RCT,共计1 047例肝硬化患者符合纳入标准.Meta分析结果显示口服BCAAs组的无事件生存率略高于对照组[RR=1.13,95% CI=(1.05,1.23),P=0.001],不能降低肝硬化患者的病死率(P=0.40).BCAAs对于降低轻微肝性脑病患者的恶化率及改善生命质量没有明确结果.口

  7. Metabolite production and kinetics of branched-chain aldehyde oxidation in Staphylococcus xylosus

    DEFF Research Database (Denmark)

    Beck, Hans Christian; Hansen, A.M.; Lauritsen, F.R.

    2002-01-01

    The metabolite production of the gram positive bacterium Staphylococcus xylosus when cultivated in a defined medium containing 18 amino acids, 6 vitamins and 2 purines was characterised. Several compounds not previously reported as metabolites of this organism were identified including 2......,5-methylpyrazine, 2-phenylethylacetate, 2-methyltetrahydrothiophen-3-one, 3-(methylthio)-propanoic acid and 3-(methylthio)-propanal. The organoleptic metabolites derived from branched-chain amino acid catabolism; 2-methylpropanal from valine, 2-methylbutanal from isoleucine and 3-methylbutanal from leucine were...... detected at levels ranging from 0.4 to 2.0μM. The concentrations of the corresponding carboxy acids were 963, 858 and 1486μM respectively. We demonstrated that α-ketoisocaproic acid was biotransformed to 3-methylbutanal which immediately was oxidised into 3-methylbutanoic acid. Kinetic studies...

  8. Infinitely dimensional control Markov branching chains in random environments

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    First of all we introduce the concepts of infinitely dimensional control Markov branching chains in random environments (β-MBCRE) and prove the existence of such chains, then we introduce the concepts of conditional generating functionals and random Markov transition functions of such chains and investigate their branching property. Base on these concepts we calculate the moments of the β-MBCRE and obtain the main results of this paper such as extinction probabilities, polarization and proliferation rate. Finally we discuss the classification ofβ-MBCRE according to the different standards.

  9. Adipose tissue monomethyl branched chain fatty acids and insulin sensitivity: effects of obesity and weight loss

    Science.gov (United States)

    Su, Xiong; Magkos, Faidon; Zhou, Dequan; Eagon, J. Christopher; Fabbrini, Elisa; Okunade, Adewole L.; Klein, Samuel

    2014-01-01

    Objective An increase in circulating branched-chain amino acids (BCAA) is associated with insulin resistance. Adipose tissue is a potentially important site for BCAA metabolism. We evaluated whether monomethyl branched chain fatty acids (mmBCFA) in adipose tissue, which are likely derived from BCAA catabolism, are associated with insulin sensitivity. Design and Methods Insulin-stimulated glucose disposal was determined by using the hyperinsulinemic-euglycemic clamp procedure with stable isotope glucose tracer infusion, in 9 lean and 9 obese subjects, and in a separate group of 9 obese subjects before and 1 year after Roux-en-Y gastric bypass (RYGB) surgery (38% weight loss). Adipose tissue mmBCFA content was measured in tissue biopsies taken in the basal state. Results Total adipose tissue mmBCFA content was ~30% lower in obese than lean subjects (P = 0.02), and increased by ~65% after weight loss in the RYGB group (P = 0.01). Adipose tissue mmBCFA content correlated positively with skeletal muscle insulin sensitivity (R2 = 35%, P = 0.01, n = 18). Conclusions These results demonstrate a novel association between adipose tissue mmBCFA content and obesity-related insulin resistance. Additional studies are needed to determine whether the association between adipose tissue mmBCFA and muscle insulin sensitivity is causal or a simple association. PMID:25328153

  10. Metabolically engineered Saccharomyces cerevisiae for branched-chain ester productions.

    Science.gov (United States)

    Yuan, Jifeng; Mishra, Pranjul; Ching, Chi Bun

    2016-12-10

    Medium branched-chain esters can be used not only as a biofuel but are also useful chemicals with various industrial applications. The development of economically feasible and environment friendly bio-based fuels requires efficient cell factories capable of producing desired products in high yield. Herein, we sought to use a number of strategies to engineer Saccharomyces cerevisiae for high-level production of branched-chain esters. Mitochondrion-based expression of ATF1 gene in a base strain with an overexpressed valine biosynthetic pathway together with expression of mitochondrion-relocalized α-ketoacid decarboxylase (encoded by ARO10) and alcohol dehydrogenase (encoded by ADH7) not only produced isobutyl acetate, but also 3-methyl-1-butyl acetate and 2-methyl-1-butyl acetate. Further segmentation of the downstream esterification step into the cytosol to utilize the cytosolic acetyl-CoA pool for acetyltransferase (ATF)-mediated condensation enabled an additional fold improvement of ester productions. The best titre attained in the present study is 260.2mg/L isobutyl acetate, 296.1mg/L 3-methyl-1-butyl acetate and 289.6mg/L 2-methyl-1-butyl acetate.

  11. Mechanisms of activation of muscle branched-chain alpha-keto acid dehydrogenase during exercise in man

    DEFF Research Database (Denmark)

    Van Hall, Gerrit; MacLean, D A; Saltin, B

    1996-01-01

    1. Exercise leads to activation (dephosphorylation) of the branched-chain alpha-keto acid dehydrogenase (BCKADH). Here we investigate the effect of low pre-exercise muscle glycogen content and of branched-chain amino acid (BCAA) ingestion on the activity of BCKADH at rest and after 90 min of one......-leg knee-extensor exercise at 65% maximal one-leg power output in five subjects. 2. Pre-exercise BCAA ingestion (308 mg BCAAs (kg body wt)-1) caused an increased muscle BCAA uptake, a higher intramuscular BCAA concentration and activation of BCKADH both at rest (9 +/- 1 versus 25 +/- 5% for the control...... and BCAA test, respectively) and after exercise (27 +/- 4 versus 54 +/- 7%). 3. At rest the percentage active BCKADH was not different, 6 +/- 2% versus 5 +/- 1%, in the normal and low glycogen content leg (392 +/- 21 and 147 +/- 34 mumol glycosyl units (g dry muscle)-1, respectively). The post...

  12. Kinetics of thermophilic, anaerobic oxidation of straight and branched chain butyrate and valerate

    DEFF Research Database (Denmark)

    Batstone, Damien J.; Pind, Peter Frode; Angelidaki, Irini

    2003-01-01

    parameter set occupied mutually exclusive parameter spaces, indicating that all were statistically different from each other. However, qualitatively, the influence on model outputs was similar, and the lumped set would be reasonable for mixed acid digestion. The main characteristic not represented by Monod......The degradation kinetics of normal and branched chain butyrate and valerate are important in protein-fed anaerobic systems, as a number of amino acids degrade to these organic acids. Including activated and primary wastewater sludge digesters, the majority of full-scale systems digest feeds...... with a significant or major fraction of COD as protein. This study assesses the validity of using a common kinetic parameter set and biological catalyst to represent butyrate, n-valerate, and i-valerate degradation in dynamic models. The i-valerate degradation stoichiometry in a continuous, mixed population system...

  13. 高支链氨基酸营养支持对肝脏手术患者肝功能保护作用的Meta分析%Meta-analysis of branched chain amino acid-enriched nutrition to improve hepatic function in patients undergoing hepatic operation

    Institute of Scientific and Technical Information of China (English)

    舒晓亮; 康凯; 钟静霞; 姬舒荣; 张勇胜; 胡怀东; 张大志

    2014-01-01

    Objective To perform a meta-analysis of randomized controlled trials (RCTs) assessing the benefit of providing branched chain amino acid (BCAA)-enriched nutrition to improve hepatic function in patients undergoing hepatic operation.Methods The electronic databases of PubMed,Springerlink,the Chinese Biomedical Database (CBM),the Cochrane Library,and the China National Knowledge Infrastructure (CNKI) were searched for relevant RCTs using the following search terms:nutritional support,enteral nutrition,parenteral nutrition,hepatic/liver surgery,liver cirrhosis,cancer,hepatectomy,and liver transplantation.The quality of the retrieved RCTs was assessed according to the scale developed by the Cochrane Collaboration.The meta-analysis was conducted using RevMan software,version 5.2.Results A total of 11 relevant RCTs,representing 510 patients,were included in the meta-analysis.Compared to patients in the control (normal nutrition) group,the patients in the BCAA group experienced an effective improvement in hepatic function,as evidenced by significant decreases in total bilirubin (by 0.07 μmol/L; 95% confidence interval (CI):-0.18 ~ 0.05,P > 0.05].In addition,the BCAA group showed improvements in plasma levels of albumin (weighted mean difference (WMD) =0.07; 95% CI:0.06,0.24,P < 0.05) and alanine aminotransferase (WMD =+5.61; 95% CI:-8.63 ~ 19.86,P > 0.05] but neither of the changes reached the threshold of a statistically significant improvement.The BCAA group did however show significantly lower complication rate after operation (65%,95% CI:0.48,0.87,P < 0.01] and mean duration of hospital stay (4.61 days; 95% CI:-6.61,-2.61,P < 0.01].Conclusion BCAA-enriched nutrition improves hepatic function in patients undergoing hepatic operation,thereby helping to reduce the complication risk,duration of hospital stay,and financial burden.BCAA-enriched nutrition is a safe and effective therapy and further clinical application may be beneficial.%目的

  14. Expression of Mitochondrial Branched-Chain Aminotransferase and α-Keto-Acid Dehydrogenase in Rat Brain: Implications for Neurotransmitter Metabolism

    Directory of Open Access Journals (Sweden)

    Jeffrey Thomas Cole

    2012-05-01

    Full Text Available In the brain, metabolism of the essential branched chain amino acids (BCAAs leucine, isoleucine and valine, is regulated in part by protein synthesis requirements. Excess BCAAs are catabolized or excreted. The first step in BCAA catabolism is catalyzed by the branched chain aminotransferase (BCAT isozymes, mitochondrial BCATm and cytosolic BCATc. A product of this reaction, glutamate, is the major excitatory neurotransmitter and precursor of the major inhibitory neurotransmitter -aminobutyric acid (GABA. The BCATs are thought to participate in an α-keto-acid nitrogen shuttle that provides nitrogen for synthesis of glutamate from -ketoglutarate. The branched-chain α-keto acid dehydrogenase enzyme complex (BCKDC catalyzes the second and first irreversible step in BCAA metabolism, which is oxidative decarboxylation of the branched-chain α-keto acid (BCKA products of the BCAT reaction. Maple Syrup Urine Disease (MSUD results from genetic defects in BCKDC, which leads to accumulation of toxic levels of BCAAs and BCKAs that result in brain swelling. Immunolocalization of BCATm and BCKDC in rats revealed that BCATm is present in astrocytes in white matter and in neuropil, while BCKDC is expressed only in neurons. BCATm appears uniformly distributed in astrocyte cell bodies throughout the brain. The segregation of BCATm to astrocytes and BCKDC to neurons provides further support for the existence of a BCAA-dependent glial-neuronal nitrogen shuttle since the data show that BCKAs produced by glial BCATm must be exported to neurons. Additionally, the neuronal localization of BCKDC suggests that MSUD is a neuronal defect involving insufficient oxidation of BCKAs, with secondary effects extending beyond the neuron.

  15. 谷氨酰胺、EPA、支链氨基酸对食管癌同步放化疗及胃癌术后辅助化疗患者的影响%Impact of glutamine, eicosapntemacnioc acid, branched-chain amino acid supplements on nutritional status and treatment compliance of esophageal cancer patients on concurrent chemoradiotherapy and gastric cancer patients on chemotherapy

    Institute of Scientific and Technical Information of China (English)

    丛明华; 宋晨鑫; 邹宝华; 邓颖冰; 李淑娈; 刘雪辉; 刘微微; 刘金英; 于雷

    2015-01-01

    Objective To explore the effects of glutamine,eicosapntemacnioc acid (EPA) and branched-chain amino acids supplements in esophageal cancer patients on concurrent chemoradiotherapy and gastric cancer patients on chemotherapy.Methods From April 2013 to April 2014,a total of 104 esophageal and gastric carcinoma patients on chemotherapy or concurrent chemoradiotherapy were recruited and randomly divided into experimental and control groups.Both groups received dietary counseling and routine nutritional supports while only experimental group received supplements of glutamine (20 g/d),EPA (3.3 g/d) and branched-chain amino acids (8 g/d).And body compositions,blood indicators,incidence of complications and completion rates of therapy were compared between two groups.Results After treatment,free fat mass and muscle weight increased significantly in experiment group while decreased in control group (P < 0.05).And albumin,red blood cell count,white blood cell count and blood platelet count remained stable in experiment group while declined significantly in control group.During treatment,compared to control group,the incidences of infection-associated complication were lower (6% vs 19%,P < 0.05) and the completion rates of therapy were significantly higher in experiment group (96% vs 83%,P < 0.05).Conclusion Supplements of glutamine,EPA and branched-chain amino acids can help maintain nutrition status,decrease the complications and improve compliance for esophageal cancer patients on concurrent chemo-radiotherapy and gastric cancer patients on postoperative adjuvant chemotherapy.%目的 观察补充谷氨酰胺、二十碳五烯酸(EPA)、支链氨基酸对食管癌同步放化疗及胃癌术后辅助化疗患者的作用.方法 2013年4月至2014年4月因食管癌、胃癌在中国医学科学院肿瘤医院和北京桓兴病区拟行化疗或同步化放疗患者104例入组,数字随机分为试验组和对照组进行营养干预,两组均进行

  16. Improved zeolite regeneration processes for preparing saturated branched-chain fatty acids

    Science.gov (United States)

    Ferrierite zeolite solid is an excellent catalyst for the skeletal isomerization of unsaturated linear-chain fatty acids (i.e., oleic acid) to unsaturated branched-chain fatty acids (i.e., iso-oleic acid) follow by hydrogenation to give saturated branched-chain fatty acids (i.e., isostearic acid). ...

  17. Glycogen depletion and acute branched-chain aminoacids supplementation on submaximal exercise in trained rats : implications on anaplerotic reactions

    OpenAIRE

    2005-01-01

    Resumo: Durante o exercício sub-máximo, a utilização de estoques de glicogênio muscular é de suma importância para possibilitar a continuidade do mesmo. A suplementação de Aminoácidos de Cadeia Ramificada (AACR ou BCAA, do inglês Branched Chain Amino Acids) tem sido experimentada como forma de gerar energia para o músculo nessas condições. Através da utilização dos seus esqueletos de carbono, podem ser gerados intermediários do ciclo dos ácidos tricarboxílicos (CAT ou TCA, do inglês Tricarbox...

  18. Adipose tissue monomethyl branched-chain fatty acids and insulin sensitivity: Effects of obesity and weight loss.

    Science.gov (United States)

    Su, Xiong; Magkos, Faidon; Zhou, Dequan; Eagon, J Christopher; Fabbrini, Elisa; Okunade, Adewole L; Klein, Samuel

    2015-02-01

    An increase in circulating branched-chain amino acids (BCAA) is associated with insulin resistance. Adipose tissue is a potentially important site for BCAA metabolism. It was evaluated whether monomethyl branched-chain fatty acids (mmBCFA) in adipose tissue, which are likely derived from BCAA catabolism, are associated with insulin sensitivity. Insulin-stimulated glucose disposal was determined by using the hyperinsulinemic-euglycemic clamp procedure with stable isotope glucose tracer infusion in nine lean and nine obese subjects, and in a separate group of nine obese subjects before and 1 year after Roux-en-Y gastric bypass (RYGB) surgery (38% weight loss). Adipose tissue mmBCFA content was measured in tissue biopsies taken in the basal state. Total adipose tissue mmBCFA content was ∼30% lower in obese than lean subjects (P=0.02) and increased by ∼65% after weight loss in the RYGB group (P=0.01). Adipose tissue mmBCFA content correlated positively with skeletal muscle insulin sensitivity (R(2) =35%, P=0.01, n=18). These results demonstrate a novel association between adipose tissue mmBCFA content and obesity-related insulin resistance. Additional studies are needed to determine whether the association between adipose tissue mmBCFA and muscle insulin sensitivity is causal or a simple association. © 2014 The Obesity Society.

  19. Novel interstrand communication systems within DNA duplexes based on 1-, 2- and 4-(phenylethynyl)pyrenes attached to 2′-amino-LNA: high-affinity hybridization and fluorescence sensing

    DEFF Research Database (Denmark)

    Astakhova, Irina; Lindegaard, Dorthe; Korshun, Vladimir A.

    2010-01-01

    Functionalisation of 2′-amino-LNA oligonucleotides with 1-, 2- and 4-(phenylethynyl)pyrene fluorophores via a carbonyl linker (PEPyc) resulted in efficient interstrand communication systems in nucleic acid duplexes, providing effective tools for stabilization of nanostructures and fluorescence...

  20. Effect of recombinant human growth hormone combined with branched chain amino acid on the levels of tumor necrosis factor-α and interleukins-10 in patients with hepatic cirrhosis%重组人生长激素联合支链氨基酸对肝硬化患者肿瘤坏死因子-α和白细胞介素-10的影响

    Institute of Scientific and Technical Information of China (English)

    马铭; 袁耀宗; 刘建生; 田怡; 张晓红; 刘海玲

    2015-01-01

    Objective:To investigate the effects of recombinant human growth hormone(r-hGH) combined with branched chain amino acid(BCAA) on the levels of Tumor necrosis factor-α(TNF-α) and interleukins-10(IL-10) in patients with decompensated cirrhosis. Methods:Eighty-two patients with decompensated cirrhosis were randomly divided into the treatment group(42 cases) and control group(40 cases). The treatment group and control group were treated with 5 IU of r-hGH by subcutaneous injection 1 time two days combined with 250 ml of BCAA by intravenous drip once a day for 20 d and 250 ml of BCAA by intravenous drip once a day for 20 d based on protecting liver and support treatment,respectively. The liver function and levels of TNF-α and IL-10 in two groups were observed before and after treatment. Results:The liver function and levels of TNF-α and IL-10 in two groups were improved,the improvement of poor appetite,lower limbs edema,and the levels of alanine aminotransferase,albumin,TNF-α and IL-10 in treatment group were better than those in control group(P < 0. 05 to P < 0. 01). Conclusions:The effects of r-hGH combined with BCAA on the improvement of serum albumin,TNF-α and IL-10 levels are better than those of BCAA alone.%目的::探讨重组人生长激素(r-hGH)联合应用支链氨基酸(BCAA)对肝硬化失代偿期患者肿瘤坏死因子-α(TNF-α)、白细胞介素-10(IL-10)的影响。方法:将82例肝硬化失代偿期患者随机分为治疗组42例和对照组40例。在相同保肝、支持治疗基础上,治疗组应用 r-hGH 5 IU,每2天1次皮下注射,同时联用 BCAA 250 ml,每天1次静脉滴注,连用20 d。对照组应用BCAA 250 ml,静脉滴注,每天1次,连用20 d。观察治疗前和治疗结束后第2天患者肝功能和 TNF-α、IL-10水平变化情况。结果:2种治疗方法对患者肝功能指标、血清 TNF-α和 IL-10水平均有改善,但治疗组在纳差、下肢水肿等症状改善及丙氨酸氨基

  1. [Odd- and branched-chain fatty acids in milk fat--characteristic and health properties].

    Science.gov (United States)

    Adamska, Agata; Rutkowska, Jarosława

    2014-08-22

    This review analyzes the current state of knowledge on odd- and branched-chain fatty acids present in milk fat. Special attention is devoted to the characteristic, synthesis in ruminants, factors affecting their content in milk fat and pro-health properties of these compounds. The group of odd- and branched-chain fatty acids includes mainly saturated fatty acids with one or more methyl branches in the iso or anteiso position. These fatty acids are largely derived from ruminal bacteria and they have been transferred to ruminant tissue (milk and meat). For that reason they have been used as biomarkers of rumen fermentation. Odd- and branched-chain fatty acids are exogenous products for humans, and therefore have specific properties. The results of research from recent decades show that odd- and branched-chain fatty acids have anti-cancer activity. Branched-chain fatty acids may reduce the incidence of necrotizing enterocolitis. Additionally, these compounds have a beneficial effect on proper tissue function and on functioning and development of the infant gut, whereas odd-chain fatty acids are considered as biomarkers of milk fat intake by humans. So far, not all the mechanisms of activity of these compounds are known thoroughly. They should be more carefully studied for application of their biological effects in prevention and treatment.

  2. Dyes with high affinity for polylactide

    Institute of Scientific and Technical Information of China (English)

    Liang He; Shu Fen Zhang; Bing Tao Tang; Li Li Wang; Jin Zong Yang

    2007-01-01

    Attempts were made to develop dyes with high affinity for polylactide as an alternative to the existent commercial disperse dyes.The dyes synthesized according to the affinity concept of dye to polylactide exhibited excellent dyeing properties on polylactide compared with the commercial disperse dyes.

  3. Naturally occurring branched-chain polyamines induce a crosslinked meshwork structure in a giant DNA

    Science.gov (United States)

    Muramatsu, Akira; Shimizu, Yuta; Yoshikawa, Yuko; Fukuda, Wakao; Umezawa, Naoki; Horai, Yuhei; Higuchi, Tsunehiko; Fujiwara, Shinsuke; Imanaka, Tadayuki; Yoshikawa, Kenichi

    2016-12-01

    We studied the effect of branched-chain polyamines on the folding transition of genome-sized DNA molecules in aqueous solution by the use of single-molecule observation with fluorescence microcopy. Detailed morphological features of polyamine/DNA complexes were characterized by atomic force microscopy (AFM). The AFM observations indicated that branched-chain polyamines tend to induce a characteristic change in the higher-order structure of DNA by forming bridges or crosslinks between the segments of a DNA molecule. In contrast, natural linear-chain polyamines cause a parallel alignment between DNA segments. Circular dichroism measurements revealed that branched-chain polyamines induce the A-form in the secondary structure of DNA, while linear-chain polyamines have only a minimum effect. This large difference in the effects of branched- and linear-chain polyamines is discussed in relation to the difference in the manner of binding of these polyamines to negatively charged double-stranded DNA.

  4. THE CONSTRUCTION OF MULTITYPE CANONICAL MARKOV BRANCHING CHAINS IN RANDOM ENVIRONMENTS

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    The investigation for branching processes has a long history by their strong physics background, but only a few authors have investigated the branching processes in random environments. First of all, the author introduces the concepts of the multitype canonical Markov branching chain in random environment (CMBCRE) and multitype Markov branching chain in random environment (MBCRE) and proved that CMBCRE must be MBCRE, and any MBCRE must be equivalent to another CMBCRE in distribution. The main results of this article are the construction of CMBCRE and some of its probability properties.

  5. The Effect of Supplement with Branched -chain Amino Acid, Creatine on Blood Biochemical IndeX of Yili Horse%补喂支链氨基酸﹑肌酸对伊犁马血浆生化指标的影响

    Institute of Scientific and Technical Information of China (English)

    李晓斌; 马军; 聂彪彪; 杨景焘; 乔春江; 杨开伦

    2015-01-01

    Objective] This study aims to determine the effects of the branched —chain supplemented with amino acid (BCAA) and creatine (Cr) on the change of the plasma antioxidant index,hormone level,the concentration of creatine,glucose,lactic acid,uric acid and creatinine in plasma of the Yili horses,thus provi_ding reference data for effective nutritious additives in horse sports training fodder.[Method]12 well —trained male horses of 3 —5 years old (average 4 years old),weight 390 —514kg (average 457 ±50 kg) and the simi_lar performance were selected as the subjects in the experiments.They were divided into 3 groups with four in each and one group was fed with basal diet (the control group).The experimental group one was fed with diet supplemented with BCAA and the experimental group two was fed with diet supplemented with Cr.On the 0 day,1 d,14 d,21 d and 31 d jugular vein blood was collected after feeding 4 hours later,and testing kit was used to measure the related indexes.[Result]The total antioxidant capacity in plasma was increased by the di_et supplemented with creatine after feeding 4 h later,and the difference was significant compared with control group (P ﹤0.05).Especially on 14 d and 31 d it was increased by 21.79% and 25.68%; BCAA reduced MDA content during the test,No significant differences were observed in the concentrations of insulin,glucagon and cortisol hormones ,creatine,glucose,lactic acid,uric acid and creatinine of plasma compared with the con_trol group (P ﹥0.05).[Conclusion]The results indicated that the diet supplemented with BCAA could re_duce lipid peroxidation products MDA concentration; Fodder supplemented with Cr could improve SOD vigor and improve the Yili horse′s serum total antioxidant capacity,but no obvious effects were found in other inde_xes.%【目的】研究补喂支链氨基酸(BCAA)﹑肌酸(Cr)对伊犁马血浆抗氧化指标﹑激素水平﹑肌酸﹑葡萄糖浓度以及乳酸﹑尿酸和肌酐含量的影

  6. Lipase-catalyzed kinetic resolution of branched chain fatty acids and their esters

    NARCIS (Netherlands)

    Heinsman, N.W.J.T.

    2000-01-01

    Flavors and fragrances make an important contribution to the taste and smell of all kinds of food products both as natural occurring components and as additional flavors or fragrances. One of these flavor components is 4-methyloctanoic acid (4-MOA). This branched chain fatty acid contributes to the

  7. A safety assessment of branched chain saturated alcohols when used as fragrance ingredients

    DEFF Research Database (Denmark)

    Belsito, D.; Bickers, D.; Bruze, M.

    2010-01-01

    The Branched Chain Saturated Alcohol (BCSA) group of fragrance ingredients was evaluated for safety. In humans, no evidence of skin irritation was found at concentrations of 2-10%. Undiluted, 11 materials evaluated caused moderate to severe eye irritation. As current end product use levels...

  8. Three new branched chain equations of state based on Wertheim's perturbation theory.

    Science.gov (United States)

    Marshall, Bennett D; Chapman, Walter G

    2013-05-07

    In this work, we present three new branched chain equations of state (EOS) based on Wertheim's perturbation theory. The first represents a slightly approximate general branched chain solution of Wertheim's second order perturbation theory (TPT2) for athermal hard chains, and the second represents the extension of first order perturbation theory with a dimer reference fluid (TPT1-D) to branched athermal hard chain molecules. Each athermal branched chain EOS was shown to give improved results over their linear counterparts when compared to simulation data for branched chain molecules with the branched TPT1-D EOS being the most accurate. Further, it is shown that the branched TPT1-D EOS can be extended to a Lennard-Jones dimer reference system to obtain an equation of state for branched Lennard-Jones chains. The theory is shown to accurately predict the change in phase diagram and vapor pressure which results from branching as compared to experimental data for n-octane and corresponding branched isomers.

  9. Branched-Chain Amino and Keto Acid Biochemistry and Cellular Biology in Central Nervous System Diseases

    Science.gov (United States)

    2009-05-21

    Grape seed proanthocyanidins inhibit the growth of human non-small cell lung cancer xenografts by targeting insulin-like growth factor binding protein...pathways, that is to say the interaction between metabolic and signaling roles in an in vitro model of cancer . BCAAs exclusively undergo irreversible...kinase would decrease the anabolic protein translational pathway to result in a decrease of cancerous phenotypes, such as proliferation, invasion

  10. Biosynthesis of branched-chain amino acids in Schizosaccharomyces pombe: properties of acetohydroxy acid synthetase.

    Science.gov (United States)

    McDonald, R A; Satyanarayana, T; Kaplan, J G

    1973-04-01

    The regulatory properties of acetohydroxy acid synthetase (AHAS), the first enzyme in the biosynthetic pathway to valine and the second in the isoleucine pathway, were investigated in the fission yeast Schizosaccharomyces pombe. The enzyme was partially purified from crude extracts by protamine sulfate treatment, ammonium sulfate fractionation, and gel filtration through Sephadex G-25. AHAS from S. pombe is unique in that its activity shows a single peak around pH 6.5; high sensitivity to feedback inhibition by valine at this pH (K(i) = 0.1 mM) indicates that the enzyme is involved in valine biosynthesis. Pyruvate saturation kinetics of AHAS extracted from cells grown on glycerol as sole carbon and energy source were normal and hyperbolic. In contrast, the enzyme from glucose-grown cells exhibited sigmoidal saturation kinetics, an effect which disappeared when the synthetase from such cells was partially purified. This phenomenon was shown to be due to competition for pyruvate between AHAS and pyruvate decarboxylase; the latter enzyme is present in large amounts in cells fermenting glucose. Valine inhibition is noncompetitive in nature, and this effector exhibits homotropic cooperative effects; isoleucine is a less-potent inhibitor of AHAS activity. Mercurial treatment reversibly desensitized the enzyme to valine inhibition. On the basis of these data, the S. pombe AHAS appears to be an allosteric regulatory enzyme with the properties of a negative V system.

  11. In Vivo Imaging of Branched Chain Amino Acid Metabolism in Prostate Cancer

    Science.gov (United States)

    2014-10-01

    5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) Daniel Spielman Betty Diamond 5d. PROJECT NUMBER 5e. TASK NUMBER E-Mail: spielman@stanford.edu 5f...substrate for monocarboxylate trans- porters such as MCT1, which are readily expressed prostate cancer cell lines and prostate tissue [27–29]. Further

  12. Milk Odd- and Branched-Chain Fatty Acids in Relation to the Rumen Fermentation Pattern

    NARCIS (Netherlands)

    Vlaeminck, B.; Fievez, V.; Tamminga, S.; Dewhurst, R.J.; Vuuren, van A.M.; Brabander, de D.; Demeyer, D.

    2006-01-01

    The objectives of this study were 1) to determine whether a relationship exists between molar proportions of volatile fatty acids in the rumen and milk odd-and branched-chain fatty acid concentrations (i.e., iso C13:0, anteiso C13:0, iso C14:0, C15:0, iso C15:0, anteiso C15:0, iso C16:0, C17:0, iso

  13. Activated-sludge nitrification in the presence of linear and branched-chain alkyl benzene sulfonates.

    Science.gov (United States)

    Baillod, C R; Boyle, W C

    1968-01-01

    The effects of biodegradable linear alkyl benzene sulfonate and branched-chain alkyl benzene sulfonate detergents on activated-sludge nitrification were investigated by administering a synthetic waste containing up to 23 mg of each detergent per liter to eight bench-scale, batch, activated-sludge units. It was found that both detergents tended to promote complete oxidation of ammonia to nitrate, whereas control units produced approximately equal amounts of nitrite and nitrate. Various hypotheses are offered to explain the phenomenon.

  14. KEJADIAN INDEL SIMULTAN PADA INTRON 7 GEN BRANCHED-CHAIN Α-KETOACID DEHYDROGENASE E1A (BCKDHA PADA SAPI MADURA

    Directory of Open Access Journals (Sweden)

    Asri Febriana

    2015-08-01

    Full Text Available Madura cattle is one of the Indonesian local cattle breeds derived from crossing between Zebu cattle (Bos indicus and banteng (Bos javanicus. Branched-chain α-ketoacid dehydrogenase (BCKDH is one of the main enzyme complexes in the inner mitochondrial membrane that metabolizes branched chain amino acid (BCAA, ie valine, leucine, and isoleucine. The diversity of the nucleotide sequences of the genes largely determine the efficiency of enzyme encoded. This paper aimed to determine the nucleotide variation contained in section intron 7, exon 8, and intron 8 genes BCKDHA on Madura cattle. This study was conducted on three Madura cattle that used as bull race (karapan, beauty contest (sonok, and beef cattle. The analysis showed that the variation in intron higher than occurred in the exon. Simultaneous indel found at base position 34 and 68 in sonok cattle. In addition, the C266T variant found in beef cattle. These variants do not cause significant changes in amino acids. There was no specific mutation in intron 7, exon 8, and intron 8 were found in Madura cattle designation. This indicated the absence of differentiation Madura cattle designation of selection pressure of BCKDHA gene.

  15. Development and characterization of high affinity leptins and leptin antagonists.

    Science.gov (United States)

    Shpilman, Michal; Niv-Spector, Leonora; Katz, Meirav; Varol, Chen; Solomon, Gili; Ayalon-Soffer, Michal; Boder, Eric; Halpern, Zamir; Elinav, Eran; Gertler, Arieh

    2011-02-11

    Leptin is a pleiotropic hormone acting both centrally and peripherally. It participates in a variety of biological processes, including energy metabolism, reproduction, and modulation of the immune response. So far, structural elements affecting leptin binding to its receptor remain unknown. We employed random mutagenesis of leptin, followed by selection of high affinity mutants by yeast surface display and discovered that replacing residue Asp-23 with a non-negatively charged amino acid leads to dramatically enhanced affinity of leptin for its soluble receptor. Rational mutagenesis of Asp-23 revealed the D23L substitution to be most effective. Coupling the Asp-23 mutation with alanine mutagenesis of three amino acids (L39A/D40A/F41A) previously reported to convert leptin into antagonist resulted in potent antagonistic activity. These novel superactive mouse and human leptin antagonists (D23L/L39A/D40A/F41A), termed SMLA and SHLA, respectively, exhibited over 60-fold increased binding to leptin receptor and 14-fold higher antagonistic activity in vitro relative to the L39A/D40A/F41A mutants. To prolong and enhance in vivo activity, SMLA and SHLA were monopegylated mainly at the N terminus. Administration of the pegylated SMLA to mice resulted in a remarkably rapid, significant, and reversible 27-fold more potent increase in body weight (as compared with pegylated mouse leptin antagonist), because of increased food consumption. Thus, recognition and mutagenesis of Asp-23 enabled construction of novel compounds that induce potent and reversible central and peripheral leptin deficiency. In addition to enhancing our understanding of leptin interactions with its receptor, these antagonists enable in vivo study of the role of leptin in metabolic and immune processes and hold potential for future therapeutic use in disease pathologies involving leptin.

  16. Development and Characterization of High Affinity Leptins and Leptin Antagonists*

    Science.gov (United States)

    Shpilman, Michal; Niv-Spector, Leonora; Katz, Meirav; Varol, Chen; Solomon, Gili; Ayalon-Soffer, Michal; Boder, Eric; Halpern, Zamir; Elinav, Eran; Gertler, Arieh

    2011-01-01

    Leptin is a pleiotropic hormone acting both centrally and peripherally. It participates in a variety of biological processes, including energy metabolism, reproduction, and modulation of the immune response. So far, structural elements affecting leptin binding to its receptor remain unknown. We employed random mutagenesis of leptin, followed by selection of high affinity mutants by yeast surface display and discovered that replacing residue Asp-23 with a non-negatively charged amino acid leads to dramatically enhanced affinity of leptin for its soluble receptor. Rational mutagenesis of Asp-23 revealed the D23L substitution to be most effective. Coupling the Asp-23 mutation with alanine mutagenesis of three amino acids (L39A/D40A/F41A) previously reported to convert leptin into antagonist resulted in potent antagonistic activity. These novel superactive mouse and human leptin antagonists (D23L/L39A/D40A/F41A), termed SMLA and SHLA, respectively, exhibited over 60-fold increased binding to leptin receptor and 14-fold higher antagonistic activity in vitro relative to the L39A/D40A/F41A mutants. To prolong and enhance in vivo activity, SMLA and SHLA were monopegylated mainly at the N terminus. Administration of the pegylated SMLA to mice resulted in a remarkably rapid, significant, and reversible 27-fold more potent increase in body weight (as compared with pegylated mouse leptin antagonist), because of increased food consumption. Thus, recognition and mutagenesis of Asp-23 enabled construction of novel compounds that induce potent and reversible central and peripheral leptin deficiency. In addition to enhancing our understanding of leptin interactions with its receptor, these antagonists enable in vivo study of the role of leptin in metabolic and immune processes and hold potential for future therapeutic use in disease pathologies involving leptin. PMID:21119198

  17. Differential redox potential between the human cytosolic and mitochondrial branched-chain aminotransferase

    Institute of Scientific and Technical Information of China (English)

    Steven J. Coles; John T. Hancock; Myra E. Conway

    2012-01-01

    The human branched-chain aminotransferase (hBCAT) isoenzymes are CXXC motif redox sensitive homodimers central to glutamate metabolism in the central nervous system.These proteins respond differently to oxidation by H2O2,NO,and S-glutathionylation,suggesting that the redox potential is distinct between isoenzymes.Using various reduced to oxidized glutathione ratios (GSH:GSSG) to alter the redox environment,we demonstrate that hBCATc (cytosolic) has an overall redox potential that is 30 mV lower than hBCATm (mitochondrial).Furthermore,the CXXC motif of hBCATc was estimated to be 80 mV lower,suggesting that hBCATm is more oxidizing in nature.Western blot analysis revealed close correlations between hBCAT S-glutathionylation and the redox status of the assay environment,offering the hBCAT isoenzymes as novel biomarkers for cytosolic and mitochondrial oxidative stress.

  18. A Chiral, Photoluminescent, and Spin-Canted {Cu(I)Re(IV)2}n Branched Chain.

    Science.gov (United States)

    Martínez-Lillo, José; Armentano, Donatella; Fortea-Pérez, Francisco R; Stiriba, Salah-Eddine; De Munno, Giovanni; Lloret, Francesc; Julve, Miguel; Faus, Juan

    2015-05-18

    A new heteroleptic 1D Cu(I)-Re(IV) coordination polymer of the formula {Cu(I)Re(IV)Cl4(μ-Cl)(μ-pyz)[Re(IV)Cl4(μ-bpym)]}n·nMeNO2 (1; pyz = pyrazine, bpym = 2,2'-bipyrimidine) has been prepared through the Cu(I)-mediated self-assembly of two different Re(IV) metalloligands, namely, [ReCl5(pyz)](-) and [ReCl4(bpym)]. 1 consists of chiral branched chains with an overall rack-type architecture displaying photoemission and magnetic ordering. These results constitute a first step toward making new multifunctional magnetic materials based on mixed 3d-5d molecular systems.

  19. Higher order classical density functional theory for branched chains and rings.

    Science.gov (United States)

    Marshall, Bennett D; Chapman, Walter G

    2011-12-22

    We develop a general density functional theory for polyatomic molecules by taking the complete association limit of Wertheim's thermodynamic perturbation theory in a general way. By enforcing the complete association limit at an early point in the derivation and introducing the concept of molecular graphs, a general form for the free energy functional and segment densities are developed for both rigid and semiflexible molecules. The theory is applicable to branched chains, rings, or any other molecular structure. As an example the theory is applied as a second order perturbation theory to the case of molecules with a rigid three segment head and a fully flexible tail in a slit pore where bond angle of the rigid portion is an independent variable.

  20. HIGH AFFINITY ACYLATING ANTAGONISTS FOR MUSCARINIC RECEPTORS

    Science.gov (United States)

    Baumgold, Jesse; Karton, Yishai; Malka, Naftali; Jacobson, Kenneth A.

    2012-01-01

    Summary The muscarinic antagonists pirenzepine and telenzepine were derivitized as alkylamino derivatives at a site on the molecules corresponding to a region of bulk tolerance in receptor binding. The distal primary amino groups were coupled to the cross-linking reagent meta-phenylene diisothiocyanate, resulting in two isothiocyanate derivatives that were found to inhibit muscarinic receptors irreversibly and in a dose-dependent fashion. Preincubation of rat forebrain membranes with an isothiocyanate derivative followed by radioligand binding using [3H]N-methylscopolamine diminished the Bmax value, but did not affect the Kd value. The receptor binding site was not restored upon repeated washing, indicating that irreversible inhibition had occurred. IC50 values for the irreversible inhibition at rat forebrain muscarinic receptors were 0.15 nM and 0.19 nM, for derivatives of pirenzepine and telenzepine, respectively. The isothiocyanate derivative of pirenzepine was non-selective as an irreversible muscarinic inhibitor, and the corresponding derivative prepared from telenzepine was 5-fold selective for forebrain (mainly m1) vs. heart (m2) muscarinic receptors. PMID:1625525

  1. Reactions of a fluorescent ATP analog, 2'-(5-dimethyl-aminonaphthalene-1-sulfonyl) amino-2'-deoxyATP, with E. coli F1-ATPase and its subunits: the roles of the high affinity binding site in the alpha subunit and the low affinity binding site in the beta subunit.

    Science.gov (United States)

    Matsuoka, I; Takeda, K; Futai, M; Tonomura, Y

    1982-11-01

    . The kinetic properties of the fluorescence change of DNS-ATP in the reaction with the reconstituted EF1-ATPase were quite similar to those of native EF1. Most of our findings are consistent with a simple mechanism that the high affinity catalytic site and low affinity regulatory site exist in the alpha subunit and beta subunit, respectively. However, the findings mentioned in (4) suggest that the binding of the alpha and beta subunit, which is mediated by the gamma subunit, induces conformational change(s) in the ATP binding site located probably in the alpha subunit, and that the conformational change(s) is essential to exert the full hydrolyzing activity.

  2. [3H]ATPA: a high affinity ligand for GluR5 kainate receptors.

    Science.gov (United States)

    Hoo, K; Legutko, B; Rizkalla, G; Deverill, M; Hawes, C R; Ellis, G J; Stensbol, T B; Krogsgaard-Larsen, P; Skolnick, P; Bleakman, D

    1999-12-01

    The pharmacological properties of [3H]ATPA ((RS)-2-amino-3(3-hydroxy-5-tert-butylisoxazol-4-yl)propanoic acid) are described. ATPA is a tert-butyl analogue of AMPA (alpha-amino-3-hydroxy-5-methyl-4-isoxazoleproprionic acid) that has been shown to possess high affinity for the GluR5 subunit of kainate receptors. [3H]ATPA exhibits saturable, high affinity binding to membranes expressing human GluR5 (GluR5) kainate receptors (Kd approximately 13 nM). No specific binding was observed in membranes expressing GluR2 and GluR6 receptors. Several compounds known to interact with the GluR5 kainate receptor inhibited [3H]ATPA binding with potencies similar to those obtained for competition of [3H]kainate binding to GluR5. Saturable, high affinity [3H]ATPA binding (Kd approximately 4 nM) was also observed in DRG neuron (DRG) membranes isolated from neonatal rats. The rank order potency of compounds to inhibit [3H]ATPA binding in rat DRG and GluR5 membranes were in agreement. These finding demonstrate that [3H]ATPA can be used as a radioligand to examine the pharmacological properties of GluR5 containing kainate receptors.

  3. Stereoselective synthesis of 2,2-bis(C-branched-chain) glucopyranosid-3-ulose via autoxidation reaction

    Institute of Scientific and Technical Information of China (English)

    LIU Hong-Min; ZHANG Fuyi; TAO Jing-Chao

    2004-01-01

    Many different approaches for synthesis of branched chain sugars have beenestablished,1 because they are very useful intermediates for synthesis of other non-sugar chiralmolecules, and usually occur in nature. Branched chain glycosidulose can be used for construction offive- and six-membered carbocyclic rings to which two chiral carbons of sugar are incorporated byintramolecular aldol condensation and Robinson annulation,2 Therefore they are useful in thesynthesis of natural products which consist of annulated carbohydrates or where a highlyfunctionalised enantiomerically pure cyclopentane or cyclohexane is required. Also, this type ofbranched chain sugar can be considered as the synthons of monoterpenoid natural products of theiridoid class which have the cyclopentan-(c)-pyran structure. In view of the importance of branchedchain glycosiduloses, it is desirable to have a general, convenient methodology to their synthesis.However, none of the literature methods was reported on their synthesis by a nuclephilic addition toa partially protected glycosidulose, due to the fact that these glycosiduloses are very difficult tosynthesize selectively and unstable;3 and what is more, one-step synthesis branched chainglycosidulose using this method is almost impossible.In this paper, we report on a general, convenient method for stereoselective syntheses of2,2-bis(C-branched-chain)glucopyranosid-3-uloses by the new reaction of 1 with various activemethylene compounds. The generality of this method was examined in detail. The optimumtemperature was 18-25℃. The solvent DMF was better than the others. In all cases he yields werehigher than 60%.All the 2,2-bis(C-branched-chain)glucopyranosid-3-uloses were characterized by X-raycrystallographic analyses. In addition, the important iintermediate in this reaction was isolated,which is the product of autoxidation of 1 at C-3 position. Thus the reaction mechanism for thesynthesis of 2,2-bis(C-branched-chain) glucopyranosid-3-uloses

  4. Inactivation of Gram-Positive Bacteria by Novel Phenolic Branched-Chain Fatty Acids.

    Science.gov (United States)

    Fan, Xuetong; Wagner, Karen; Sokorai, Kimberly J B; Ngo, Helen

    2017-01-01

    Novel phenolic branched-chain fatty acids (PBC-FAs) were evaluated for their antimicrobial properties against both gram-positive ( Listeria innocua , Bacillus subtilis , Enterococcus faecium ) and gram-negative ( Escherichia coli , Salmonella Typhimurium, and Pseudomonas tolaasii ) bacteria. In addition, PBC-FA derivatives, such as PBC-FA methyl ester mixture, methyl-branched fatty acid mixtures, and trimethylsilyl-PBC-FA methyl esters, were synthesized to study the structure activity relationship. Results showed that PBC-FAs were a potent antimicrobial against gram-positive bacteria with MICs of 1.8 to 3.6 μg/ml. The compounds were less effective against gram-negative bacteria. Derivatives of PBC-FAs and an equimolar mixture of oleic acid and phenol all had MICs above 233 μg/ml against both gram-positive and gram-negative bacteria. Comparison of antimicrobial activities of the PBC-FAs with those of the derivatives suggests that the carboxylic group in the fatty acid moiety and the hydroxyl group on the phenol moiety were responsible for the antimicrobial efficacy. Growth curves of L. innocua revealed that PBC-FAs prevented bacterial growth, while MBC-FAs only delayed the onset of rapid growth of L. innocua . Our results demonstrated that the novel PBC-FAs have potential for use as antimicrobials against gram-positive bacteria.

  5. Branched chain fatty acid content of United States retail cow's milk and implications for dietary intake.

    Science.gov (United States)

    Ran-Ressler, R R; Sim, D; O'Donnell-Megaro, A M; Bauman, D E; Barbano, D M; Brenna, J T

    2011-07-01

    Branched chain fatty acids (BCFA) have recently been shown to be a major component of the normal human newborn gastrointestinal tract and have long been known to be a component of human milk. Ruminant food products are major sources of fat in the American diet, but there are no studies of milkfat BCFA content in retail milk. We report here the profile and concentrations of BCFA in a representative sampling of retail milk in the 48 contiguous United States (US), and their estimated intake in the American diet. Conventionally produced whole fluid milk samples were obtained from 56 processing plants across the contiguous 48 states. Retail milk samples contain exclusively iso- and anteiso-BCFA with 14-18 carbons. BCFA were 2.05 ± 0.14%, w/w of milkfat fatty acids (mean ± SD), and anteiso-BCFA comprised more than half this total. Based on these data and USDA food availability data, the average per capita BCFA intake of Americans is estimated to be about 220 mg/d from dairy; if current dietary recommendations were followed, BCFA intake would be about 400 mg/d. Adding intake from beef consumption, these estimates rise to approximately 400 and 575 mg/d, respectively. These results indicate that BCFA intake is a substantial fraction of daily fat intake, in amounts exceeding those of many bioactive fatty acids.

  6. Branched Chain Fatty Acid (BCFA) Content of Foods and Estimated Intake in the United States

    Science.gov (United States)

    Bae, SangEun; Lawrence, Peter; Wang, Dong Hao

    2015-01-01

    Branched chain fatty acids (BCFA) are bioactive food components that constitute about 2% of fatty acids in cow’s milk fat. Little systematic information on the BCFA content of other foods is available to estimate dietary intakes. We report BCFA distribution and content of fresh and processed foods representing the major foods of Americans and estimate BCFA intake. BCFA are primarily components of dairy and ruminant foods, and were absent from chicken, pork, and salmon. Dairy and beef delivered most of the 500 mg per day mean intake; in comparison, intake of the widely studied long chain polyunsaturates eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) is estimated to average 100 mg per day. Common adjustments in diet can double BCFA daily intake. The fermented foods sauerkraut and miso had appreciable fractions of BCFA but overall are low fat foods providing very small amounts in the diet, and other fermented foods did not contain BCFA as might have been expected from microbial exposure. These data support the quantitative importance of BCFA delivered primarily from dairy and beef and highlight the need for research into their health effects. PMID:24830474

  7. Metabolic engineering of Saccharomyces cerevisiae for the overproduction of short branched-chain fatty acids.

    Science.gov (United States)

    Yu, Ai-Qun; Juwono, Nina Kurniasih Pratomo; Foo, Jee Loon; Leong, Susanna Su Jan; Chang, Matthew Wook

    2016-03-01

    Short branched-chain fatty acids (SBCFAs, C4-6) are versatile platform intermediates for the production of value-added products in the chemical industry. Currently, SBCFAs are mainly synthesized chemically, which can be costly and may cause environmental pollution. In order to develop an economical and environmentally friendly route for SBCFA production, we engineered Saccharomyces cerevisiae, a model eukaryotic microorganism of industrial significance, for the overproduction of SBCFAs. In particular, we employed a combinatorial metabolic engineering approach to optimize the native Ehrlich pathway in S. cerevisiae. First, chromosome-based combinatorial gene overexpression led to a 28.7-fold increase in the titer of SBCFAs. Second, deletion of key genes in competing pathways improved the production of SBCFAs to 387.4 mg/L, a 31.2-fold increase compared to the wild-type. Third, overexpression of the ATP-binding cassette (ABC) transporter PDR12 increased the secretion of SBCFAs. Taken together, we demonstrated that the combinatorial metabolic engineering approach used in this study effectively improved SBCFA biosynthesis in S. cerevisiae through the incorporation of a chromosome-based combinatorial gene overexpression strategy, elimination of genes in competitive pathways and overexpression of a native transporter. We envision that this strategy could also be applied to the production of other chemicals in S. cerevisiae and may be extended to other microbes for strain improvement.

  8. Production of C4 and C5 branched-chain alcohols by engineered Escherichia. coli.

    Science.gov (United States)

    Chen, Xiaoyan; Xu, Jingliang; Yang, Liu; Yuan, Zhenhong; Xiao, Shiyuan; Zhang, Yu; Liang, Cuiyi; He, Minchao; Guo, Ying

    2015-11-01

    Higher alcohols, longer chain alcohols, contain more than 3 carbon atoms, showed close energy advantages as gasoline, and were considered as the next generation substitution for chemical fuels. Higher alcohol biosynthesis by native microorganisms mainly needs gene expression of heterologous keto acid decarboxylase and alcohol dehydrogenases. In the present study, branched-chain α-keto acid decarboxylase gene from Lactococcus lactis subsp. lactis CICC 6246 (Kivd) and alcohol dehydrogenases gene from Zymomonas mobilis CICC 41465 (AdhB) were transformed into Escherichia coli for higher alcohol production. SDS-PAGE results showed these two proteins were expressed in the recombinant strains. The resulting strain was incubated in LB medium at 37 °C in Erlenmeyer flasks and much more 3-methyl-1-butanol (104 mg/L) than isobutanol (24 mg/L) was produced. However, in 5 g/L glucose-containing medium, the production of two alcohols was similar, 156 and 161 mg/L for C4 (isobutanol) and C5 (3-methyl-1-butanol) alcohol, respectively. Effects of fermentation factors including temperature, glucose content, and α-keto acid on alcohol production were also investigated. The increase of glucose content and the adding of α-keto acids facilitated the production of C4 and C5 alcohols. The enzyme activities of pure Kivd on α-ketoisovalerate and α-ketoisocaproate were 26.77 and 21.24 μmol min(-1) mg(-1), respectively. Due to its ability on decarboxylation of α-ketoisovalerate and α-ketoisocaproate, the recombinant E. coli strain showed potential application on isoamyl alcohol and isobutanol production.

  9. Altered Expression of Human Mitochondrial Branched Chain Aminotransferase in Dementia with Lewy Bodies and Vascular Dementia.

    Science.gov (United States)

    Ashby, Emma L; Kierzkowska, Marta; Hull, Jonathon; Kehoe, Patrick G; Hutson, Susan M; Conway, Myra E

    2017-01-01

    Cytosolic and mitochondrial human branched chain aminotransferase (hBCATc and hBCATm, respectively) play an integral role in brain glutamate metabolism. Regional increased levels of hBCATc in the CA1 and CA4 region of Alzheimer's disease (AD) brain together with increased levels of hBCATm in frontal and temporal cortex of AD brains, suggest a role for these proteins in glutamate excitotoxicity. Glutamate toxicity is a key pathogenic feature of several neurological disorders including epilepsy associated dementia, AD, vascular dementia (VaD) and dementia with Lewy bodies (DLB). To further understand if these increases are specific to AD, the expression profiles of hBCATc and hBCATm were examined in other forms of dementia including DLB and VaD. Similar to AD, levels of hBCATm were significantly increased in the frontal and temporal cortex of VaD cases and in frontal cortex of DLB cases compared to controls, however there were no observed differences in hBCATc between groups in these areas. Moreover, multiple forms of hBCATm were observed that were particular to the disease state relative to matched controls. Real-time PCR revealed similar expression of hBCATm mRNA in frontal and temporal cortex for all cohort comparisons, whereas hBCATc mRNA expression was significantly increased in VaD cases compared to controls. Collectively our results suggest that hBCATm protein expression is significantly increased in the brains of DLB and VaD cases, similar to those reported in AD brain. These findings indicate a more global response to altered glutamate metabolism and suggest common metabolic responses that might reflect shared neurodegenerative mechanisms across several forms of dementia.

  10. High affinity retinoic acid receptor antagonists: analogs of AGN 193109.

    Science.gov (United States)

    Johnson, A T; Wang, L; Gillett, S J; Chandraratna, R A

    1999-02-22

    A series of high affinity retinoic acid receptor (RAR) antagonists were prepared based upon the known antagonist AGN 193109 (2). Introduction of various phenyl groups revealed a preference for substitution at the para-position relative to the meta-site. Antagonists with the highest affinities for the RARs possessed hydrophobic groups, however, the presence of polar functionality was also well tolerated.

  11. High affinity ligands for 'diazepam-insensitive' benzodiazepine receptors.

    Science.gov (United States)

    Wong, G; Skolnick, P

    1992-01-14

    Structurally diverse compounds have been shown to possess high affinities for benzodiazepine receptors in their 'diazepam-sensitive' (DS) conformations. In contrast, only the imidazobenzodiazepinone Ro 15-4513 has been shown to exhibit a high affinity for the 'diazepam-insensitive' (DI) conformation of benzodiazepine receptors. We examined a series of 1,4-diazepines containing one or more annelated ring systems for their affinities at DI and DS benzodiazepine receptors, several 1,4-diazepinone carboxylates including Ro 19-4603, Ro 16-6028 and Ro 15-3505 were found to possess high affinities (Ki approximately 2.6-20 nM) for DI. Nonetheless, among the ligands examined, Ro 15-4513 was the only substance with a DI/DS potency ratio approximately 1; other substances had ratios ranging from 13 to greater than 1000. Ligands with high to moderate affinities at DI were previously classified as partial agonists, antagonists, or partial inverse agonists at DS benzodiazepine receptors, but behaved as 'GABA neutral' (antagonist) substances at DI. The identification of several additional high affinity ligands at DI benzodiazepine receptors may be helpful in elucidating the pharmacological and physiological importance of these sites.

  12. Metabolic engineering of Saccharomyces cerevisiae for production of fatty acid short- and branched-chain alkyl esters biodiesel.

    Science.gov (United States)

    Teo, Wei Suong; Ling, Hua; Yu, Ai-Qun; Chang, Matthew Wook

    2015-01-01

    Biodiesel is a mixture of fatty acid short-chain alkyl esters of different fatty acid carbon chain lengths. However, while fatty acid methyl or ethyl esters are useful biodiesel produced commercially, fatty acid esters with branched-chain alcohol moieties have superior fuel properties. Crucially, this includes improved cold flow characteristics, as one of the major problems associated with biodiesel use is poor low-temperature flow properties. Hence, microbial production as a renewable, nontoxic and scalable method to produce fatty acid esters with branched-chain alcohol moieties from biomass is critical. We engineered Saccharomyces cerevisiae to produce fatty acid short- and branched-chain alkyl esters, including ethyl, isobutyl, isoamyl and active amyl esters using endogenously synthesized fatty acids and alcohols. Two wax ester synthase genes (ws2 and Maqu_0168 from Marinobacter sp.) were cloned and expressed. Both enzymes were found to catalyze the formation of fatty acid esters, with different alcohol preferences. To boost the ability of S. cerevisiae to produce the aforementioned esters, negative regulators of the INO1 gene in phospholipid metabolism, Rpd3 and Opi1, were deleted to increase flux towards fatty acyl-CoAs. In addition, five isobutanol pathway enzymes (Ilv2, Ilv5, Ilv3, Aro10, and Adh7) targeted into the mitochondria were overexpressed to enhance production of alcohol precursors. By combining these engineering strategies with high-cell-density fermentation, over 230 mg/L fatty acid short- and branched-chain alkyl esters were produced, which is the highest titer reported in yeast to date. In this work, we engineered the metabolism of S. cerevisiae to produce biodiesels in the form of fatty acid short- and branched-chain alkyl esters, including ethyl, isobutyl, isoamyl and active amyl esters. To our knowledge, this is the first report of the production of fatty acid isobutyl and active amyl esters in S. cerevisiae. Our findings will be useful for

  13. The influence of environmental parameters on the catabolism of branched-chain amino acids by Staphylococcus xylosus and Staphylococcus carnosus

    DEFF Research Database (Denmark)

    Olesen, Pelle Thonning; Stahnke, Louise Heller

    2004-01-01

    detection (GC/FID). Main volatile catabolic products of leucine, isoleucine and valine were 3-methylbutanoic, 2-methylbutanoic and 2-methylpropanoic acids, respectively. The generation of branched flavour compounds was influenced significantly by most of the investigated environmental parameters...

  14. The influence of precultivation parameters on the catabolism of branched-chain amino acids by Staphylococcus xylosus and Staphylococcus carnosus

    DEFF Research Database (Denmark)

    Olesen, Pelle Thonning; Stahnke, Louise Heller

    2003-01-01

    precultivation parameters were: growth phase, temperature, NaCl concentration and the concentration of leucine, isoleucine and valine (only for S. xylosus). Flavour compounds were sampled by automatic static headspace collection and separated/quantified using gas chromatography/flame ionization detection (GC/FID...

  15. The Relationship between Branched-Chain Amino Acid Related Metabolomic Signature and Insulin Resistance: A Systematic Review

    Directory of Open Access Journals (Sweden)

    Xue Zhao

    2016-01-01

    Full Text Available Recent studies have shown the positive association between increased circulating BCAAs (valine, leucine, and isoleucine and insulin resistance (IR in obese or diabetic patients. However, results seem to be controversial in different races, diets, and distinct tissues. Our aims were to evaluate the relationship between BCAA and IR as well as later diabetes risk and explore the phenotypic and genetic factors influencing BCAA level based on available studies. We performed systematic review, searching MEDLINE, EMASE, ClinicalTrials.gov, the Cochrane Library, and Web of Science from inception to March 2016. After selection, 23 studies including 20,091 participants were included. Based on current evidence, we found that BCAA is a useful biomarker for early detection of IR and later diabetic risk. Factors influencing BCAA level can be divided into four parts: race, gender, dietary patterns, and gene variants. These factors might not only contribute to the elevated BCAA level but also show obvious associations with insulin resistance. Genes related to BCAA catabolism might serve as potential targets for the treatment of IR associated metabolic disorders. Moreover, these factors should be controlled properly during study design and data analysis. In the future, more large-scale studies with elaborate design addressing BCAA and IR are required.

  16. Weight loss and weight maintenance obtained with or without GLP-1 analogue treatment decrease branched chain amino acid levels

    DEFF Research Database (Denmark)

    Engelbrechtsen, Line; Iepsen, Eva Pers Winning; Galijatovic, Ehm Astrid Andersson

    2016-01-01

    -eight obese individuals underwent a diet-induced 12 % body weight loss during 8 weeks. Participants were randomized to weight maintenance with or without administration of the GLP-1 RA liraglutide (1.2 mg/day) for 52 weeks. Metabolomic profiling by high-throughput proton nuclear magnetic resonance...

  17. Effect of lipid supplementation on milk odd- and branched-chain fatty acids in dairy cows.

    Science.gov (United States)

    Baumann, E; Chouinard, P Y; Lebeuf, Y; Rico, D E; Gervais, R

    2016-08-01

    Eight ruminally fistulated, multiparous Holstein cows were arranged in a double 4×4 Latin square with 14-d periods to investigate the effects of lipid supplementation on performance, rumen parameters, the milk odd- and branched-chain fatty acid (OBCFA) profile, and the relationships between milk OBCFA and rumen parameters. Lipid supplementation is known to inhibit microbial growth in the rumen, decrease de novo microbial fatty acid synthesis, and increase the uptake of circulating fatty acids by the mammary gland; treatments were selected to isolate these effects on the milk OBCFA profile. The 4 treatments were (1) a lipid-free emulsion medium infused in the rumen (CTL), (2) soybean oil as a source of polyunsaturated fatty acids infused in the rumen (RSO), (3) saturated fatty acids (38% 16:0, 40% 18:0) infused in the rumen (RSF), and (4) saturated fatty acids infused in the abomasum (ASF). Fat supplements were provided continuously as emulsions at a rate of 450g/d. Preplanned contrasts compared CTL to RSO, RSO to RSF, and RSF to ASF. Infusing RSO slightly decreased ruminal pH, but did not affect volatile fatty acids profile and milk fat concentration as compared with CTL. The yields of energy-corrected milk, fat, and protein were greater with RSF compared with RSO. The concentration of odd-chain fatty acids was decreased by RSO, whereas even-chain iso fatty acids were not affected. Milk fat concentration of 17:0 + cis-9 17:1 was higher for RSF than for RSO, due to the saturated fatty acids supplement containing 2% 17:0 + cis-9 17:1. Limited differences were observed in the milk OBCFA profile between RSF and ASF. A multiple regression analysis yielded the following equation for predicting rumen pH based on milk fatty acids: pH=6.24 - (0.56×4:0) + (1.67 × iso 14:0) + (4.22 × iso 15:0) + (9.41×22:0). Rumen propionate concentration was negatively correlated with milk fat concentration of iso 14:0 and positively correlated with milk 15:0, whereas the acetate

  18. Sequence Classification: 183986 [

    Lifescience Database Archive (English)

    Full Text Available Non-TMB TMH Non-TMB TMB Non-TMB Non-TMB >gi|17988464|ref|NP_541097.1| HIGH-AFFINITY... BRANCHED-CHAIN AMINO ACID TRANSPORT SYSTEM PERMEASE PROTEIN LIVM / HIGH-AFFINITY BRANCHED-CHAIN AMINO ACID

  19. Gamma scintigraphy of {sup 111}In-labelled branched chain polypeptides (BCP) with a poly(L-lysine) backbone in mice with mammary carcinoma. Effect of charge on biodistribution and tumour imaging potential

    Energy Technology Data Exchange (ETDEWEB)

    Pimm, M.V.; Gribben, S.J. [University of Nottingham (United Kingdom); Perkins, A.C.; Mezo, G.; Gaal, D.; Hudecz, F.

    1995-11-01

    Radiolabelled synthetic branched chain polypeptides (BCP) represent a new and novel range of materials with potential as radiopharmaceuticals. Preliminary imaging studies have been undertaken with {sup 111}In-labelled BCP in mice with subcutaneously transplanted mammary carcinoma. Four polypeptides each with a poly(L-lysine) backbone and side chains of DL-alanine residues were studied. These were AK, which is polycationic, EAK which is amphoteric, having additional glutamic acid residues at the end of the side chains, and AcEAK (anionic) and SucEAK (highly polyanionic) where the terminal glutamic acid amino groups were acetylated or succinylated respectively. Radiolabelling was achieved by previous conjugation with DTPA. Serial images up to 48 hours showed marked retention of {sup 111}In-labelled polycationic AK and polyanionic SucEAK in the liver and spleen, with renal uptake also being visible in the case of AK. {sup 111}In-labelled EAK and AcEAK showed longer blood survival with some liver uptake, but tumour uptake was also visualized by 24 hours with both of these polypeptides. These studies demonstrate the feasibility of using {sup 111}In-labelled synthetic branched chain polypeptides as radiopharmaceuticals for gamma scintigraphy and the visualization of tumours by modification of the side chain structure. These materials warrant further study. (author).

  20. Branched chain aldehydes: production and breakdown pathways and relevance for flavour in foods

    NARCIS (Netherlands)

    Smit, B.A.; Engels, W.J.M.; Smit, G.

    2009-01-01

    Branched aldehydes, such as 2-methyl propanal and 2- and 3-methyl butanal, are important flavour compounds in many food products, both fermented and non-fermented (heat-treated) products. The production and degradation of these aldehydes from amino acids is described and reviewed extensively in lite

  1. Peroxisome protein transportation affects metabolism of branched-chain fatty acids that critically impact growth and development of C. elegans.

    Directory of Open Access Journals (Sweden)

    Rencheng Wang

    Full Text Available The impact of specific lipid molecules, including fatty acid variants, on cellular and developmental regulation is an important research subject that remains under studied. Monomethyl branched-chain fatty acids (mmBCFAs are commonly present in multiple organisms including mammals, however our understanding of mmBCFA functions is very limited. C. elegans has been the premier model system to study the functions of mmBCFAs and their derived lipids, as mmBCFAs have been shown to play essential roles in post-embryonic development in this organism. To understand more about the metabolism of mmBCFAs in C. elegans, we performed a genetic screen for suppressors of the L1 developmental arrest phenotype caused by mmBCFA depletion. Extensive characterization of one suppressor mutation identified prx-5, which encodes an ortholog of the human receptor for the type-1 peroxisomal targeting signal protein. Our study showed that inactivating prx-5 function compromised the peroxisome protein import, resulting in an increased level of branched-chain fatty acid C17ISO in animals lacking normal mmBCFA synthesis, thereby restoring wild-type growth and development. This work reveals a novel connection between peroxisomal functions and mmBCFA metabolism.

  2. AGP2 encodes the major permease for high affinity polyamine import in Saccharomyces cerevisiae.

    Science.gov (United States)

    Aouida, Mustapha; Leduc, Anick; Poulin, Richard; Ramotar, Dindial

    2005-06-24

    Polyamines play essential functions in many aspects of cell biology. Plasma membrane transport systems for the specific uptake of polyamines exist in most eukaryotic cells but have been very recently identified at the molecular level only in the parasite Leishmania. We now report that the high affinity polyamine permease in Saccharomyces cerevisiae is identical to Agp2p, a member of the yeast amino acid transporter family that was previously identified as a carnitine transporter. Deletion of AGP2 dramatically reduces the initial velocity of spermidine and putrescine uptake and confers strong resistance to the toxicity of exogenous polyamines, and transformation with an AGP2 expression vector restored polyamine transport in agp2delta mutants. Yeast mutants deficient in polyamine biosynthesis required >10-fold higher concentrations of exogenous putrescine to restore cell proliferation upon deletion of the AGP2 gene. Disruption of END3, a gene required for an early step of endocytosis, increased the abundance of Agp2p, an effect that was paralleled by a marked up-regulation of spermidine transport velocity. Thus, AGP2 encodes the first eukaryotic permease that preferentially uses spermidine over putrescine as a high affinity substrate and plays a central role in the uptake of polyamines in yeast.

  3. 01-ERD-111 - The Development of Synthetic High Affinity Ligands

    Energy Technology Data Exchange (ETDEWEB)

    Perkins, J; Balhorn, R; Cosman, M; Lightstone, F; Zeller, L

    2004-02-05

    The aim of this project was to develop Synthetic High-Affinity Ligands (SHALs), which bind with high affinity and specificity to proteins of interest for national security and cancer therapy applications. The aim of producing synthetic ligands for sensory devices as an alternative to antibody-based detection assays and therapeutic agents is to overcome the drawbacks associated with antibody-based in next-generation sensors and systems. The focus area of the project was the chemical synthesis of the SHALs. The project concentrated on two different protein targets. (a) The C fragment of tetanus and botulinum toxin, potential biowarfare agents. A SHAL for tetanus or botulinum toxin would be incorporated into a sensory device for the toxins. (b) HLA-DR10, a protein found in high abundance on the surface of Non-Hodgkins Lymphoma. A SHAL specific to a tumor marker, labeled with a radionuclide, would enable the targeted delivery of radiation therapy to metastatic disease. The technical approach used to develop a SHAL for each protein target will be described in more detail below. However, in general, the development of a SHAL requires a combination of computational modeling techniques, modern nuclear magnetic resonance spectroscopy (NMR) and synthetic chemistry.

  4. Short branched-chain C6 carboxylic acids result in increased growth, novel 'unnatural' fatty acids and increased membrane fluidity in a Listeria monocytogenes branched-chain fatty acid-deficient mutant.

    Science.gov (United States)

    Sen, Suranjana; Sirobhushanam, Sirisha; Hantak, Michael P; Lawrence, Peter; Brenna, J Thomas; Gatto, Craig; Wilkinson, Brian J

    2015-10-01

    Listeria monocytogenes is a psychrotolerant food borne pathogen, responsible for the high fatality disease listeriosis, and expensive food product recalls. Branched-chain fatty acids (BCFAs) of the membrane play a critical role in providing appropriate membrane fluidity and optimum membrane biophysics. The fatty acid composition of a BCFA-deficient mutant is characterized by high amounts of straight-chain fatty acids and even-numbered iso fatty acids, in contrast to the parent strain where odd-numbered anteiso fatty acids predominate. The presence of 2-methylbutyrate (C5) stimulated growth of the mutant at 37°C and restored growth at 10°C along with the content of odd-numbered anteiso fatty acids. The C6 branched-chain carboxylic acids 2-ethylbutyrate and 2-methylpentanoate also stimulated growth to a similar extent as 2-methylbutyrate. However, 3-methylpentanoate was ineffective in rescuing growth. 2-Ethylbutyrate and 2-methylpentanoate led to novel major fatty acids in the lipid profile of the membrane that were identified as 12-ethyltetradecanoic acid and 12-methylpentadecanoic acid respectively. Membrane anisotropy studies indicated that growth of strain MOR401 in the presence of these precursors increased its membrane fluidity to levels of the wild type. Cells supplemented with 2-methylpentanoate or 2-ethylbutyrate at 10°C shortened the chain length of novel fatty acids, thus showing homeoviscous adaptation. These experiments use the mutant as a tool to modulate the membrane fatty acid compositions through synthetic precursor supplementation, and show how existing enzymes in L. monocytogenes adapt to exhibit non-native activity yielding unique 'unnatural' fatty acid molecules, which nevertheless possess the correct biophysical properties for proper membrane function in the BCFA-deficient mutant.

  5. Evidence for a precursor of the high-affinity metastasis-associated murine laminin receptor

    DEFF Research Database (Denmark)

    Rao, C N; Castronovo, V; Schmitt, M C;

    1989-01-01

    The high-affinity cellular receptor for the basement membrane component laminin is differentially expressed during tumor invasion and metastasis. A cDNA clone encoding the murine laminin receptor was isolated and identified on the basis of sequence homology to the human laminin receptor [Wewer et...... al. (1986) Proc. Natl. Acad. Sci. U.S.A. 83, 7137-7141]. Primer extension experiments demonstrated that the clone contained the complete 5' sequence of the murine laminin receptor mRNA. RNA blot data demonstrated a single-sized laminin receptor mRNA, approximately 1400 bases long, in human, mouse......, and rat. The nascent laminin receptor predicted from the cDNA sequence is 295 amino acids long, with a molecular weight of 33,000, and contains one intradisulfide bridge, a short putative transmembrane domain, and an extracellular carboxy-terminal region which has abundant glutamic acid residues...

  6. PREPARATION OF IMMUNOGEN AND PURIFICA¬TION OF HIGH AFFINITY AND SPECIFICITY FAB FRAGMENT OF ANTI-DIGOXIN POLYCLONAL ANTIBODIES

    Directory of Open Access Journals (Sweden)

    M. Pour-Amir

    2000-01-01

    Full Text Available In this study we produced and purified a high titer of specific and high affin¬ity Fab fragments of anti-digoxin antibody. Immunization of rabbits with a conju¬gate of the cardiac glycoside digoxin, coupled by a periodate oxidation method to the amino group of lysine in bovine serum albumin resulted in the production of this type of high titer digoxin-specific antibodies with exceptionally high affinity (109 L/mol and specificity in immune response. Increase in titer was found in steps of purification ending up with the highest titer for Fab fragment to be at 1.75 ug of purified Fab (for 50% binding of I25I-digoxin. High specificity for antigenic determinants of the steroid nucleus of digoxin was observed such that much less cross-reaction with digoxin (2.3% and no cross-reaction with ouabaine, estradiol, Cortisol, progesterone and testosterone were detected.

  7. The high-affinity immunoglobulin E receptor as pharmacological target.

    Science.gov (United States)

    Blank, Ulrich; Charles, Nicolas; Benhamou, Marc

    2016-05-05

    The high-affinity receptor for immunoglobulin E is expressed mainly on mast cells and basophils, but also on neutrophils, eosinophils, platelets, monocytes, Langerhans and dendritic cells, airway smooth muscle cells and some nerve cells. Its main function is, upon its engagement by IgE and specific antigen, to trigger a powerful defense against invading pathogens and a rapid neutralization of dangerous toxic substances introduced in the body. This powerful response could be wielded against tumors. But, when control over this receptor is lost, its unchecked activation can induce an array of diseases, some of which can lead to death. In this review we will summarize the pharmacological approaches and strategies that are currently used, or under study, to harness or wield activation of this receptor for therapeutic purposes.

  8. Structural characterization of saturated branched chain fatty acid methyl esters by collisional dissociation of molecular ions generated by electron ionization.

    Science.gov (United States)

    Ran-Ressler, Rinat R; Lawrence, Peter; Brenna, J Thomas

    2012-01-01

    Saturated branched chain fatty acids (BCFA) are present as complex mixtures in numerous biological samples. The traditional method for structure elucidation, electron ionization (EI) mass spectrometry, sometimes does not unambiguously enable assignment of branching in isomeric BCFA. Zirrolli and Murphy (Zirrolli , J. A. , and R. A. Murphy. 1993. Low-energy tandem mass spectrometry of the molecular ion derived from fatty acid methyl esters: a novel method for analysis of branched-chain fatty acids. J. Am. Soc. Mass Spectrom. 4: 223-229.) showed that the molecular ions of four BCFA methyl ester (BCFAME) yield highly characteristic fragments upon collisional dissociation using a triple quadrupole instrument. Here, we confirm and extend these results by analysis using a tabletop 3-D ion trap for activated molecular ion EI-MS/MS to 30 BCFAME. iso-BCFAME produces a prominent ion (30-100% of base peak) for [M-43] (M-C₃H₇), corresponding to the terminal isopropyl moiety in the original iso-BCFAME. Anteiso-FAME yield prominent ions (20-100% of base peak) corresponding to losses on both side of the methyl branch, [M-29] and [M-57], and tend to produce more prominent m/z 115 peaks corresponding to a cyclization product around the ester. Dimethyl and tetramethyl FAME, with branches separated by at least one methylene group, yield fragment on both sides of the sites of methyl branches that are more than 6 C away from the carboxyl carbon. EI-MS/MS yields uniquely specific ions that enable highly confident structural identification and quantification of BCFAME.

  9. Peptide array-based characterization and design of ZnO-high affinity peptides.

    Science.gov (United States)

    Okochi, Mina; Sugita, Tomoya; Furusawa, Seiji; Umetsu, Mitsuo; Adschiri, Tadafumi; Honda, Hiroyuki

    2010-08-15

    Peptides with both an affinity for ZnO and the ability to generate ZnO nanoparticles have attracted attention for the self-assembly and templating of nanoscale building blocks under ambient conditions with compositional uniformity. In this study, we have analyzed the specific binding sites of the ZnO-binding peptide, EAHVMHKVAPRP, which was identified using a phage display peptide library. The peptide binding assay against ZnO nanoparticles was performed using peptides synthesized on a cellulose membrane using the spot method. Using randomized rotation of amino acids in the ZnO-binding peptide, 125 spot-synthesized peptides were assayed. The peptide binding activity against ZnO nanoparticles varied greatly. This indicates that ZnO binding does not depend on total hydrophobicity or other physical parameters of these peptides, but rather that ZnO recognizes the specific amino acid alignment of these peptides. In addition, several peptides were found to show higher binding ability compared with that of the original peptides. Identification of important binding sites in the EAHVMHKVAPRP peptide was investigated by shortened, stepwise sequence from both termini. Interestingly, two ZnO-binding sites were found as 6-mer peptides: HVMHKV and HKVAPR. The peptides identified by amino acid substitution of HKVAPR were found to show high affinity and specificity for ZnO nanoparticles.

  10. Detection of Waterborne Viruses Using High Affinity Molecularly Imprinted Polymers.

    Science.gov (United States)

    Altintas, Zeynep; Gittens, Micah; Guerreiro, Antonio; Thompson, Katy-Anne; Walker, Jimmy; Piletsky, Sergey; Tothill, Ibtisam E

    2015-07-07

    Molecularly imprinted polymers (MIPs) are artificial receptor ligands which can recognize and specifically bind to a target molecule. They are more resistant to chemical and biological damage and inactivation than antibodies. Therefore, target specific-MIP nanoparticles are aimed to develop and implemented to biosensors for the detection of biological toxic agents such as viruses, bacteria, and fungi toxins that cause many diseases and death due to the environmental contamination. For the first time, a molecularly imprinted polymer (MIP) targeting the bacteriophage MS2 as the template was investigated using a novel solid-phase synthesis method to obtain the artificial affinity ligand for the detection and removal of waterborne viruses through optical-based sensors. A high affinity between the artificial ligand and the target was found, and a regenerative MIP-based virus detection assay was successfully developed using a new surface plasmon resonance (SPR)-biosensor which provides an alternative technology for the specific detection and removal of waterborne viruses that lead to high disease and death rates all over the world.

  11. Production and Identification of High Affinity Monoclonal Antibodies Against Pesticide Carbofuran

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    To produce high-affinity monoclonal antibodies against pesticide carbofuran, and the develop immunochemical assays for people's health and environmental protection, the hapten 4-[[(2,3-dihydro-2,2-dimethyl-7-benzofuranyloxy) carbonyl]-amino]-butanoic acid (BFNB) of carbofuran was synthesized and Balb/c mice were immunized by the hapten-carrier (BFNB-bovine serum albumin, BFNB-BSA) conjugates. The splenocytes of immunized mice were fused with Sp2/0 cells and the cultural supernatants of hybridoma cells were screened by the indirect enzyme-linked immunoabsorbent assay (ELISA), based on BFNB-ovoalbumin conjugates (BFNB-OVA). Purified monoclonal antibody (McAb) was obtained from fluids of ascites, deposited by octanoic acid and ammonium sulfate. The affinity and the specificity of McAb were characterized by ELISA or indirect competitive ELISA. A hybridoma cell line (5D3) secreting anti-carbofuran McAb had been established. The titer of culture medium and ascites was up to 1:2.048 × 103 and 1:1.024 × 106, respectively, and the subtype of the McAb was IgG1. The affinity constant of the McAb was about 2.54 × 109 L mol-1, with an IC50 value of 1.18 ng mL-1 and a detection limit of 0.01 ng mL-1. Cross-reactivity studies showed that the McAb was quiet specific for carbofuran, as among the four analogous compounds, they were all hardly recognized (4.59 × 10-4% for 2,3-dihydro-2,2-dimethyl-7-benzofuranol and less than 3.0 × 10-4% for others). The prepared McAb had a very high affinity and specificity,and it could be used to develop ELISA for rapid determination of carbofuran.

  12. Analysis of high affinity self-association by fluorescence optical sedimentation velocity analytical ultracentrifugation of labeled proteins: opportunities and limitations.

    Directory of Open Access Journals (Sweden)

    Huaying Zhao

    Full Text Available Sedimentation velocity analytical ultracentrifugation (SV is a powerful first-principle technique for the study of protein interactions, and allows a rigorous characterization of binding stoichiometry and affinities. A recently introduced commercial fluorescence optical detection system (FDS permits analysis of high-affinity interactions by SV. However, for most proteins the attachment of an extrinsic fluorophore is an essential prerequisite for analysis by FDS-SV. Using the glutamate receptor GluA2 amino terminal domain as a model system for high-affinity homo-dimerization, we demonstrate how the experimental design and choice of fluorescent label can impact both the observed binding constants as well as the derived hydrodynamic parameter estimates for the monomer and dimer species. Specifically, FAM (5,6-carboxyfluorescein was found to create different populations of artificially high-affinity and low-affinity dimers, as indicated by both FDS-SV and the kinetics of dimer dissociation studied using a bench-top fluorescence spectrometer and Förster Resonance Energy Transfer. By contrast, Dylight488 labeled GluA2, as well as GluA2 expressed as an EGFP fusion protein, yielded results consistent with estimates for unlabeled GluA2. Our study suggests considerations for the choice of labeling strategies, and highlights experimental designs that exploit specific opportunities of FDS-SV for improving the reliability of the binding isotherm analysis of interacting systems.

  13. Rumen odd and branched chain fatty acids in relation to in vitro rumen volatile fatty acid productions and dietary characteristics of incubated substrates

    NARCIS (Netherlands)

    Vlaeminck, B.; Fievez, V.; Laar, van H.; Demeyer, D.

    2004-01-01

    A first aim of this batch in vitro experiment (21 h) was to use changes in odd and branched chain fatty acid (OBCFA) patterns to suggest shifts in microbial populations, associated with four types of incubated whole dairy cow diets. Principal component analysis suggested higher dietary starch increa

  14. cDNA heterogeneity suggests structural variants related to the high-affinity IgE receptor.

    OpenAIRE

    Liu, F T; Albrandt, K; Robertson, M W

    1988-01-01

    The high-affinity IgE receptor present on mast cells and basophils is responsible for the IgE-mediated activation of these cells. The current model for this receptor depicts a four-subunit structure, alpha beta gamma 2. A cDNA for the alpha subunit was recently cloned and predicts a structure consisting of two homologous extracellular domains, a transmembrane segment, and a cytoplasmic tail. Using a synthetic oligonucleotide corresponding to the amino-terminal sequence of the alpha subunit, w...

  15. Amino acids as regulators of gene expression

    Directory of Open Access Journals (Sweden)

    Kimball SR

    2004-08-01

    Full Text Available The role of amino acids as substrates for protein synthesis is well documented. However, a function for amino acids in modulating the signal transduction pathways that regulate mRNA translation has only recently been described. Interesting, some of the signaling pathways regulated by amino acids overlap with those classically associated with the cellular response to hormones such as insulin and insulin-like growth factors. The focus of this review is on the signaling pathways regulated by amino acids, with a particular emphasis on the branched-chain amino acid leucine, and the steps in mRNA translation controlled by the signaling pathways.

  16. A high-affinity molybdate transporter in eukaryotes.

    Science.gov (United States)

    Tejada-Jiménez, Manuel; Llamas, Angel; Sanz-Luque, Emanuel; Galván, Aurora; Fernández, Emilio

    2007-12-11

    Molybdenum is an essential element for almost all living beings, which, in the form of a molybdopterin-cofactor, participates in the active site of enzymes involved in key reactions of carbon, nitrogen, and sulfur metabolism. This metal is taken up by cells in form of the oxyanion molybdate. Bacteria acquire molybdate by an ATP-binding-cassette (ABC) transport system in a widely studied process, but how eukaryotic cells take up molybdenum is unknown because molybdate transporters have not been identified so far. Here, we report a eukaryotic high-affinity molybdate transporter, encoded by the green alga Chlamydomonas reinhardtii gene MoT1. An antisense RNA strategy over the MoT1 gene showed that interference of the expression of this gene leads to the inhibition of molybdate transport activity and, in turn, of the Mo-containing enzyme nitrate reductase, indicating a function of MoT1 in molybdate transport. MOT1 functionality was also shown by heterologous expression in Saccharomyces cerevisiae. Molybdate uptake mediated by MOT1 showed a K(m) of approximately 6 nM, which is the range of the lowest K(m) values reported and was activated in the presence of nitrate. Analysis of deduced sequence from the putative protein coded by MoT1 showed motifs specifically conserved in similar proteins present in the databases, and defines a family of membrane proteins in both eukaryotes and prokaryotes probably involved in molybdate transport and distantly related to plant sulfate transporters SULTR. These findings represent an important step in the understanding of molybdate transport, a crucial process in eukaryotic cells.

  17. Branched Chain Fatty Acids (BCFA) in the neonatal gut, and estimated dietary intake in infancy and adulthood

    Science.gov (United States)

    Ran-Ressler, Rinat R.; Glahn, Raymond P; Bae, SangEun; Brenna, J. Thomas

    2014-01-01

    Branched chain fatty acids (BCFA) are primarily saturated fatty acids (FA) with a methyl branch, usually near the terminal methyl group. BCFA are abundant in bacteria, skin, and vernix caseosa but have seldom been studied with respect to human nutrition. They are constituents of the term newborn infant gut lumen, being swallowed as vernix particulate components of amniotic fluid in the last trimester of normal pregnancy. We recently showed that BCFA protect against Necrotizing Enterocolitis (NEC) in the rat pup model. Dietary BCFA at levels similar to those found in human vernix reduced NEC incidence by more than 50%, increased the abundance of BCFA-containing bacteria, and increased the expression of ileal anti-inflammatory IL-10. The few published reports of BCFA in human milk enable an estimate that breast fed infants consume 19 mg BCFA per 100ml milk. Dietary BCFA consumption from milkfat and other ruminant products, the main sources of dietary BCFA, is more than 400 mg BCFA per day in adult Americans. This estimate exceeds by several fold the average dietary intake of bioactive FA, such as docosahexaenoic acid. BCFA are bioactive, abundant but neglected components of the human food supply. PMID:24107503

  18. The relationship between odd- and branched-chain fatty acids and microbial nucleic acid bases in rumen.

    Science.gov (United States)

    Liu, Keyuan; Hao, Xiaoyan; Li, Yang; Luo, Guobin; Zhang, Yonggen; Xin, Hangshu

    2017-06-26

    This study aims to identify the relationship between odd- and branched-chain fatty acids (OBCFAs) and microbial nucleic acid base in the rumen, and to establish a model to accurately predict microbial protein flow by using OBCFA. To develop the regression equations, data on the rumen contents of individual cows was obtained from 2 feeding experiments. In the first experiment, 3 rumen-fistulated dry dairy cows arranged in a 3×3 Latin square were fed diets of differing forage to concentration ratios (F:C). The second experiment consisted of 9 lactating Holstein dairy cows of similar body weights at the same fetal time. For each milk stage, 3 cows with similar milk production were selected. The rumen contents were sampled at 4 time points of every two hours after morning feeding 6 h, and then to analyse the concentrations of OBCFA and microbial nucleic acid bases in the rumen samples. The ruminal bacteria nucleic acid bases were significantly influenced by feeding diets of differing forge to concentration ratios and milk stages of dairy (P acids, especially odd-chain fatty acids and C15:0 isomers, strongly correlated with the microbial nucleic acid bases in the rumen (P acid bases which established by ruminal OBCFAs contents showed a good a good predictive capacity, as indicated by reasonably low SEs and high R-squared values. This finding suggests that the rumen OBCFA composition could be used as an internal marker of rumen microbial matter.

  19. Effect of whey protein on plasma amino acids in diabetic mice.

    Science.gov (United States)

    Han, Ting; Cai, Donglian; Geng, Shanshan; Wang, Ying; Zhen, Hui; Wu, Peiying

    2013-12-01

    The aim of this study was to investigate the effect of whey protein on plasma amino acid levels in a mouse model of type II diabetes, using high-performance liquid chromatography (HPLC). The composition and content of amino acids in the whey proteins were analyzed using HPLC. Type I and type II diabetic mouse models were prepared using streptozotocin (STZ) and normal mice were used as a control. The ICR mice in each group were then randomly divided into four subgroups, to which 0, 10, 20 and 40% whey protein, respectively, was administered for four weeks. Changes in the plasma amino acid levels were observed in each group. The proportions of leucine, isoleucine and valine in the whey proteins were 14.40, 5.93 and 5.32% of the total amino acids, respectively, that is, the branched-chain amino acid content was 25.65%. The levels of branched-chain amino acids increased in the plasma of the normal and model mice following the administration of whey proteins by gavage and the amino acid levels increased as the concentration of the administered protein increased. In addition, the branched-chain amino acid levels in the blood of the model mice were higher than those in the normal mice. The levels of plasma amino acids in diabetic mice increased following gavage with whey protein, which is rich in branched-chain amino acids.

  20. High Affinity Binding of Indium and Ruthenium Ions by Gastrins.

    Directory of Open Access Journals (Sweden)

    Graham S Baldwin

    Full Text Available The peptide hormone gastrin binds two ferric ions with high affinity, and iron binding is essential for the biological activity of non-amidated forms of the hormone. Since gastrins act as growth factors in gastrointestinal cancers, and as peptides labelled with Ga and In isotopes are increasingly used for cancer diagnosis, the ability of gastrins to bind other metal ions was investigated systematically by absorption spectroscopy. The coordination structures of the complexes were characterized by extended X-ray absorption fine structure (EXAFS spectroscopy. Changes in the absorption of gastrin in the presence of increasing concentrations of Ga3+ were fitted by a 2 site model with dissociation constants (Kd of 3.3 x 10-7 and 1.1 x 10-6 M. Although the absorption of gastrin did not change upon the addition of In3+ ions, the changes in absorbance on Fe3+ ion binding in the presence of indium ions were fitted by a 2 site model with Kd values for In3+ of 6.5 x 10-15 and 1.7 x 10-7 M. Similar results were obtained with Ru3+ ions, although the Kd values for Ru3+ of 2.6 x 10-13 and 1.2 x 10-5 M were slightly larger than observed for In3+. The structures determined by EXAFS all had metal:gastrin stoichiometries of 2:1 but, while the metal ions in the Fe, Ga and In complexes were bridged by a carboxylate and an oxygen with a metal-metal separation of 3.0-3.3 Å, the Ru complex clearly demonstrated a short range Ru-Ru separation, which was significantly shorter, at 2.4 Å, indicative of a metal-metal bond. We conclude that gastrin selectively binds two In3+ or Ru3+ ions, and that the affinity of the first site for In3+ or Ru3+ ions is higher than for ferric ions. Some of the metal ion-gastrin complexes may be useful for cancer diagnosis and therapy.

  1. Branched-chain fatty acids produced by mutants of Streptomyces fradiae, putative precursors of the lactone ring of tylosin.

    Science.gov (United States)

    Huber, M L; Paschal, J W; Leeds, J P; Kirst, H A; Wind, J A; Miller, F D; Turner, J R

    1990-01-01

    Three branched-chain fatty acids (7-hydroxy-4,6-dimethylnona-2,4-dienoic acid [compound 1], its 7-epimer [compound 2], and 7-keto-4,6-dimethylnona-2,4-dienoic acid [compound 3]) and a ketone (9-hydroxy-6,8-dimethylundeca-4,6-dien-3-one [compound 4]) were isolated from the culture broth of mutants of Streptomyces fradiae which were blocked in the biosynthesis of the macrolide antibiotic tylosin. Two phenotypic classes of mutants of this organism which were blocked in the addition of mycaminose to tylactone (compound 6) accumulated these compounds. These compounds were not produced by mutants which were blocked in lactone synthesis, in steps beyond mycaminose addition, or by the wild-type strain. Synthesis of these compounds, like synthesis of tylosin, was inhibited by the addition of cerulenin. Compounds 1, 2, and 3 were partially interconvertible by these mutants; but they were not produced from the degradation of tylactone and they were not directly incorporated into tylosin by intact cells. The structures of compounds 1 and 2 were equivalent to that of a predicted intermediate (S. Yue, J. S. Duncan, Y. Yamamoto, and C. R. Hutchinson, J. Am. Chem. Soc. 109:1253-1255, 1987) in the biosynthesis of tylactone. The ketone (compound 4) reported previously (N. D. Jones, M. O. Chaney, H. A. Kirst, G. M. Wild, R. H. Baltz, R. L. Hamill, and J. W. Paschal, J. Antibiot. 35:420-425, 1982) appears to be the decarboxylation product of the intermediate following that represented by compound 1. This represents the first report of the isolation of putative precursors of tylactone from tylosin-producing organisms. PMID:2221862

  2. Isovaleryl-homoserine lactone, an unusual branched-chain quorum-sensing signal from the soybean symbiont Bradyrhizobium japonicum.

    Science.gov (United States)

    Lindemann, Andrea; Pessi, Gabriella; Schaefer, Amy L; Mattmann, Margrith E; Christensen, Quin H; Kessler, Aline; Hennecke, Hauke; Blackwell, Helen E; Greenberg, E Peter; Harwood, Caroline S

    2011-10-01

    Many species of Proteobacteria communicate by using LuxI-LuxR-type quorum-sensing systems that produce and detect acyl-homoserine lactone (acyl-HSL) signals. Most of the known signals are straight-chain fatty acyl-HSLs, and evidence indicates that LuxI homologs prefer fatty acid-acyl carrier protein (ACP) over fatty acyl-CoA as the acyl substrate for signal synthesis. Two related LuxI homologs, RpaI and BtaI from Rhodopseudomonas palustris and photosynthetic stem-nodulating bradyrhizobia, direct production of the aryl-HSLs p-coumaroyl-HSL and cinnamoyl-HSL, respectively. Here we report that BjaI from the soybean symbiont Bradyrhizobium japonicum USDA110 is closely related to RpaI and BtaI and catalyzes the synthesis of isovaleryl-HSL (IV-HSL), a branched-chain fatty acyl-HSL. We show that IV-HSL induces expression of bjaI, and in this way IV-HSL functions like many other acyl-HSL quorum-sensing signals. Purified histidine-tagged BjaI was an IV-HSL synthase, which was active with isovaleryl-CoA but not detectably so with isovaleryl-ACP. This suggests that the RpaI-BtaI-BjaI subfamily of acyl-HSL synthases may use CoA- rather than ACP-linked substrates for acyl-HSL synthesis. The bjaI-linked bjaR(1) gene is involved in the response to IV-HSL, and BjaR(1) is sensitive to IV-HSL at concentrations as low as 10 pM. Low but sufficient levels of IV-HSL (about 5 nM) accumulate in B. japonicum culture fluid. The low levels of IV-HSL synthesis have likely contributed to the fact that the quorum-sensing signal from this bacterium has not been described elsewhere.

  3. In brain mitochondria the branched-chain fatty acid phytanic acid impairs energy transduction and sensitizes for permeability transition.

    Science.gov (United States)

    Schönfeld, Peter; Kahlert, Stefan; Reiser, Georg

    2004-10-01

    Phytanic acid (3,7,11,15-tetramethylhexadecanoic acid) accumulates at high levels throughout the body in the adult form of Refsum disease, a peroxisomal genetic disorder. However, it is still unclear why increased levels of phytanic acid have cytotoxic effects. In the present study, we examined the influence of non-esterified phytanic acid on energy-related functions of mitochondria from adult rat brain. Phytanic acid at low concentrations (5-20 microM, i.e. 5-20 nmol/mg of mitochondrial protein) de-energized mitochondria, as indicated by depolarization, stimulation of non-phosphorylating oxygen uptake and inhibition of the reduction of the tetrazolium dye 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide. The unbranched homologue palmitic acid exerted much smaller effects. In addition, phytanic acid reduced state 3 respiration, which was partly due to inhibition of the ADP/ATP carrier. Phytanic acid decreased the rate of adenine nucleotide exchange and increased the degree of control, which the ADP/ATP carrier has on state 3 respiration. Important for functional consequences is the finding that mitochondria, which are preloaded with small amounts of Ca2+ (100 nmol/mg of protein), became highly sensitized to rapid permeability transition even when only low concentrations of phytanic acid (below 5 microM) were applied. In conclusion, the incorporation of phytanic acid into the inner mitochondrial membrane increases the membrane H+ conductance and disturbs the protein-linked functions in energy coupling. This is most probably essential for the short-term toxicity of phytanic acid. Thus in neural tissue, which becomes enriched with phytanic acid, the reduction in mitochondrial ATP supply and the facilitation of the opening of the permeability transition pore are two major mechanisms by which the branched-chain fatty acid phytanic acid induces the onset of degenerative processes.

  4. The Mycobacterium tuberculosis high-affinity iron importer, IrtA, contains an FAD-binding domain.

    Science.gov (United States)

    Ryndak, Michelle B; Wang, Shuishu; Smith, Issar; Rodriguez, G Marcela

    2010-02-01

    Iron is an essential nutrient not freely available to microorganisms infecting mammals. To overcome iron deficiency, bacteria have evolved various strategies including the synthesis and secretion of high-affinity iron chelators known as siderophores. The siderophores produced and secreted by Mycobacterium tuberculosis, exomycobactins, compete for iron with host iron-binding proteins and, together with the iron-regulated ABC transporter IrtAB, are required for the survival of M. tuberculosis in iron deficient conditions and for normal replication in macrophages and in mice. This study further characterizes the role of IrtAB in M. tuberculosis iron acquisition. Our results demonstrate a role for IrtAB in iron import and show that the amino terminus domain of IrtA is a flavin-adenine dinucleotide-binding domain essential for iron acquisition. These results suggest a model in which the amino terminus of IrtA functions to couple iron transport and assimilation.

  5. Molecular evolutionary analysis of the high-affinity K+ transporter gene family in angiosperms.

    Science.gov (United States)

    Yang, P; Hua, C; Zhou, F; Zhang, B-J; Cai, X-N; Chen, Q-Z; Wang, R-L

    2016-07-15

    The high-affinity K(+) transporter (HKT) family comprises a group of multifunctional cation transporters widely distributed in organisms ranging from Bacteria to Eukarya. In angiosperms, the HKT family consists primarily of nine types, whose evolutionary relationships are not fully understood. The available sequences from 31 plant species were used to perform a comprehensive evolutionary analysis, including an examination of selection pressure and estimating phylogenetic tree and gene duplication events. Our results show that a gene duplication in the HKT1;5/HKT1;4 cluster might have led to the divergence of the HKT1;5 and HKT1;4 subfamilies. Additionally, maximum likelihood analysis revealed that the HKT family has undergone a strong purifying selection. An analysis of the amino acids provided strong statistical evidence for a functional divergence between subfamilies 1 and 2. Our study was the first to provide evidence of this functional divergence between these two subfamilies. Analysis of co-evolution in HKT identified 25 co-evolved groups. These findings expanded our understanding of the evolutionary mechanisms driving functional diversification of HKT proteins.

  6. Plasma amino acid relationships during parenteral nutrition.

    Science.gov (United States)

    Wells, F E; Smits, B J

    1980-01-01

    The plasma amino acidfs of 17 patients were studied before and during total parenteral nutrition (TPN). The amino acid (AA) pattern changed similarly for all patients. The AA concentration changes relative to preinfusion (PAER) were the most informative index of change. Two groups of AA were defined, the "branched chain" group (five amino acids) and the "hepatic" group (four amino acids) based on the correlation of PAER values. Comparison of PAER values with the ratio of AA intake to requirement indicated that the requirements of the sick patients were more similar to those of children than those of healthy adults.

  7. Utilization of milk amino acids for body gain in suckling mink (Mustela vison) kits

    DEFF Research Database (Denmark)

    Tauson, Anne-Helene; Fink, Rikke; Hansen, Niels E

    2005-01-01

    The efficiency of utilization of milk amino acids for body gain in suckling mink kits from small (n = 3), medium (n = 6) and large litters (n = 9) was investigated by using 36 mink dams and their litters for measurements during lactation weeks 1 through 4. Measurements on each dam and litter were...... performed once, hence three dams per litter size each week (n = 9). Individual milk intake of kits was determined, milk samples were collected and kits were killed for determination of amino acid composition. The most abundant amino acids in milk were glutamate, leucine and aspartate making up about 40......% of total amino acids. Branched chained amino acids made up slightly more than 20% and sulphur containing amino acids less than 5% of total milk amino acids. In kit bodies the sum of glutamate, aspartate and leucine made up about 32% of amino acids, branched chain amino acids about 16% and sulphur...

  8. Crystallographic analysis reveals the structural basis of the high-affinity binding of iophenoxic acid to human serum albumin.

    Science.gov (United States)

    Ryan, Ali J; Chung, Chun-Wa; Curry, Stephen

    2011-04-18

    Iophenoxic acid is an iodinated radiocontrast agent that was withdrawn from clinical use because of its exceptionally long half-life in the body, which was due in part to its high-affinity binding to human serum albumin (HSA). It was replaced by Iopanoic acid, which has an amino rather than a hydroxyl group at position 3 on the iodinated benzyl ring and, as a result, binds to albumin with lower affinity and is excreted more rapidly from the body. To understand how iophenoxic acid binds so tightly to albumin, we wanted to examine the structural basis of its interaction with HSA. We have determined the co-crystal structure of HSA in complex with iophenoxic acid at 2.75 Å resolution, revealing a total of four binding sites, two of which--in drugs sites 1 and 2 on the protein--are likely to be occupied at clinical doses. High-affinity binding of iophenoxic acid occurs at drug site 1. The structure reveals that polar and apolar groups on the compound are involved in its interactions with drug site 1. In particular, the 3-hydroxyl group makes three hydrogen bonds with the side-chains of Tyr 150 and Arg 257. The mode of binding to drug site 2 is similar except for the absence of a binding partner for the hydroxyl group on the benzyl ring of the compound. The HSA-iophenoxic acid structure indicates that high-affinity binding to drug site 1 is likely to be due to extensive desolvation of the compound, coupled with the ability of the binding pocket to provide a full set of salt-bridging or hydrogen bonding partners for its polar groups. Consistent with this interpretation, the structure also suggests that the lower-affinity binding of iopanoic acid arises because replacement of the 3-hydroxyl by an amino group eliminates hydrogen bonding to Arg 257. This finding underscores the importance of polar interactions in high-affinity binding to albumin.

  9. Crystallographic analysis reveals the structural basis of the high-affinity binding of iophenoxic acid to human serum albumin

    Directory of Open Access Journals (Sweden)

    Chung Chun-wa

    2011-04-01

    Full Text Available Abstract Background Iophenoxic acid is an iodinated radiocontrast agent that was withdrawn from clinical use because of its exceptionally long half-life in the body, which was due in part to its high-affinity binding to human serum albumin (HSA. It was replaced by Iopanoic acid, which has an amino rather than a hydroxyl group at position 3 on the iodinated benzyl ring and, as a result, binds to albumin with lower affinity and is excreted more rapidly from the body. To understand how iophenoxic acid binds so tightly to albumin, we wanted to examine the structural basis of its interaction with HSA. Results We have determined the co-crystal structure of HSA in complex with iophenoxic acid at 2.75 Å resolution, revealing a total of four binding sites, two of which - in drugs sites 1 and 2 on the protein - are likely to be occupied at clinical doses. High-affinity binding of iophenoxic acid occurs at drug site 1. The structure reveals that polar and apolar groups on the compound are involved in its interactions with drug site 1. In particular, the 3-hydroxyl group makes three hydrogen bonds with the side-chains of Tyr 150 and Arg 257. The mode of binding to drug site 2 is similar except for the absence of a binding partner for the hydroxyl group on the benzyl ring of the compound. Conclusions The HSA-iophenoxic acid structure indicates that high-affinity binding to drug site 1 is likely to be due to extensive desolvation of the compound, coupled with the ability of the binding pocket to provide a full set of salt-bridging or hydrogen bonding partners for its polar groups. Consistent with this interpretation, the structure also suggests that the lower-affinity binding of iopanoic acid arises because replacement of the 3-hydroxyl by an amino group eliminates hydrogen bonding to Arg 257. This finding underscores the importance of polar interactions in high-affinity binding to albumin.

  10. Cerebral net exchange of large neutral amino acids after lipopolysaccharide infusion in healthy humans

    DEFF Research Database (Denmark)

    Berg, Ronan Mg; Taudorf, Sarah; Bailey, Damian M;

    2010-01-01

    Alterations in circulating large neutral amino acids (LNAAs), leading to a decrease in the plasma ratio between branched-chain and aromatic amino acids (BCAA/AAA ratio), may be involved in sepsis-associated encephalopathy. We hypothesised that a decrease in the BCAA/AAA ratio occurs along with a ...

  11. Cerebral net exchange of large neutral amino acids after lipopolysaccharide infusion in healthy humans

    DEFF Research Database (Denmark)

    2010-01-01

    Alterations in circulating large neutral amino acids (LNAAs), leading to a decrease in the plasma ratio between branched-chain and aromatic amino acids (BCAA/AAA ratio), may be involved in sepsis-associated encephalopathy. We hypothesised that a decrease in the BCAA/AAA ratio occurs along...

  12. Effect of carbohydrate ingestion on brain exchange of amino acids during sustained exercise in human subjects

    DEFF Research Database (Denmark)

    Blomstrand, Eva; Møller, Kirsten; Secher, Niels Henry

    2005-01-01

    AIM: This study investigated the effect of prolonged exercise with and without carbohydrate intake on the brain exchange of amino acids, especially focussing on tryptophan and branched-chain amino acids (BCAA). METHODS: Five male subjects exercised for 3 h on a cycle ergometer at 200 +/- 7 W on t...

  13. A pharmacological profile of the high-affinity GluK5 kainate receptor.

    Science.gov (United States)

    Møllerud, Stine; Kastrup, Jette Sandholm; Pickering, Darryl S

    2016-10-05

    Mouse GluK5 was expressed in Sf9 insect cells and radiolabelled with [(3)H]-kainate in receptor binding assays (Kd=6.9nM). Western immunoblotting indicated an Sf9 GluK5 band doublet corresponding to the glycosylated (128kDa) and deglycosylated (111kDa) protein, which was identical to the band pattern of native rat brain GluK5. A pharmacological profile of the high-affinity kainate receptor GluK5 is described which is distinct from the profiles of other kainate receptors (GluK1-3). The 27 tested ligands generally show a preferential affinity to GluK1 over GluK5, the exceptions being: dihydrokainate, (S)-5-fluorowillardiine, (S)-glutamate and quisqualate, where the affinity is similar at GluK1 and GluK5. In contrast, quisqualate shows 40-fold higher affinity at GluK5 over GluK3 whereas (S)-1-(2'-amino-2'-caboxyethyl)thienol[3,4-d]pyrimidin-2,4-dione (NF1231), (RS)-2-amino-3-(5-tert-butyl-3-hydroxyisoxazol-4-yl)propionate (ATPA), dihydrokainate and (2S,4R)-4-methyl-glutamate (SYM2081) have higher affinity at GluK3 compared to GluK5. Since some studies have indicated that GluK5 is associated with various diseases in the central nervous system (e.g. schizophrenia, temporal lobe epilepsy, bipolar disorder), selective GluK5 ligands could have therapeutic potential. The distinct pharmacological profile of GluK5 suggests that it would be possible to design ligands with selectivity towards GluK5.

  14. Postruminal synthesis modifies the odd- and branched-chain fatty acid profile from the duodenum to milk.

    Science.gov (United States)

    Vlaeminck, B; Gervais, R; Rahman, M M; Gadeyne, F; Gorniak, M; Doreau, M; Fievez, V

    2015-07-01

    Milk odd- and branched-chain fatty acids (OBCFA) have been suggested as potential biomarkers for rumen function. The potential of milk OBCFA as a biomarker depends on whether their profile reflects the profile observed in the duodenum. The objective of this study was to evaluate whether the OBCFA profile in duodenum samples is reflected in plasma and milk. For this, 2 dairy cattle experiments were used. In experiment 1, 4 Holstein cows fitted with rumen and proximal duodenum cannulas were used in a 4×4 Latin square design. The treatments consisted of 2 nitrogen levels (143 vs. 110g of crude protein/kg of dry matter for high and low N, respectively) combined with either 1 of the 2 energy sources (i.e., starch from barley, corn, and wheat or fiber from soybean hulls and dehydrated beet pulp). In experiment 2, 4 Holstein cows fitted with rumen and proximal duodenum cannulas were used in a 3×3 Latin square design, with the treatments consisting of 3 diets: (1) RNB-, a diet with a crude protein content of 122g/kg of dry matter, predicted to provide protein digested in the small intestine according to the requirement of the animals, but with a shortage of rumen degradable protein; (2) RNB- to which 6g/d of niacin was added through inclusion in the mineral and vitamin premix, and (3) RNB- to which urea was added to balance rumen degradable N supply resulting in a CP content of 156g/kg of dry matter. In both experiments, samples of duodenal digesta, plasma, and milk were collected and analyzed for fatty acids. Additionally, lipids in plasma samples were separated in lipid classes and analyzed for fatty acids. The OBCFA profile in milk was enriched in 15:0, iso-17:0, anteiso-17:0, and cis-9-17:1 as compared with duodenal samples, and milk secretions even exceeded duodenal flows, which suggests occurrence of postruminal synthesis, such as de novo synthesis, desaturation, and elongation. The postruminal modification of the OBCFA profile might hamper the application of OBCFA

  15. Brain amino acid requirements and toxicity: the example of leucine.

    Science.gov (United States)

    Yudkoff, Marc; Daikhin, Yevgeny; Nissim, Ilana; Horyn, Oksana; Luhovyy, Bohdan; Luhovyy, Bogdan; Lazarow, Adam; Nissim, Itzhak

    2005-06-01

    Glutamic acid is an important excitatory neurotransmitter of the brain. Two key goals of brain amino acid handling are to maintain a very low intrasynaptic concentration of glutamic acid and also to provide the system with precursors from which to synthesize glutamate. The intrasynaptic glutamate level must be kept low to maximize the signal-to-noise ratio upon the release of glutamate from nerve terminals and to minimize the risk of excitotoxicity consequent to excessive glutamatergic stimulation of susceptible neurons. The brain must also provide neurons with a constant supply of glutamate, which both neurons and glia robustly oxidize. The branched-chain amino acids (BCAAs), particularly leucine, play an important role in this regard. Leucine enters the brain from the blood more rapidly than any other amino acid. Astrocytes, which are in close approximation to brain capillaries, probably are the initial site of metabolism of leucine. A mitochondrial branched-chain aminotransferase is very active in these cells. Indeed, from 30 to 50% of all alpha-amino groups of brain glutamate and glutamine are derived from leucine alone. Astrocytes release the cognate ketoacid [alpha-ketoisocaproate (KIC)] to neurons, which have a cytosolic branched-chain aminotransferase that reaminates the KIC to leucine, in the process consuming glutamate and providing a mechanism for the "buffering" of glutamate if concentrations become excessive. In maple syrup urine disease, or a congenital deficiency of branched-chain ketoacid dehydrogenase, the brain concentration of KIC and other branched-chain ketoacids can increase 10- to 20-fold. This leads to a depletion of glutamate and a consequent reduction in the concentration of brain glutamine, aspartate, alanine, and other amino acids. The result is a compromise of energy metabolism because of a failure of the malate-aspartate shuttle and a diminished rate of protein synthesis.

  16. High-affinity binding of (/sup 3/H)acetylcholine to muscarinic cholinergic receptors

    Energy Technology Data Exchange (ETDEWEB)

    Kellar, K.J.; Martino, A.M.; Hall, D.P. Jr.; Schwartz, R.D.; Taylor, R.L.

    1985-06-01

    High-affinity binding of (/sup 3/H)acetylcholine to muscarinic cholinergic sites in rat CNS and peripheral tissues was measured in the presence of cytisin, which occupies nicotinic cholinergic receptors. The muscarinic sites were characterized with regard to binding kinetics, pharmacology, anatomical distribution, and regulation by guanyl nucleotides. These binding sites have characteristics of high-affinity muscarinic cholinergic receptors with a Kd of approximately 30 nM. Most of the muscarinic agonist and antagonist drugs tested have high affinity for the (/sup 3/H)acetylcholine binding site, but pirenzepine, an antagonist which is selective for M-1 receptors, has relatively low affinity. The ratio of high-affinity (/sup 3/H)acetylcholine binding sites to total muscarinic binding sites labeled by (/sup 3/H)quinuclidinyl benzilate varies from 9 to 90% in different tissues, with the highest ratios in the pons, medulla, and heart atrium. In the presence of guanyl nucleotides, (/sup 3/H) acetylcholine binding is decreased, but the extent of decrease varies from 40 to 90% in different tissues, with the largest decreases being found in the pons, medulla, cerebellum, and heart atrium. The results indicate that (/sup 3/H)acetylcholine binds to high-affinity M-1 and M-2 muscarinic receptors, and they suggest that most M-2 sites have high affinity for acetylcholine but that only a small fraction of M-1 sites have such high affinity.

  17. Sequence Classification: 184741 [

    Lifescience Database Archive (English)

    Full Text Available Non-TMB TMH Non-TMB Non-TMB Non-TMB Non-TMB >gi|17989219|ref|NP_541852.1| HIGH-AFFI...NITY BRANCHED-CHAIN AMINO ACID TRANSPORT SYSTEM PERMEASE PROTEIN LIVH / HIGH-AFFINITY BRANCHED-CHAIN AMINO A

  18. cDNA heterogeneity suggests structural variants related to the high-affinity IgE receptor.

    Science.gov (United States)

    Liu, F T; Albrandt, K; Robertson, M W

    1988-08-01

    The high-affinity IgE receptor present on mast cells and basophils is responsible for the IgE-mediated activation of these cells. The current model for this receptor depicts a four-subunit structure, alpha beta gamma 2. A cDNA for the alpha subunit was recently cloned and predicts a structure consisting of two homologous extracellular domains, a transmembrane segment, and a cytoplasmic tail. Using a synthetic oligonucleotide corresponding to the amino-terminal sequence of the alpha subunit, we identified a number of cDNA clones from a rat basophilic leukemia cell cDNA library. Nucleotide sequencing established four different forms of cDNA: one is nearly identical to the published cDNA; the second differs from the first in the 5' untranslated sequence; the other two forms use either one or the other of the 5'-end sequences as above and lack 163 base pairs in the region coding for the second extracellular domain. RNase protection analysis with radioactive RNA probes established the heterogeneity of rat basophilic leukemia cell mRNA with regard to both the 5' and the internal sequences. Our results suggest the existence of at least four different protein forms related to the alpha subunit of the high-affinity IgE receptor.

  19. Lactococcal aminotransferases AraT and BcaT are key enzymes for the formation of aroma compounds from amino acids in cheese

    NARCIS (Netherlands)

    Rijnen, L.; Yvon, M.; Kranenburg, van R.; Courtin, P.; Verheul, A.; Chambellon, E.; Smit, G.

    2003-01-01

    Amino acid catabolism plays a major role in cheese aroma development. Previously, we showed that the lactococcal aminotransferases AraT and BcaT initiate the conversion of aromatic amino acids, branched-chain amino acids and methionine to aroma compounds. In this study, we evaluated the importance o

  20. High affinity group III mGluRs regulate mossy fiber input to CA3 interneurons.

    Science.gov (United States)

    Cosgrove, Kathleen E; Meriney, Stephen D; Barrionuevo, Germán

    2011-12-01

    Stratum lacunosum-moleculare interneurons (L-Mi) in hippocampal area CA3 target the apical dendrite of pyramidal cells providing feedforward inhibition. Here we report that selective activation of group III metabotropic glutamate receptors (mGluRs) 4/8 with L(+)-2-amino-4-phosphnobytyric acid (L-AP4; 10 μM) decreased the probability of glutamate release from the mossy fiber (MF) terminals synapsing onto L-Mi. Consistent with this interpretation, application of L-AP4 in the presence of 3 mM strontium decreased the frequency of asynchronous MF EPSCs in L-Mi. Furthermore, the dose response curve showed that L-AP4 at 400 μM produced no further decrease in MF EPSC amplitude compared with 20 μM L-AP4, indicating the lack of mGluRs 7 at these MF terminals. We also found that one mechanism of mGluRs 4/8-mediated inhibition of release is linked to N-type voltage gated calcium channels at MF terminals. Application of the group III mGluR antagonist MSOP (100 μM) demonstrated that mGluRs 4/8 are neither tonically active nor activated by low and moderate frequencies of activity. However, trains of stimuli to the MF at 20 and 40 Hz delivered during the application of MSOP revealed a relief of inhibition of transmitter release and an increase in the overall probability of action potential firing in the postsynaptic L-Mi. Interestingly, the time to first action potential was significantly shorter in the presence of MSOP, indicating that mGluR 4/8 activation delays L-Mi firing in response to MF activity. Taken together, our data demonstrate that the timing and probability of action potentials in L-Mi evoked by MF synaptic input is regulated by the activation of presynaptic high affinity group III mGluRs.

  1. Peptide-based protein capture agents with high affinity, selectivity, and stability as antibody replacements in biodetection assays

    Science.gov (United States)

    Coppock, Matthew B.; Farrow, Blake; Warner, Candice; Finch, Amethist S.; Lai, Bert; Sarkes, Deborah A.; Heath, James R.; Stratis-Cullum, Dimitra

    2014-05-01

    Current biodetection assays that employ monoclonal antibodies as primary capture agents exhibit limited fieldability, shelf life, and performance due to batch-to-batch production variability and restricted thermal stability. In order to improve upon the detection of biological threats in fieldable assays and systems for the Army, we are investigating protein catalyzed capture (PCC) agents as drop-in replacements for the existing antibody technology through iterative in situ click chemistry. The PCC agent oligopeptides are developed against known protein epitopes and can be mass produced using robotic methods. In this work, a PCC agent under development will be discussed. The performance, including affinity, selectivity, and stability of the capture agent technology, is analyzed by immunoprecipitation, western blotting, and ELISA experiments. The oligopeptide demonstrates superb selectivity coupled with high affinity through multi-ligand design, and improved thermal, chemical, and biochemical stability due to non-natural amino acid PCC agent design.

  2. Direct determination of flavor relevant and further branched-chain fatty acids from sheep subcutaneous adipose tissue by gas chromatography with mass spectrometry.

    Science.gov (United States)

    Kaffarnik, Stefanie; Preuß, Siegfried; Vetter, Walter

    2014-07-11

    4-Methyloctanoic acid, 4-ethyloctanoic acid and 4-methylnonanoic acid are key-flavor compounds of sheep and goat. Yet, the low amounts of these volatile branched-chain fatty acids (vBCFAs) in the fat of the ruminants hampered their determination on a routine basis. In this work we developed a gas chromatography/mass spectrometry selected ion monitoring (GC/MS-SIM) method for the direct determination of the vBCFAs as methyl esters, which were obtained after transesterification of subcutaneous adipose tissue of sheep. The excellent sensitivity of the GC/MS-SIM method (limit of quantification, LOQ, 3.6-4.8μg/g; limit of detection, LOD, 1.1-1.4μg/g for 4-Me-8:0, 4-Et-8:0 and 4-Me-9:0) enabled us to determine the three vBCFAs without an enrichment step. Subcutaneous adipose sheep tissue of three different breeds contained 23-88μg/g 4-methyloctanoic acid, 13-26μg/g 4-ethyloctanoic acid and ∼2.9-18μg/g 4-methylnonanoic acid. Since all fatty acids were present in the FAME fraction, the samples could be screened for further branched-chain FAMEs. After elimination of unsaturated fatty acids by hydrogenation (the presence of some unsaturated fatty acids could be verified by this measure), additional measurements in SIM and full scan modes of methyl esters and picolinyl esters enabled the determination of further 97 saturated fatty acids in the samples with eight to 20 carbons. The method is suited for routine analysis and may be useful to investigate the reasons for the abundance/absence of 4-methyloctanoic acid, 4-ethyloctanoic acid and 4-methylnonanoic acid and further branched-chain fatty acids in sheep and goat.

  3. The Branched-Chain Dodecylbenzene Sulfonate Degradation Pathway of Pseudomonas aeruginosa W51D Involves a Novel Route for Degradation of the Surfactant Lateral Alkyl Chain

    OpenAIRE

    Campos-García, Jesús; Esteve, Abraham; Vázquez-Duhalt, Rafael; Ramos, Juán Luis; Soberón-Chávez, Gloria

    1999-01-01

    Pseudomonas aeruginosa W51D is able to grow by using branched-chain dodecylbenzene sulfonates (B-DBS) or the terpenic alcohol citronellol as a sole source of carbon. A mutant derived from this strain (W51M1) is unable to degrade citronellol but still grows on B-DBS, showing that the citronellol degradation route is not the main pathway involved in the degradation of the surfactant alkyl moiety. The structures of the main B-DBS isomers and of some intermediates were identified by gas chromatog...

  4. The Branched-Chain Dodecylbenzene Sulfonate Degradation Pathway of Pseudomonas aeruginosa W51D Involves a Novel Route for Degradation of the Surfactant Lateral Alkyl Chain

    Science.gov (United States)

    Campos-García, Jesús; Esteve, Abraham; Vázquez-Duhalt, Rafael; Ramos, Juán Luis; Soberón-Chávez, Gloria

    1999-01-01

    Pseudomonas aeruginosa W51D is able to grow by using branched-chain dodecylbenzene sulfonates (B-DBS) or the terpenic alcohol citronellol as a sole source of carbon. A mutant derived from this strain (W51M1) is unable to degrade citronellol but still grows on B-DBS, showing that the citronellol degradation route is not the main pathway involved in the degradation of the surfactant alkyl moiety. The structures of the main B-DBS isomers and of some intermediates were identified by gas chromatography-mass spectrometric analysis, and a possible catabolic route is proposed. PMID:10427075

  5. Sertraline and its metabolite desmethylsertraline, but not bupropion or its three major metabolites, have high affinity for P-glycoprotein.

    Science.gov (United States)

    Wang, Jun-Sheng; Zhu, Hao-Jie; Gibson, Bryan Bradford; Markowitz, John Seth; Donovan, Jennifer Lyn; DeVane, Carl Lindsay

    2008-02-01

    The ATP-binding cassette (ABC) transporter protein subfamily B1 line (ABCB1) transporter P-glycoprotein (P-gp) plays an important role in the blood-brain barrier limiting a broad spectrum of substrates from entering the central nervous system. In the present study, the transport activity of P-gp for sertraline, desmethylsertraline, bupropion, and the major metabolites of bupropion, threo-amino alcohol (TB), erythro-amino alcohol (EB), and hydroxy metabolite (HB) was studied using an ATPase assay in expressed human P-gp membranes by measuring concentrations of inorganic P(i) in expressed human P-gp membranes. Verapamil was included as a positive control. The Michaelis-Menten equation was used for characterizing the kinetic data. Sertraline and desmethylsertraline showed high affinity for P-gp. The V(max)/K(m) values of sertraline (1.6 min(-1) x 10(-3)) and desmethylsertraline (1.4 min(-1) x 10(-3)) were comparable with that of verapamil (1.7 min(-1) x 10(-3)). Bupropion and its three metabolites showed very weak affinity for P-gp, with V(max)/K(m) values lower than 0.01 min(-1) x 10(-3). The results of the present study indicate that sertraline and desmethylsertraline have high affinity for P-gp, whereas bupropion and its three major metabolites TB, EB, and HB have very weak affinity for P-gp. These findings may help to explain observed drug-drug interactions among antidepressants.

  6. Structural characterization of a high affinity mononuclear site in the copper(II)-α-synuclein complex.

    Science.gov (United States)

    Bortolus, Marco; Bisaglia, Marco; Zoleo, Alfonso; Fittipaldi, Maria; Benfatto, Maurizio; Bubacco, Luigi; Maniero, Anna Lisa

    2010-12-29

    Human α-Synuclein (aS), a 140 amino acid protein, is the main constituent of Lewy bodies, the cytoplasmatic deposits found in the brains of Parkinson's disease patients, where it is present in an aggregated, fibrillar form. Recent studies have shown that aS is a metal binding protein. Moreover, heavy metal ions, in particular divalent copper, accelerate the aggregation process of the protein. In this work, we investigated the high affinity binding mode of truncated aS (1-99) (aS99) with Cu(II), in a stoichiometric ratio, to elucidate the residues involved in the binding site and the role of copper ions in the protein oligomerization. We used Electron Paramagnetic Resonance spectroscopy on the Cu(II)-aS99 complex at pH 6.5, performing both multifrequency continuous wave experiments and pulsed experiments at X-band. The comparison of 9.5 and 95 GHz data showed that at this pH only one binding mode is present. To identify the nature of the ligands, we performed Electron Spin Echo Envelope Modulation, Hyperfine Sublevel Correlation Spectroscopy, and pulsed Davies Electron-Nuclear Double Resonance (Davies-ENDOR) experiments. We determined that the EPR parameters are typical of a type-II copper complex, in a slightly distorted square planar geometry. Combining the results from the different pulsed techniques, we obtained that the equatorial coordination is {N(Im), N(-), H(2)O, O}, where N(im) is the imino nitrogen of His50, N(-) a deprotonated amido backbone nitrogen that we attribute to His50, H(2)O an exchangeable water molecule, and O an unidentified oxygen ligand. Moreover, we propose that the free amino terminus (Met1) participates in the complex as an axial ligand. The MXAN analysis of the XAS k-edge absorption data allowed us to independently validate the structural features proposed on the basis of the magnetic parameters of the Cu(II)-aS99 complex and then to further refine the quality of the proposed structural model.

  7. Cadmium inhibits the induction of high-affinity nitrate uptake in maize (Zea mays L.) roots.

    Science.gov (United States)

    Rizzardo, Cecilia; Tomasi, Nicola; Monte, Rossella; Varanini, Zeno; Nocito, Fabio F; Cesco, Stefano; Pinton, Roberto

    2012-12-01

    Cadmium (Cd) detoxification involves glutathione and phytochelatins biosynthesis: the higher need of nitrogen should require increased nitrate (NO(3)(-)) uptake and metabolism. We investigated inducible high-affinity NO(3)(-) uptake across the plasma membrane (PM) in maize seedlings roots upon short exposure (10 min to 24 h) to low Cd concentrations (0, 1 or 10 μM): the activity and gene transcript abundance of high-affinity NO(3)(-) transporters, NO(3)(-) reductases and PM H(+)-ATPases were analyzed. Exposure to 1 mM NO(3)(-) led to a peak in high-affinity (0.2 mM) NO(3)(-) uptake rate (induction), which was markedly lowered in Cd-treated roots. Plasma membrane H(+)-ATPase activity was also strongly limited, while internal NO(3)(-) accumulation and NO(3)(-) reductase activity in extracts of Cd treated roots were only slightly lowered. Kinetics of high- and low-affinity NO(3)(-) uptake showed that Cd rapidly (10 min) blocked the inducible high-affinity transport system; the constitutive high-affinity transport system appeared not vulnerable to Cd and the low-affinity transport system appeared to be less affected and only after a prolonged exposure (12 h). Cd-treatment also modified transcript levels of genes encoding high-affinity NO(3)(-) transporters (ZmNTR2.1, ZmNRT2.2), PM H(+)-ATPases (ZmMHA3, ZmMHA4) and NO(3)(-) reductases (ZmNR1, ZmNADH:NR). Despite an expectable increase in NO(3)(-) demand, a negative effect of Cd on NO(3)(-) nutrition is reported. Cd effect results in alterations at the physiological and transcriptional levels of NO(3)(-) uptake from the external solution and it is particularly severe on the inducible high-affinity anion transport system. Furthermore, Cd would limit the capacity of the plant to respond to changes in NO(3) (-) availability.

  8. Ecological genomics of Boechera stricta: Identification of a QTL controlling the allocation of methionine- vs branched-chain amino acid-derived glucosinolates and levels of insect herbivory

    NARCIS (Netherlands)

    Schranz, M.E.; Manzaneda, A.J.; Windsor, A.J.; Clauss, M.; Mitchell-Olds, T.

    2009-01-01

    In the Brassicaceae, glucosinolates influence the feeding, reproduction and development of many insect herbivores. Glucosinolate production and effects on herbivore feeding have been extensively studied in the model species, Arabidopsis thaliana and Brassica crops, both of which constitutively produ

  9. Branched-chain amino acid restriction in Zucker-fatty rats improves muscle insulin sensitivity by enhancing efficiency of fatty acid oxidation and acyl-glycine export

    Directory of Open Access Journals (Sweden)

    Phillip J. White

    2016-07-01

    Conclusions: Our data are consistent with a model wherein elevated circulating BCAA contribute to development of obesity-related insulin resistance by interfering with lipid oxidation in skeletal muscle. BCAA-dependent lowering of the skeletal muscle glycine pool appears to contribute to this effect by slowing acyl-glycine export to the urine.

  10. Regulation of aflatoxin biosynthesis and branched-chain amino acids metabolism in Aspergillus flavus by 2-phenylethanol reveal biocontrol mechanism of Pichia anomala

    Science.gov (United States)

    Pichia anomala WRL-076 is a biocontrol yeast which has been shown to inhibit growth and aflatoxin production of A. flavus. Using the SPME-GC/MS analysis we identified that the volatile, 2-phenylethanol (2-PE) produced by this yeast and demonstrated that the compound inhibited aflatoxin production. W...

  11. Modulation of fatty acid composition and growth in Sporosarcina species in response to temperatures and exogenous branched-chain amino acids.

    Science.gov (United States)

    Tsuda, Kentaro; Nagano, Hideaki; Ando, Akinori; Shima, Jun; Ogawa, Jun

    2017-03-22

    Psychrotolerant endospore-forming Sporosarcina species have been predominantly isolated from minced fish meat (surimi), which is stored under refrigeration after heat treatment. To develop a better method for preserving surimi-based food products, we studied the growth and fatty acid compositions of the isolated strain S92h as well as Sporosarcina koreensis and Sporosarcina aquimarina at cold and moderate temperatures. The growth rates of strain S92h and S. koreensis were the fastest and slowest at cold temperatures, respectively, although these strains grew at a similar rate at moderate temperatures. In all three strains, the proportions of anteiso-C15:0 and unsaturated fatty acids (UFAs) were significantly higher at cold temperatures than at moderate temperatures. Furthermore, supplementation with valine, leucine, and isoleucine resulted in proportional increases in iso-C16:0, iso-C15:0, and anteiso-C15:0, respectively, among the fatty acid compositions of these strains. The proportions of the UFAs were also altered by the supplementation. At cold temperatures, the growth rates of strain S92h and S. koreensis, but not of S. aquimarina, were affected by supplementation with leucine. Supplementation with isoleucine enhanced the growth of S. koreensis at cold temperatures but not that of the other strains. Valine did not affect the growth of any strain. These results indicate that anteiso-C15:0 and UFAs both play important roles in the cold tolerance of the genus Sporosarcina and that these bacteria modulate their fatty acid compositions in response to the growth environment.

  12. GHB receptor targets in the CNS: focus on high-affinity binding sites.

    Science.gov (United States)

    Bay, Tina; Eghorn, Laura F; Klein, Anders B; Wellendorph, Petrine

    2014-01-15

    γ-Hydroxybutyric acid (GHB) is an endogenous compound in the mammalian brain with both low- and high-affinity receptor targets. GHB is used clinically in the treatment of symptoms of narcolepsy and alcoholism, but also illicitly abused as the recreational drug Fantasy. Major pharmacological effects of exogenous GHB are mediated by GABA subtype B (GABAB) receptors that bind GHB with low affinity. The existence of GHB high-affinity binding sites has been known for more than three decades, but the uncovering of their molecular identity has only recently begun. This has been prompted by the generation of molecular tools to selectively study high-affinity sites. These include both genetically modified GABAB knock-out mice and engineered selective GHB ligands. Recently, certain GABA subtype A (GABAA) receptor subtypes emerged as high-affinity GHB binding sites and potential physiological mediators of GHB effects. In this research update, a description of the various reported receptors for GHB is provided, including GABAB receptors, certain GABAA receptor subtypes and other reported GHB receptors. The main focus will thus be on the high-affinity binding targets for GHB and their potential functional roles in the mammalian brain.

  13. Anatomic study of branches-chain base on the superficial peroneal artery%腓浅动脉皮支链的应用解剖

    Institute of Scientific and Technical Information of China (English)

    秦向征; 刘鹏; 许东元; 刘环宇; 金昱; 延光海; 方今女; 丁自海

    2015-01-01

    Objective To definite the specific area and morphology of the branches-chain base on the superficial peroneal artery,provide the Anatomical information for Taking the branches-chain flap.Methods From October,2012 to September,2013,a total of 15 fresh adult cadaveric lower limb were used in the research:which were 3 females and 12 males.There was no history of trauma and surgery.Thirteen specimens were perfused with red latex for microanatomy to observe linking form of the the cutaneous perforators,measuring the outer diameter of perforators at the deep fascia.2 specimens were perfused with self-curing denture acrylic to be vascular cast mold.Results A total of 61 cutaneous branches issued fom the chain of the superficial peroneal nerve nutrient vessels,each side was (4.38 ± 0.77) cutaneous branches,which were the diameter of wearing the deep fascia was (0.57 ± 0.14) mm; Fibula length was (347.76 ± 17.78) mm,The number of cutaneous branches participating the branches-chain was 28,averaged on each side of (2.15 ± 0.38).The diameter was (0.45 ± 0.13) mm,and the area of cutaneous branches-chain of the superficial peroneal artery was (45.95 ± 13.08)% to (76.91 ± 8.71)%.Conclusion Taking the dislocation or free cutaneous flap is safe with branches-chain base on the superficial peroneal artery as the axis in the medium and lower segment of anterolateral of crus.%目的 明确腓浅动脉皮支链形成的区域和形态,为临床切取小腿前外侧皮支链皮瓣提供解剖学基础. 方法 2012年10月-2013年9月,解剖新鲜成年尸体标本下肢15侧,女性3侧,男性12侧,皮肤观察无外伤及手术史.13侧下肢经股动脉灌注红色乳胶,手术显微镜下精细解剖,显示各皮支来源、链状吻合部位和形态,于穿深筋膜处测量各皮支的外径.2侧下肢制做动脉铸型标本. 结果 腓浅动脉皮支共61支,每侧(4.38±0.77)支,各皮支穿过深筋膜处的直径为(0.57±0.14)mm;参与构成腓浅

  14. Amino acid uptake and energy coupling dependent on photosynthesis in Anacystis nidulans.

    OpenAIRE

    Lee-Kaden, J; Simonis, W

    1982-01-01

    The photoautotrophic cyanobacterium Anacystis nidulans was used to investigate the membrane transport of branched-chain, neutral amino acids and its dependence on photosynthetic reactions. The uptake of alpha-amino [1-14C]isobutyric acid and L-[1-14C]leucine followed Michaelis, Menten kinetics and resulted in an energy-dependent accumulation. As in bacteria, different uptake systems for neutral amino acids were present: two DAG (D-alanine, aminoisobutyric acid, and glycine) systems responsibl...

  15. Muscle protein degradation and amino acid metabolism during prolonged knee-extensor exercise in humans

    DEFF Research Database (Denmark)

    Van Hall, Gerrit; Saltin, B; Wagenmakers, A J

    1999-01-01

    to a substantial increase in net muscle protein degradation, and that a lowering of the starting muscle glycogen content leads to a further increase. The carbon atoms of the branched-chain amino acids (BCAA), glutamate, aspartate and asparagine, liberated by protein degradation, and the BCAA and glutamate...

  16. High-affinity olfactory receptor for the death-associated odor cadaverine

    OpenAIRE

    2013-01-01

    Cadaverine and putrescine, two diamines emanating from decaying flesh, are strongly repulsive odors to humans but serve as innate attractive or social cues in other species. Here we show that zebrafish, a vertebrate model system, exhibit powerful and innate avoidance behavior to both diamines, and identify a high-affinity olfactory receptor for cadaverine.

  17. Isolation and cloning of the gene encoding high affinity phosphate transporter in rice

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    High affinity phosphate transporter plays an important role in plant adapting to low phosphorus. Isolation of genes coding this kind of protein has attracted worldwide scholars to accomplish. We aimed to isolate the gene and transfer it to target plants for breeding.

  18. Supramolecular surface immobilization of knottin derivatives for dynamic display of high affinity binders

    NARCIS (Netherlands)

    Sankaran, S.; Ruiter, de M.V.; Cornelissen, J.J.L.M.; Jonkheijm, P.

    2015-01-01

    Knottins are known as a robust and versatile class of miniprotein scaffolds for the presentation of high-affinity binding peptides; however, to date their application in biomaterials, biological coatings, and surface applications have not been explored. We have developed a strategy to recombinantly

  19. GHB receptor targets in the CNS: Focus on high-affinity binding sites

    DEFF Research Database (Denmark)

    Bay, Tina; Eghorn, Laura Friis; Klein, Anders Bue;

    2014-01-01

    γ-Hydroxybutyric acid (GHB) is an endogenous compound in the mammalian brain with both low- and high-affinity receptor targets. GHB is used clinically in the treatment of symptoms of narcolepsy and alcoholism, but also illicitly abused as the recreational drug Fantasy. Major pharmacological effects...

  20. N-Oxide analogs of WAY-100635 : new high affinity 5-HT (1A) receptor antagonists

    NARCIS (Netherlands)

    Oberwinkler - Marchais, Sandrine; Nowicki, B; Pike, VW; Halldin, C; Sandell, J; Chou, YH; Gulyas, B; Brennum, LT; Farde, L; Wikstrom, H V

    2005-01-01

    WAY-100635 [N-(2-(1-(4-(2-methoxyphenyl)piperazinyl)ethyl))-N-(2-pyridinyl)cyclohexanecarboxamide] 1 and its O-des-methyl derivative DWAY 2 are well-known high affinity 5-HT1A receptor antagonists. which when labeled with carbon-II (beta(+): t(1/2) 20.4min) in the carbonyl group are effective radiol

  1. N-Oxide analogs of WAY-100635 : new high affinity 5-HT1A receptor antagonists

    NARCIS (Netherlands)

    Marchais-Oberwinkler, S; Nowicki, B; Pike, VW; Halldin, C; Sandell, J; Chou, YH; Gulyas, B; Brennum, LT; Farde, L; Wikstrom, HV

    2005-01-01

    WAY-100635 [N-(2-(1-(4-(2-methoxyphenyl)piperazinyl)ethyl))-N-(2-pyridinyl)cyclohexanecarboxamide] 1 and its O-des-methyl derivative DWAY 2 are well-known high affinity 5-HT1A receptor antagonists. which when labeled with carbon-II (beta(+): t(1/2) 20.4min) in the carbonyl group are effective radiol

  2. Supramolecular surface immobilization of knottin derivatives for dynamic display of high affinity binders

    NARCIS (Netherlands)

    Sankaran, S.; de Ruiter, Mark Vincent; Cornelissen, Jeroen Johannes Lambertus Maria; Jonkheijm, Pascal

    2015-01-01

    Knottins are known as a robust and versatile class of miniprotein scaffolds for the presentation of high-affinity binding peptides; however, to date their application in biomaterials, biological coatings, and surface applications have not been explored. We have developed a strategy to recombinantly

  3. High Affinity Iron Permease is Required for Virulence of Rhizopus oryzae

    Science.gov (United States)

    Rhizopus oryzae is the most common cause of mucormycosis. Clinical and animal model data clearly demonstrate that the presence of elevated available serum iron predisposes the host to develop mucormycosis. The high affinity iron permease gene (rFTR1) is required for R. oryzae iron transport in iro...

  4. The dual aptamer approach: rational design of a high-affinity FAD aptamer.

    Science.gov (United States)

    Merkle, T; Holder, I T; Hartig, J S

    2016-01-14

    A design strategy for high-affinity aptamers of complex biomolecules is presented. We developed an RNA with FAD-binding properties by combining known ATP- and FMN-aptamers. Cooperative binding of FAD was shown by SPR spectroscopy and fluorescence assays. The strategy should be transferable to several other biomolecules.

  5. Triazoloquinazolinediones as novel high affinity ligands for the benzodiazepine site of GABA(A) receptors

    DEFF Research Database (Denmark)

    Nilsson, Jakob; Gidlöf, Ritha; Nielsen, Elsebet Østergaard

    2011-01-01

    Based on a pharmacophore model of the benzodiazepine-binding site of GABA(A) receptors, a series of 2-aryl-2,6-dihydro[1,2,4]triazolo[4,3-c]quinazoline-3,5-diones (structure type I) were designed, synthesized, and identified as high-affinity ligands of the binding site. For several compounds, K...

  6. Novel cyclen-based linear polymer as a high-affinity binding material for DNA condensation

    Institute of Scientific and Technical Information of China (English)

    XIANG YongZhe; WANG Na; ZHANG Ji; LI Kun; ZHANG ZhongWei; LIN HongHui; YU XiaoQi

    2009-01-01

    A novel cyclen-based linear polyamine (POGEC) was designed and synthesized from the reaction be-tween 1,3-propanediol diglycidyl ether and 1,7-bis(diethoxyphosphory)-1,4,7,10-tetraazacyclod- odecane.High-affinity binding between POGEC and DNA was demonstrated by agarose gel electrophoresis and scanning electron microscopy (SEM). Moreover, the formed POGEC/DNA complex (termed polyplex) could be disassociated to release the free DNA through addition of the physiological concentration of NaCl solution. Fluorescence spectrum was used to measure the high-affinity binding and DNA con-densation capability of POGEC. Circular dichroism (CD) spectrum indicates that the DNA conformation did not change after binding to POEGC.

  7. Novel cyclen-based linear polymer as a high-affinity binding material for DNA condensation

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    A novel cyclen-based linear polyamine (POGEC) was designed and synthesized from the reaction between 1,3-propanediol diglycidyl ether and 1,7-bis(diethoxyphosphory)-1,4,7,10-tetraazacyclod-odecane. High-affinity binding between POGEC and DNA was demonstrated by agarose gel electrophoresis and scanning electron microscopy (SEM). Moreover,the formed POGEC/DNA complex (termed polyplex) could be disassociated to release the free DNA through addition of the physiological concentration of NaCl solution. Fluorescence spectrum was used to measure the high-affinity binding and DNA condensation capability of POGEC. Circular dichroism (CD) spectrum indicates that the DNA conformation did not change after binding to POEGC.

  8. The fourth dimension in immunological space: how the struggle for nutrients selects high-affinity lymphocytes.

    Science.gov (United States)

    Wensveen, Felix M; van Gisbergen, Klaas P J M; Eldering, Eric

    2012-09-01

    Lymphocyte activation via the antigen receptor is associated with radical shifts in metabolism and changes in requirements for nutrients and cytokines. Concomitantly, drastic changes occur in the expression of pro-and anti-apoptotic proteins that alter the sensitivity of lymphocytes to limiting concentrations of key survival factors. Antigen affinity is a primary determinant for the capacity of activated lymphocytes to access these vital resources. The shift in metabolic needs and the variable access to key survival factors is used by the immune system to eliminate activated low-affinity cells and to generate an optimal high-affinity response. In this review, we focus on the control of apoptosis regulators in activated lymphocytes by nutrients, cytokines, and costimulation. We propose that the struggle among individual clones that leads to the formation of high-affinity effector cell populations is in effect an 'invisible' fourth signal required for effective immune responses.

  9. A High-Affinity Metal-Binding Peptide From Escherichia Coli Hypb

    Energy Technology Data Exchange (ETDEWEB)

    Chung, K.C.Chan; Cao, L.; Dias, A.V.; Pickering, I.J.; George, G.N.; Zamble, D.B.

    2009-05-12

    The high-affinity nickel-binding site of the Escherichia coli [NiFe]-hydrogenase accessory protein HypB was localized to residues at the immediate N-terminus of the protein. Modification of a metal-binding fusion protein, site-directed mutagenesis experiments, and DFT calculations were used to identify the N-terminal amine as a ligand as well as the three cysteine residues in the CXXCGCXXX motif. This sequence can be removed from the protein and both a synthesized peptide and a protein fusion bind nickel with a similar affinity and the same structure as the parent metalloprotein, indicating the self-sufficiency of this high-affinity nickel-binding sequence.

  10. Engineering high-affinity PD-1 variants for optimized immunotherapy and immuno-PET imaging.

    Science.gov (United States)

    Maute, Roy L; Gordon, Sydney R; Mayer, Aaron T; McCracken, Melissa N; Natarajan, Arutselvan; Ring, Nan Guo; Kimura, Richard; Tsai, Jonathan M; Manglik, Aashish; Kruse, Andrew C; Gambhir, Sanjiv S; Weissman, Irving L; Ring, Aaron M

    2015-11-24

    Signaling through the immune checkpoint programmed cell death protein-1 (PD-1) enables tumor progression by dampening antitumor immune responses. Therapeutic blockade of the signaling axis between PD-1 and its ligand programmed cell death ligand-1 (PD-L1) with monoclonal antibodies has shown remarkable clinical success in the treatment of cancer. However, antibodies have inherent limitations that can curtail their efficacy in this setting, including poor tissue/tumor penetrance and detrimental Fc-effector functions that deplete immune cells. To determine if PD-1:PD-L1-directed immunotherapy could be improved with smaller, nonantibody therapeutics, we used directed evolution by yeast-surface display to engineer the PD-1 ectodomain as a high-affinity (110 pM) competitive antagonist of PD-L1. In contrast to anti-PD-L1 monoclonal antibodies, high-affinity PD-1 demonstrated superior tumor penetration without inducing depletion of peripheral effector T cells. Consistent with these advantages, in syngeneic CT26 tumor models, high-affinity PD-1 was effective in treating both small (50 mm(3)) and large tumors (150 mm(3)), whereas the activity of anti-PD-L1 antibodies was completely abrogated against large tumors. Furthermore, we found that high-affinity PD-1 could be radiolabeled and applied as a PET imaging tracer to efficiently distinguish between PD-L1-positive and PD-L1-negative tumors in living mice, providing an alternative to invasive biopsy and histological analysis. These results thus highlight the favorable pharmacology of small, nonantibody therapeutics for enhanced cancer immunotherapy and immune diagnostics.

  11. Quantifying high-affinity binding of hydrophobic ligands by isothermal titration calorimetry.

    Science.gov (United States)

    Krainer, Georg; Broecker, Jana; Vargas, Carolyn; Fanghänel, Jörg; Keller, Sandro

    2012-12-18

    A fast and reliable quantification of the binding thermodynamics of hydrophobic high-affinity ligands employing a new calorimetric competition experiment is described. Although isothermal titration calorimetry is the method of choice for a quantitative characterization of intermolecular interactions in solution, a reliable determination of a dissociation constant (K(D)) is typically limited to the range 100 μM > K(D) > 1 nM. Interactions displaying higher or lower K(D) values can be assessed indirectly, provided that a suitable competing ligand is available whose K(D) falls within the directly accessible affinity window. This established displacement assay, however, requires the high-affinity ligand to be soluble at high concentrations in aqueous buffer and, consequently, poses serious problems in the study of protein binding involving small-molecule ligands dissolved in organic solvents--a familiar case in many drug-discovery projects relying on compound libraries. The calorimetric competition assay introduced here overcomes this limitation, thus allowing for a detailed thermodynamic description of high-affinity receptor-ligand interactions involving poorly water-soluble compounds. Based on a single titration of receptor into a dilute mixture of the two competing ligands, this competition assay provides accurate and precise values for the dissociation constants and binding enthalpies of both high- and moderate-affinity ligands. We discuss the theoretical background underlying the approach, demonstrate its practical application to metal ion chelation and high-affinity protein-inhibitor interactions, and explore its potential and limitations with the aid of simulations and statistical analyses.

  12. Nuclear Choline Acetyltransferase Activates Transcription of a High-affinity Choline Transporter*

    OpenAIRE

    Matsuo, Akinori; Bellier, Jean-Pierre; Nishimura, Masaki; YASUHARA, Osamu; Saito, Naoaki; Kimura, Hiroshi

    2010-01-01

    Choline acetyltransferase (ChAT) synthesizes the neurotransmitter, acetylcholine, at cholinergic nerve terminals. ChAT contains nuclear localization signals and is also localized in the nuclei of neural and non-neuronal cells. Nuclear ChAT might have an as yet unidentified function, such as transcriptional regulation. In this study, we investigated the alteration of candidate gene transcription by ChAT. We chose high affinity choline transporter (CHT1) and vesicular acetylcholine transporter ...

  13. Neurotransmitter/sodium symporter orthologue LeuT has a single high-affinity substrate site.

    Science.gov (United States)

    Piscitelli, Chayne L; Krishnamurthy, Harini; Gouaux, Eric

    2010-12-23

    Neurotransmitter/sodium symporters (NSSs) couple the uptake of neurotransmitter with one or more sodium ions, removing neurotransmitter from the synaptic cleft. NSSs are essential to the function of chemical synapses, are associated with multiple neurological diseases and disorders, and are the targets of therapeutic and illicit drugs. LeuT, a prokaryotic orthologue of the NSS family, is a model transporter for understanding the relationships between molecular mechanism and atomic structure in a broad range of sodium-dependent and sodium-independent secondary transporters. At present there is a controversy over whether there are one or two high-affinity substrate binding sites in LeuT. The first-reported crystal structure of LeuT, together with subsequent functional and structural studies, provided direct evidence for a single, high-affinity, centrally located substrate-binding site, defined as the S1 site. Recent binding, flux and molecular simulation studies, however, have been interpreted in terms of a model where there are two high-affinity binding sites: the central, S1, site and a second, the S2 site, located within the extracellular vestibule. Furthermore, it was proposed that the S1 and S2 sites are allosterically coupled such that occupancy of the S2 site is required for the cytoplasmic release of substrate from the S1 site. Here we address this controversy by performing direct measurement of substrate binding to wild-type LeuT and to S2 site mutants using isothermal titration calorimetry, equilibrium dialysis and scintillation proximity assays. In addition, we perform uptake experiments to determine whether the proposed allosteric coupling between the putative S2 site and the S1 site manifests itself in the kinetics of substrate flux. We conclude that LeuT harbours a single, centrally located, high-affinity substrate-binding site and that transport is well described by a simple, single-substrate kinetic mechanism.

  14. Reconstitution of high-affinity opioid agonist binding in brain membranes

    Energy Technology Data Exchange (ETDEWEB)

    Remmers, A.E.; Medzihradsky, F. (Univ. of Michigan Medical School, Ann Arbor (United States))

    1991-03-15

    In synaptosomal membranes from rat brain cortex, the {mu} selective agonist ({sup 3}H)dihydromorphine in the absence of sodium, and the nonselective antagonist ({sup 3}H)naltrexone in the presence of sodium, bound to two populations of opioid receptor sites with K{sub d} values of 0.69 and 8.7 nM for dihydromorphine, and 0.34 and 5.5 nM for naltrexone. The addition of 5 {mu}M guanosine 5{prime}-({gamma}-thio)triphosphate (GTP({gamma}S)) strongly reduced high-affinity agonist but not antagonist binding. Exposure of the membranes to high pH reduced the number of GTP({gamma}-{sup 35}S) binding sites by 90% and low K{sub m}, opioid-sensitive GTPase activity by 95%. In these membranes, high-affinity agonist binding was abolished and modulation of residual binding by GTP({gamma}S) was diminished. Alkali treatment of the glioma cell membranes prior to fusion inhibited most of the low K{sub m} GTPase activity and prevented the reconstitution of agonist binding. The results show that high-affinity opioid agonist binding reflects the ligand-occupied receptor - guanine nucleotide binding protein complex.

  15. Enhanced selection of high affinity DNA-reactive B cells following cyclophosphamide treatment in mice.

    Directory of Open Access Journals (Sweden)

    Daisuke Kawabata

    Full Text Available A major goal for the treatment of patients with systemic lupus erythematosus with cytotoxic therapies is the induction of long-term remission. There is, however, a paucity of information concerning the effects of these therapies on the reconstituting B cell repertoire. Since there is recent evidence suggesting that B cell lymphopenia might attenuate negative selection of autoreactive B cells, we elected to investigate the effects of cyclophosphamide on the selection of the re-emerging B cell repertoire in wild type mice and transgenic mice that express the H chain of an anti-DNA antibody. The reconstituting B cell repertoire in wild type mice contained an increased frequency of DNA-reactive B cells; in heavy chain transgenic mice, the reconstituting repertoire was characterized by an increased frequency of mature, high affinity DNA-reactive B cells and the mice expressed increased levels of serum anti-DNA antibodies. This coincided with a significant increase in serum levels of BAFF. Treatment of transgene-expressing mice with a BAFF blocking agent or with DNase to reduce exposure to autoantigen limited the expansion of high affinity DNA-reactive B cells during B cell reconstitution. These studies suggest that during B cell reconstitution, not only is negative selection of high affinity DNA-reactive B cells impaired by increased BAFF, but also that B cells escaping negative selection are positively selected by autoantigen. There are significant implications for therapy.

  16. Single-experiment displacement assay for quantifying high-affinity binding by isothermal titration calorimetry.

    Science.gov (United States)

    Krainer, Georg; Keller, Sandro

    2015-04-01

    Isothermal titration calorimetry (ITC) is the gold standard for dissecting the thermodynamics of a biomolecular binding process within a single experiment. However, reliable determination of the dissociation constant (KD) from a single titration is typically limited to the range 100 μM>KD>1 nM. Interactions characterized by a lower KD can be assessed indirectly by so-called competition or displacement assays, provided that a suitable competitive ligand is available whose KD falls within the directly accessible window. However, this protocol is limited by the fact that it necessitates at least two titrations to characterize one high-affinity inhibitor, resulting in considerable consumption of both sample material and time. Here, we introduce a fast and efficient ITC displacement assay that allows for the simultaneous characterization of both a high-affinity ligand and a moderate-affinity ligand competing for the same binding site on a receptor within a single experiment. The protocol is based on a titration of the high-affinity ligand into a solution containing the moderate-affinity ligand bound to the receptor present in excess. The resulting biphasic binding isotherm enables accurate and precise determination of KD values and binding enthalpies (ΔH) of both ligands. We discuss the theoretical background underlying the approach, demonstrate its practical application to metal ion chelation, explore its potential and limitations with the aid of simulations and statistical analyses, and elaborate on potential applications to protein-inhibitor interactions.

  17. Isolation of Anti-Ricin Protective Antibodies Exhibiting High Affinity from Immunized Non-Human Primates

    Directory of Open Access Journals (Sweden)

    Tal Noy-Porat

    2016-03-01

    Full Text Available Ricin, derived from the castor bean plant Ricinus communis, is one of the most potent and lethal toxins known, against which there is no available antidote. To date, the use of neutralizing antibodies is the most promising post-exposure treatment for ricin intoxication. The aim of this study was to isolate high affinity anti-ricin antibodies that possess potent toxin-neutralization capabilities. Two non-human primates were immunized with either a ricin-holotoxin- or subunit-based vaccine, to ensure the elicitation of diverse high affinity antibodies. By using a comprehensive set of primers, immune scFv phage-displayed libraries were constructed and panned. A panel of 10 antibodies (five directed against the A subunit of ricin and five against the B subunit was isolated and reformatted into a full-length chimeric IgG. All of these antibodies were found to neutralize ricin in vitro, and several conferred full protection to ricin-intoxicated mice when given six hours after exposure. Six antibodies were found to possess exceptionally high affinity toward the toxin, with KD values below pM (koff < 1 × 10−7 s−1 that were well correlated with their ability to neutralize ricin. These antibodies, alone or in combination, could be used for the development of a highly-effective therapeutic preparation for post-exposure treatment of ricin intoxication.

  18. Characterization of a genetically reconstituted high-affinity system for serotonin transport

    Energy Technology Data Exchange (ETDEWEB)

    Chang, A.S.S.; Lam, D.M.K. (Baylor College of Medicine, Woodlands, TX (USA) Baylor College of Medicine, Houston, TX (USA)); Frnka, J.V.; Chen, D. (Baylor College of Medicine, Woodlands, TX (USA))

    1989-12-01

    By transfecting mouse fibroblast L-M cells with human genomic DNA, the authors have established and identified several clonal cell lines that stably express a high-affinity serotonin (5-HT)-uptake mechanism absent in untransfected host cells. One such cell line, L-S1, possesses features of 5-({sup 3}H)HT uptake similar to those previously characterized in the central nervous system and blood platelets: (i) specificity for 5-HT; (ii) antagonism by imipramine, a known inhibitor of high-affinity 5-HT uptake; (iii) both Na{sup +} and temperature dependence; (iv) kinetic saturability; and (v) high affinity for 5-HT. This cell line can be used to compare the relative efficacies of known blockers of 5-HT uptake and thereby offers a rapid and reliable assay system for testing novel inhibitors of this system. Since L-S1 contains stably integrated human DNA in its genome, they postulate that the observed 5-HT-uptake system resulted from the expression of human gene(s) coding for the 5-HT transporter. Thus, cell lines such as L-S1 may represent novel means for screening and developing therapeutic agents specific for neutrotransmitter-uptake systems as well as substrate for the cloning and elucidation of the genes encoding the various neurotransmitter transporters.

  19. Structural Basis for High-Affinity Peptide Inhibition of Human Pin1

    Science.gov (United States)

    Zhang, Yan; Daum, Sebastian; Wildemann, Dirk; Zhou, Xiao Zhen; Verdecia, Mark A.; Bowman, Marianne E.; Lücke, Christian; Hunter, Tony; Lu, Kun-Ping; Fischer, Gunter; Noel, Joseph P.

    2009-01-01

    Human Pin1 is a key regulator of cell-cycle progression and plays growth-promoting roles in human cancers. High-affinity inhibitors of Pin1 may provide a unique opportunity for disrupting oncogenic pathways. Here we report two high-resolution X-ray crystal structures of human Pin1 bound to non-natural peptide inhibitors. The structures of the bound high-affinity peptides identify a type-I β-turn conformation for Pin1 prolyl peptide isomerase domain–peptide binding and an extensive molecular interface for high-affinity recognition. Moreover, these structures suggest chemical elements that may further improve the affinity and pharmacological properties of future peptide-based Pin inhibitors. Finally, an intramolecular hydrogen bond observed in both peptide complexes mimics the cyclic conformation of FK506 and rapamycin. Both FK506 and rapamycin are clinically important inhibitors of other peptidyl-prolyl cis-trans isomerases. This comparative discovery suggests that a cyclic peptide polyketide bridge, like that found in FK506 and rapamycin or a similar linkage, may significantly improve the binding affinity of structure-based Pin1 inhibitors. PMID:17518432

  20. Plasma amino acid and metabolite signatures tracking diabetes progression in the UCD-T2DM rat model

    Science.gov (United States)

    Elevations of plasma concentrations of branched-chain amino acids (BCAAs) are observed in human insulin resistance and type 2 diabetes mellitus (T2DM); however, there has been some controversy with respect to the passive or causative nature of the BCAA phenotype. Using untargeted metabolomics, plasm...

  1. 仔猪支链氨基酸营养效果%Piglets and Nutritional Effect of Chain Amino Acid

    Institute of Scientific and Technical Information of China (English)

    方俊

    2003-01-01

    @@ 支链氨基酸 (branched chain amino acid,简称 BCAA)是一组在碳链上具有支链结构的脂肪族中性氨基酸,包括亮氨酸 (Leu)、异亮氨酸 (Ile)和缬氨酸 (Val),均为必需氨基酸.

  2. G196 epitope tag system: a novel monoclonal antibody, G196, recognizes the small, soluble peptide DLVPR with high affinity

    Science.gov (United States)

    Tatsumi, Kasumi; Sakashita, Gyosuke; Nariai, Yuko; Okazaki, Kosuke; Kato, Hiroaki; Obayashi, Eiji; Yoshida, Hisashi; Sugiyama, Kanako; Park, Sam-Yong; Sekine, Joji; Urano, Takeshi

    2017-01-01

    The recognition specificity of monoclonal antibodies (mAbs) has made mAbs among the most frequently used tools in both basic science research and in clinical diagnosis and therapies. Precise determination of the epitope allows the development of epitope tag systems to be used with recombinant proteins for various purposes. Here we describe a new family of tag derived from the epitope recognized by a highly specific mAb G196. The minimal epitope was identified as the five amino acid sequence Asp-Leu-Val-Pro-Arg. Permutation analysis was used to characterize the binding requirements of mAb G196, and the variable regions of the mAb G196 were identified and structurally analyzed by X-ray crystallography. Isothermal titration calorimetry revealed the high affinity (Kd = 1.25 nM) of the mAb G196/G196-epitope peptide interaction, and G196-tag was used to detect several recombinant cytosolic and nuclear proteins in human and yeast cells. mAb G196 is valuable for developing a new peptide tagging system for cell biology and biochemistry research. PMID:28266535

  3. Functional identification of activity-regulated, high-affinity glutamine transport in hippocampal neurons inhibited by riluzole.

    Science.gov (United States)

    Erickson, Jeffrey D

    2017-07-01

    Glutamine (Gln) is considered the preferred precursor for the neurotransmitter pool of glutamate (Glu), the major excitatory transmitter in the mammalian CNS. Here, an activity-regulated, high-affinity Gln transport system is described in developing and mature neuron-enriched hippocampal cultures that is potently inhibited by riluzole (IC50 1.3 ± 0.5 μM), an anti-glutamatergic drug, and is blocked by low concentrations of 2-(methylamino)isobutyrate (MeAIB), a system A transport inhibitor. K(+) -stimulated MeAIB transport displays an affinity (Km ) for MeAIB of 37 ± 1.2 μM, saturates at ~ 200 μM, is dependent on extracellular Ca(2+) , and is blocked by inhibition of voltage-gated Ca(2+) channels. Spontaneous MeAIB transport is also dependent on extracellullar Ca(2+) and voltage-gated calcium channels, but is also blocked by the Na(+) channel blocker tetrodotoxin, by Glu receptor antagonists, and by GABA indicating its dependence on intact neural circuits driven by endogenous glutamatergic activity. The transport of MeAIB itself does not rely on Ca(2+) , but on Na(+) ions, and is pH sensitive. Activity-regulated, riluzole-sensitive spontaneous and K(+) -stimulated transport is minimal at 7-8 days in vitro, coordinately induced during the next 2 weeks and is maximally expressed by days in vitro > 20; the known period for maturation of the Glu/Gln cycle and regulated pre-synaptic Glu release. Competition analyses with various amino acids indicate that Gln is the most likely physiological substrate. Activity-regulated Gln/MeAIB transport is not observed in astrocytes. The functional identification of activity-regulated, high-affinity, riluzole-sensitive Gln/MeAIB transport in hippocampal neurons may have important ramifications in the neurobiology of activity-stimulated pre-synaptic Glu release, the Glu/Gln cycle between astrocytes and neurons, and neuronal Glu-induced excitotoxicity. Cover Image for this issue: doi: 10.1111/jnc.13805. © 2017

  4. The Refsum disease marker phytanic acid, a branched chain fatty acid, affects Ca2+ homeostasis and mitochondria, and reduces cell viability in rat hippocampal astrocytes.

    Science.gov (United States)

    Kahlert, Stefan; Schönfeld, Peter; Reiser, Georg

    2005-02-01

    The saturated branched chain fatty acid, phytanic acid, a degradation product of chlorophyll, accumulates in Refsum disease, an inherited peroxisomal disorder with neurological clinical features. To elucidate the pathogenic mechanism, we investigated the influence of phytanic acid on cellular physiology of rat hippocampal astrocytes. Phytanic acid (100 microM) induced an immediate transient increase in cytosolic Ca2+ concentration, followed by a plateau. The peak of this biphasic Ca2+ response was largely independent of extracellular Ca2+, indicating activation of cellular Ca2+ stores by phytanic acid. Phytanic acid depolarized mitochondria without causing in situ swelling of mitochondria. The slow decrease of mitochondrial potential is not consistent with fast and simultaneous opening of the mitochondrial permeability transition pore. However, phytanic acid induced substantial generation of reactive oxygen species. Phytanic acid caused astroglia cell death after a few hours of exposure. We suggest that the cytotoxic effect of phytanic acid seems to be due to a combined action on Ca2+ regulation, mitochondrial depolarization, and increased ROS generation in brain cells.

  5. Cistus ladanifer L. Shrub is Rich in Saturated and Branched Chain Fatty Acids and their Concentration Increases in the Mediterranean Dry Season.

    Science.gov (United States)

    Guerreiro, Olinda; Alves, Susana P; Duarte, Maria F; Bessa, Rui J B; Jerónimo, Eliana

    2015-05-01

    The Cistus ladanifer L. shrub is a widespread species of the Mediterranean region that is available as a feed source for ruminants all the year round, constituting a source of energy and nutrients when most of the vegetation is dry. However, there is no trustworthy information about the fatty acid composition of C. ladanifer, as well as no information about the seasonal and age related changes in their fatty acid composition. Thus, we collected the aerial parts of C. ladanifer plants of two age groups [young vs. older ones (2-6 years old)] during four consecutive seasons to characterize their fatty acid composition. The fatty acid composition of C. ladanifer is dominated by saturated fatty acids including the occurrence of two methyl branched chain fatty acids (BCFA), the iso-19:0 and iso-21:0, which as far as we know were detected for the first time in shrubs. Also, we demonstrated that several labdane type compounds might interfere with the fatty acid analysis of C. ladanifer. Marked seasonal changes in BCFA and polyunsaturated fatty acids (PUFA) were found, suggesting that BCFA can replace PUFA in plant lipids at high environmental temperatures.

  6. Evaluation of FT-NIR and ATR-FTIR spectroscopy techniques for determination of minor odd- and branched-chain saturated and trans unsaturated milk fatty acids.

    Science.gov (United States)

    Stefanov, Ivan; Baeten, Vincent; Abbas, Ouissam; Vlaeminck, Bruno; De Baets, Bernard; Fievez, Veerle

    2013-04-10

    Determination of nutritionally important trans MUFA, CLA, and OBCFA milk fatty acids (often present in amounts lower than 1.0 g/100 g of total fat) using fast and nondestructive analytical methods would enhance their use as diagnostic tools in dairy herd and human health management. Here, PLS regression using ATR/FTIR spectra indicated potential for determination of trans-11 C18:1 and trans-12 C18:1 (Rcv² ≥ 0.80), and trans-9 C18:1 in very minor concentration (Rcv² > 0.82), as well as anteiso C15:0 (Rcv² = 0.57) and iso C17:0 (Rcv² = 0.61). Furthermore, the main cis-9,trans-11 CLA isomer was predicted well despite the high trans MUFA concentration. Differentiation between the CLA and the trans MUFA signals was evident (based on specific cis/trans bands), and branched-chain saturated fatty acid methyl esters revealed specific iso and anteiso ATR/FTIR absorbance bands. None of the minor FA PLS results with FT-NIR showed interesting potential, except satisfactory predictions for trans-9 C18:1 and cis-9,trans-11 CLA. Overall, ATR/FTIR resulted in better calibrations and provided more specific information for determination of minor milk fatty acids.

  7. Identification and quantification of even and odd chained 5-n alkylresorcinols, branched chain-alkylresorcinols and methylalkylresorcinols in Quinoa (Chenopodium quinoa).

    Science.gov (United States)

    Ross, Alastair B; Svelander, Cecilia; Karlsson, Göran; Savolainen, Otto I

    2017-04-01

    Quinoa is a pseudocereal grown in the Andean region of South America that is of increasing interest worldwide as an alternative staple food. We have detected a complex mixture of both odd- and even-alkyl chain alkylresorcinols (AR), branched-chain alkylresorcinols (bcAR) and methylalkylresorcinols (mAR) in ethyl acetate extracts of quinoa. We quantified the content of AR in 17 commercial samples of quinoa, and found that the mean±SD content of AR was 58±16μg/g, bcAR was 182±52μg/g, and mAR was 136±40μg/g. AR from quinoa could also be detected in plasma after eating quinoa, indicating that some of these unique AR could be used as biomarkers of quinoa intake in humans. Further work is required to understand the role of these ARs in the quinoa plant and whether any of the novel ARs may be of particular interest in human nutrition.

  8. Human epidermal Langerhans cells express the high affinity receptor for immunoglobulin E (Fc epsilon RI)

    OpenAIRE

    1992-01-01

    It has been suggested that epidermal Langerhans cells (LC) bearing immunoglobulin E (IgE) may be involved in the genesis of atopic disease. The identity of the IgE receptor(s) on LC remained unclear, although it represents a crucial point in understanding cellular events linked to the binding of allergens to LC via IgE. In this report, we demonstrate that epidermal LC express the high affinity receptor for the Fc fragment of IgE (Fc epsilon RI) which has, so far, only been described on mast c...

  9. High-affinity benzodiazepine receptor ligands among benzodiazepines and betacarbolines with different intrinsic activity

    Energy Technology Data Exchange (ETDEWEB)

    Yliniemelae, A.; Gynther, J. (Univ. of Kuopio (Finland)); Konschin, H.; Tylli, H. (Univ. of Helsinki (Finland)); Rouvinen, J. (Univ. of Joensuu (Finland))

    1989-01-01

    Structural and electrostatic features of diazepam, flumazenil, and methyl betacarboline-3-carboxylate (BCCM) have been investigated using the molecular superimposition method. These high-affinity benzodiazepine (BZ) receptor ligands are structurally unrelated and they have different intrinsic activity. These ligands are superimposed in such a way that common structural and electrostatic features essential for the high receptor binding affinity overlap. In addition to this binding pharmacophore, there are roughly three separate binding zones in the BZ receptor, one for each class of ligands. The intrinsic activity of BZ receptor ligands depends on the molecular structures and the way the ligand approaches the receptor.

  10. A high affinity monoclonal antibody recognizing the light chain of human coagulating factor VII.

    Science.gov (United States)

    Sarial, Sheila; Asadi, Farzad; Jeddi-Tehrani, Mahmood; Hadavi, Reza; Bayat, Ali Ahmad; Mahmoudian, Jafar; Taghizadeh-Jahed, Masoud; Shokri, Fazel; Rabbani, Hodjattallah

    2012-12-01

    Factor VII (FVII) is a serine protease-coagulating element responsible for the initiation of an extrinsic pathway of clot formation. Here we generated and characterized a high affinity monoclonal antibody that specifically recognizes human FVII. Recombinant human FVII (rh-FVII) was used for the production of a monoclonal antibody using BALB/c mice. The specificity of the antibody was determined by Western blot using plasma samples from human, mouse, sheep, goat, bovine, rabbit, and rat. Furthermore, the antibody was used to detect transiently expressed rh-FVII in BHK21 cell line using Western blot and sandwich ELISA. A mouse IgG1 (kappa chain) monoclonal antibody clone 1F1-B11 was produced against rh-FVII. The affinity constant (K(aff)) of the antibody was calculated to be 6.4×10(10) M(-1). The antibody could specifically recognize an epitope on the light chain of hFVII, with no reactivity with factor VII from several other animals. In addition, transiently expressed rh-FVII in BHK21 cells was recognized by 1F1-B11. The high affinity as well as the specificity of 1F1-B11 for hFVII will facilitate the affinity purification of hFVII and also production of FVII deficient plasma and minimizes the risk of bovine FVII contamination when fetal bovine serum-supplemented media are used for production and subsequent purification of rh-FVII.

  11. Acylated heptapeptide binds albumin with high affinity and application as tag furnishes long-acting peptides

    Science.gov (United States)

    Zorzi, Alessandro; Middendorp, Simon J.; Wilbs, Jonas; Deyle, Kaycie; Heinis, Christian

    2017-07-01

    The rapid renal clearance of peptides in vivo limits this attractive platform for the treatment of a broad range of diseases that require prolonged drug half-lives. An intriguing approach for extending peptide circulation times works through a `piggy-back' strategy in which peptides bind via a ligand to the long-lived serum protein albumin. In accordance with this strategy, we developed an easily synthesized albumin-binding ligand based on a peptide-fatty acid chimera that has a high affinity for human albumin (Kd=39 nM). This ligand prolongs the elimination half-life of cyclic peptides in rats 25-fold to over seven hours. Conjugation to a peptide factor XII inhibitor developed for anti-thrombotic therapy extends the half-life from 13 minutes to over five hours, inhibiting coagulation for eight hours in rabbits. This high-affinity albumin ligand could potentially extend the half-life of peptides in human to several days, substantially broadening the application range of peptides as therapeutics.

  12. Discovery of Compounds that Positively Modulate the High Affinity Choline Transporter

    Science.gov (United States)

    Choudhary, Parul; Armstrong, Emma J.; Jorgensen, Csilla C.; Piotrowski, Mary; Barthmes, Maria; Torella, Rubben; Johnston, Sarah E.; Maruyama, Yuya; Janiszewski, John S.; Storer, R. Ian; Skerratt, Sarah E.; Benn, Caroline L.

    2017-01-01

    Cholinergic hypofunction is associated with decreased attention and cognitive deficits in the central nervous system in addition to compromised motor function. Consequently, stimulation of cholinergic neurotransmission is a rational therapeutic approach for the potential treatment of a variety of neurological conditions. High affinity choline uptake (HACU) into acetylcholine (ACh)-synthesizing neurons is critically mediated by the sodium- and pH-dependent high-affinity choline transporter (CHT, encoded by the SLC5A7 gene). This transporter is comparatively well-characterized but otherwise unexplored as a potential drug target. We therefore sought to identify small molecules that would enable testing of the hypothesis that positive modulation of CHT mediated transport would enhance activity-dependent cholinergic signaling. We utilized existing and novel screening techniques for their ability to reveal both positive and negative modulation of CHT using literature tools. A screening campaign was initiated with a bespoke compound library comprising both the Pfizer Chemogenomic Library (CGL) of 2,753 molecules designed specifically to help enable the elucidation of new mechanisms in phenotypic screens and 887 compounds from a virtual screening campaign to select molecules with field-based similarities to reported negative and positive allosteric modulators. We identified a number of previously unknown active and structurally distinct molecules that could be used as tools to further explore CHT biology or as a starting point for further medicinal chemistry. PMID:28289374

  13. Humanization of high-affinity antibodies targeting glypican-3 in hepatocellular carcinoma

    Science.gov (United States)

    Zhang, Yi-Fan; Ho, Mitchell

    2016-01-01

    Glypican-3 (GPC3) is a cell-surface heparan sulfate proteoglycan highly expressed in hepatocellular carcinoma (HCC). We have generated a group of high-affinity mouse monoclonal antibodies targeting GPC3. Here, we report the humanization and testing of these antibodies for clinical development. We compared the affinity and cytotoxicity of recombinant immunotoxins containing mouse single-chain variable regions fused with a Pseudomonas toxin. To humanize the mouse Fvs, we grafted the combined KABAT/IMGT complementarity determining regions (CDR) into a human IgG germline framework. Interestingly, we found that the proline at position 41, a non-CDR residue in heavy chain variable regions (VH), is important for humanization of mouse antibodies. We also showed that two humanized anti-GPC3 antibodies (hYP7 and hYP9.1b) in the IgG format induced antibody-dependent cell-mediated cytotoxicity and complement-dependent-cytotoxicity in GPC3-positive cancer cells. The hYP7 antibody was tested and showed inhibition of HCC xenograft tumor growth in nude mice. This study successfully humanizes and validates high affinity anti-GPC3 antibodies and sets a foundation for future development of these antibodies in various clinical formats in the treatment of liver cancer. PMID:27667400

  14. High affinity binding of (/sup 3/H)cocaine to rat liver microsomes

    Energy Technology Data Exchange (ETDEWEB)

    El-Maghrabi, E.A.; Calligaro, D.O.; Eldefrawi, M.E.

    1988-01-01

    )/sup 3/H)cocaine bound reversible, with high affinity and stereospecificity to rat liver microsomes. Little binding was detected in the lysosomal, mitochondrial and nuclear fractions. The binding kinetics were slow and the kinetically calculated K/sub D/ was 2 nM. Induction of mixed function oxidases by phenobarbital did not produce significant change in (/sup 3/H)cocaine binding. On the other hand, chronic administration of cocaine reduced (/sup 3/H)cocaine binding drastically. Neither treatment affected the affinity of the liver binding protein for cocaine. Microsomes from mouse and human livers had less cocaine-binding protein and lower affinity for cocaine than those from rat liver. Binding of (/sup 3/H)cocaine to rat liver microsomes was insensitive to monovalent cations and > 10 fold less sensitive to biogenic amines than the cocaine receptor in rat striatum. However, the liver protein had higher affinity for cocaine and metabolites except for norcocaine. Amine uptake inhibitors displaced (/sup 3/H)cocaine binding to liver with a different rank order of potency than their displacement of (/sup 3/H)cocaine binding to striatum. This high affinity (/sup 3/H)cocaine binding protein in liver is not likely to be monooxygenase, but may have a role in cocaine-induced hepatotoxicity

  15. High affinity binding site-mediated prevention of chemical absorption across the gastrointestinal tract.

    Science.gov (United States)

    Rasmussen, M V; Barker, T T; Silbart, L K

    2001-12-15

    Preventing mucosal absorption of low-molecular weight compounds such as carcinogens, toxins and drugs could help prevent many diseases. To characterize the effects of dose and timing on high-affinity binding site mediated sequestration of specific chemical ligands in the gastrointestinal tract, avidin was perorally-administered to mice either prior to or mixed with 3H-biotin. Avidin enhanced fecal 3H-biotin excretion in a dose-dependent manner, consistent with the accepted mechanism of egg white-induced biotin deficiency syndrome. Avidin administration up to 4 h before 3H-biotin administration also enhanced fecal 3H-biotin excretion. Activated charcoal (AC) reduced 3H-biotin absorption when mixed with 3H-biotin before ingestion, but was ineffective when ingested prior to 3H-biotin. These studies suggest that ingestion of high-affinity protein binding sites can establish an absorptive barrier at the gastrointestinal mucosa to prevent the uptake of unwanted low molecular-weight chemicals.

  16. High affinity mouse-human chimeric Fab against Hepatitis B surface antigen

    Institute of Scientific and Technical Information of China (English)

    Biplab Bose; Navin Khanna; Subrat K Acharya; Subrata Sinha

    2005-01-01

    AIM: Passive immunotherapy using antibody against hepatitis B surface antigen (HBsAg) has been advocated in certain cases of Hepatitis B infection. We had earlier reported on the cloning and expression of a high affinity scFv derived from a mouse monoclonal (5S) against HBsAg. However this mouse antibody cannot be used for therapeutic purposes as it may elicit anti-mouse immune responses. Chimerization by replacing mouse constant domains with human ones can reduce the immunogenicity of this antibody.METHODS: We cloned the VH and VL genes of this mouse antibody; and fused them with CH1 domain of human IgG1 and CL domain of human kappa chain respectively. These chimeric genes were cloned into a phagemid vector. After initial screening using the phage display system, the chimeric Fab was expressed in soluble form in E. Coli.RESULTS: The chimeric Fab was purified from the bacterial periplasmic extract. We characterized the chimeric Fab using several in vitro techniques and it was observed that the chimeric molecule retained the high affinity and specificity of the original mouse monoclonal.This chimeric antibody fragment was further expressed in different strains of E> coli to increase the yield.CONCLUSION: We have generated a mouse-human chimeric Fab against HBsAg without any significant loss in binding and epitope specificity. This chimeric Fab fragment can be further modified to generate a fulllength chimeric antibody for therapeutic uses.

  17. Genetic evidence of a high-affinity cyanuric acid transport system in Pseudomonas sp. ADP.

    Science.gov (United States)

    Platero, Ana I; Santero, Eduardo; Govantes, Fernando

    2014-03-01

    The Pseudomonas sp. ADP plasmid pADP-1 encodes the activities involved in the hydrolytic degradation of the s-triazine herbicide atrazine. Here, we explore the presence of a specific transport system for the central intermediate of the atrazine utilization pathway, cyanuric acid, in Pseudomonas sp. ADP. Growth in fed-batch cultures containing limiting cyanuric acid concentrations is consistent with high-affinity transport of this substrate. Acquisition of the ability to grow at low cyanuric acid concentrations upon conjugal transfer of pADP1 to the nondegrading host Pseudomonas putida KT2442 suggests that all activities required for this phenotype are encoded in this plasmid. Co-expression of the pADP1-borne atzDEF and atzTUVW genes, encoding the cyanuric acid utilization pathway and the subunits of an ABC-type solute transport system, in P. putida KT2442 was sufficient to promote growth at cyanuric acid concentrations as low as 50 μM in batch culture. Taken together, our results strongly suggest that the atzTUVW gene products are involved in high-affinity transport of cyanuric acid.

  18. Purification of high affinity benzodiazepine receptor binding site fragments from rat brain

    Energy Technology Data Exchange (ETDEWEB)

    Klotz, K.L.

    1984-01-01

    In central nervous system benzodiazepine recognition sites occur on neuronal cell surfaces as one member of a multireceptor complex, including recognition sites for benzodiazepines, gamma aminobutyric acid (GABA), barbiturates and a chloride ionophore. During photoaffinity labelling, the benzodiazepine agonist, /sup 3/H-flunitrazepam, is irreversibly bound to central benzodiazepine high affinity recognition sites in the presence of ultraviolet light. In these studies a /sup 3/H-flunitrazepam radiolabel was used to track the isolation and purification of high affinity agonist binding site fragments from membrane-bound benzodiazepine receptor in rat brain. The authors present a method for limited proteolysis of /sup 3/H-flunitrazepam photoaffinity labeled rat brain membranes, generating photolabeled benzodiazepine receptor fragments containing the agonist binding site. Using trypsin chymotrypsin A/sub 4/, or a combination of these two proteases, they have demonstrated the extent and time course for partial digestion of benzodiazepine receptor, yielding photolabeled receptor binding site fragments. These photolabeled receptor fragments have been further purified on the basis of size, using ultrafiltration, gel permeation chromatography, and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) as well as on the basis of hydrophobicity, using a high performance liquid chromatography (HPLC) precolumn, several HPLC elution schemes, and two different HPLC column types. Using these procedures, they have purified three photolabeled benzodiazepine receptor fragments containing the agonist binding site which appear to have a molecular weight of less than 2000 daltons each.

  19. Mechanism of toxicity of the branched-chain fatty acid phytanic acid, a marker of Refsum disease, in astrocytes involves mitochondrial impairment.

    Science.gov (United States)

    Reiser, Georg; Schönfeld, Peter; Kahlert, Stefan

    2006-01-01

    Phytanic acid is a saturated branched-chain fatty acid, which is formed by bacterial degradation of chlorophyll in the intestinal tract of ruminants. The methyl group in beta-position prevents degradation of phytanic acid by the beta-oxidation pathway. Therefore, degradation of phytanic acid is initiated by alpha-oxidation in peroxisomes. The inherited peroxisomal disorder Refsum disease is characterised by accumulation of phytanic acid. Unusually high concentrations of phytanic acid can be found in the plasma of Refsum disease patients, who suffer from neurodegeneration and muscle dystrophy. Phytanic acid has been suggested to be causally involved in the clinical symptoms. To elucidate the pathogenic mechanism, we investigated the influence of phytanic acid in rat hippocampal astrocytes by monitoring the cytosolic Ca(2+) concentration, the mitochondrial membrane potential (Deltapsi(m)), the generation of reactive oxygen species as well as the cellular ATP level. In response to phytanic acid (100 microM) cytosolic Ca(2+) was quickly increased. The phytanic acid-evoked Ca(2+) response was transient and involved activation of intracellular Ca(2+) stores. In isolated rat brain mitochondria, phytanic acid dissipated Deltapsi(m) in a reversible and dose-dependent manner. Moreover, phytanic acid released cytochrome c from mitochondria. Depending on the mitochondrial activity state, phytanic acid either stimulated or inhibited the electron flux within the respiratory chain. In addition, phytanic acid induced substantial generation of reactive oxygen species in isolated mitochondria as well as in intact cells. Phytanic acid caused cell death of astrocytes within a few hours of exposure. In conclusion, we suggest that phytanic acid initiates astrocyte cell death by activating the mitochondrial route of apoptosis.

  20. Branched-chain fatty acid composition of human milk and the impact of maternal diet: the Global Exploration of Human Milk (GEHM) Study.

    Science.gov (United States)

    Dingess, Kelly A; Valentine, Christina J; Ollberding, Nicholas J; Davidson, Barbara S; Woo, Jessica G; Summer, Suzanne; Peng, Yongmei M; Guerrero, M Lourdes; Ruiz-Palacios, Guillermo M; Ran-Ressler, Rinat R; McMahon, Robert J; Brenna, J Thomas; Morrow, Ardythe L

    2017-01-01

    An understudied component of the diet, branched-chain fatty acids (BCFAs) are distinctive saturated fatty acids that may have an important influence on health. Human-milk fatty acid composition is known to differ worldwide, but comparative data are lacking on BCFAs. We tested the hypotheses that concentrations of BCFAs in human milk differ between populations and are associated with maternal diet. We surveyed the BCFA composition of samples collected as part of a standardized, prospective study of human-milk composition. Mothers were enrolled from 3 urban populations with differing diets: Cincinnati, Ohio; Shanghai, China; and Mexico City, Mexico. Enrollment was limited to healthy mothers of term singleton infants. We undertook a cross-sectional analysis of milk from all women with samples at postpartum week 4 (n = 359; ∼120 women/site). Fatty acids were extracted from milk by using a modified Bligh-Dyer technique and analyzed by gas chromatography. Statistical analysis was performed by ANOVA and Tobit regression. For Cincinnati mothers, 24-h diet recalls were analyzed in relation to the individual BCFA concentrations measured in milk samples. Total BCFAs in milk differed by site, with the highest concentration in Cincinnati followed by Mexico City and Shanghai (mean ± SE: 7.90 ± 0.41, 6.10 ± 0.36, and 4.27 ± 0.25 mg/100 mL, respectively; P Milk concentrations of iso-14:0 and anteiso-15:0 were associated with maternal intake of dairy; iso-16:0 was associated with maternal intakes of dairy and beef. BCFA concentrations in milk at 4 wk postpartum differed between mothers from Cincinnati, Shanghai, and Mexico City. Variations in human-milk BCFAs are influenced by diet. The impact of BCFAs on infant health warrants investigation. © 2017 American Society for Nutrition.

  1. Identification of a Key Gene Involved in Branched-Chain Short Fatty Acids Formation in Natto by Transcriptional Analysis and Enzymatic Characterization in Bacillus subtilis.

    Science.gov (United States)

    Hong, Chenlu; Chen, Yangyang; Li, Lu; Chen, Shouwen; Wei, Xuetuan

    2017-03-01

    Natto as a fermented soybean product has many health benefits for human due to its rich nutritional and functional components. However, the unpleasant odor of natto, caused by the formation of branched-chain short fatty acids (BCFAs), prohibits the wide acceptance of natto products. This work is to identify the key gene of BCFAs formation and develop the guidance to reduce natto odor. Transcriptional analysis of BCFAs synthesis pathway genes was conducted in two Bacillus subtilis strains with obvious different BCFAs synthesis abilities. The transcriptional levels of bcd, bkdAA, and ptb in B. subtilis H-9 were 2.7-fold, 0.7-fold, and 8.9-fold higher than that of B. subtilis H-4, respectively. Therefore, the ptb gene with the highest transcriptional change was considered as the key gene in BCFAs synthesis. The ptb encoded enzyme Ptb was further characterized by inducible expression in Escherichia coli. The recombinant Ptb protein (about 32 kDa) was verified by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis analysis. The catalysis functions of Ptb were confirmed on substrates of isovaleryl-CoA and isobutyryl-CoA, and the higher catalysis efficiency of Ptb on isovaleryl-CoA explained the higher level of isovaleric acid in natto. The optimal activities of Ptb were observed at 50 °C and pH 8.0, and the enzymatic activity was inhibited by Ca(2+), Zn(2+), Ba(2+), Mn(2+), Cu(2+), SDS, and EDTA. Collectively, this study reports a key gene responsible for BCFAs formation in natto fermentation and provides potential strategies to solve the odor problem.

  2. Branched-Chain in situ Hybridization for κ and λ Light Chains: A Powerful Ancillary Technique for Determining B-Cell Clonality in Cytology Samples

    Science.gov (United States)

    Arora, Kshitij; Chebib, Ivan; Zukerberg, Lawrence; Gandhi, Manoj; Rivera, Miguel; Ting, David; Deshpande, Vikram

    2017-01-01

    BACKGROUND Current immunohistochemical and in situ hybridization (ISH) assays are generally inconclusive for clonality unless plasmacytic differentiation is present. This study examined a series of cytology specimens and explored the ability of a branched-chain RNA (bRNA) ISH assay for immunoglobulin κ constant (IGKC) and immunoglobulin λ constant (IGLC) to detect a clonal population of B lymphocytes. METHODS Pathology databases were used to identify fine-needle aspiration biopsies (n = 28) and exfoliative cytology samples (n = 20). Demographic, flow cytometry, and excision biopsy results were recorded. bRNA ISH was performed on the Leica Bond platform with the following probes: IGKC, IGLC, immunoglobulin λ-like polypeptide 5 (IGLL5), and a housekeeping gene (HKG). RESULTS The bRNA ISH assay was validated with 30 surgical biopsies. On bRNA ISH, a clonal B-cell population (light-chain ratio > 10:1) was detected in 22 of 28 cases with a final diagnosis of lymphoma. In 2 cases, a κ predominance was present, although the ratio was <10:1. Eleven of the 17 κ-clonal lymphomas also expressed IGLL5, the latter recognized by the presence of an intranuclear signal. Two B-cell lymphomas lacked IGKC and IGLC, whereas 2 cases were negative for the HKG. In 12 of the 20 cases with reactive lymphoid tissue, bRNA ISH identified a polyclonal lymphoid population. No light-chain messenger RNA was detected in 6 cases (typically those associated with very few B cells). CONCLUSIONS The automated bRNA ISH platform is a robust technique for detecting a clonal B-cell population in cytology material. PMID:26524200

  3. A specific protein-enriched enteral formula decreases cortisolemia and improves plasma albumin and amino acid concentrations in elderly patients

    Directory of Open Access Journals (Sweden)

    Pérez de la Cruz Antonio

    2010-07-01

    Full Text Available Abstract Background Old age is associated with an involuntary and progressive but physiological loss of muscle mass. The aim of this study was to evaluate the effects of exclusive consumption for 6 months of a protein-enriched enteral diet with a relatively high content of branched-chain amino acids on albuminemia, cortisolemia, plasma amino acids, insulin resistance, and inflammation biomarkers in elderly patients. Methods Thirty-two patients from the Clinical Nutrition Outpatient Unit at our hospital exclusively consumed a protein-enriched enteral diet for 6 months. Data were collected at baseline and at 3 and 6 months on anthropometric and biochemical parameters and on plasma concentrations of amino acids, cortisol, adrenocorticotropic hormone, urea, creatinine, insulin resistance, and inflammation biomarkers. Results The percentage of patients with albumin concentration below normal cut-off values decreased from 18% to 0% by the end of the study. At 6 months, concentrations of total plasma (p = 0.008 and essential amino acids (p = 0.011, especially branched-chain amino acids (p = 0.031, were higher versus baseline values, whereas 3-methylhistidine (p = 0.001, cortisol (p = 0.001 and adrenocorticotropic hormone (p = 0.004 levels were lower. Conclusions Regular intake of specific protein-enriched enteral formula increases plasma essential amino acids, especially branched-chain amino acids, and decreases cortisol and 3-methylhistidine, while plasma urea and creatinine remain unchanged.

  4. New Synthesis and Tritium Labeling of a Selective Ligand for Studying High-affinity γ-Hydroxybutyrate (GHB) Binding Sites

    OpenAIRE

    Vogensen, Stine B.; Marek, Aleš; Bay, Tina; Wellendorph, Petrine; Kehler, Jan; Bundgaard, Christoffer; Frølund, Bente; Pedersen, Martin H. F.; Clausen, Rasmus P.

    2013-01-01

    3-Hydroxycyclopent-1-enecarboxylic acid (HOCPCA, 1) is a potent ligand for the high-affinity GHB binding sites in the CNS. An improved synthesis of 1 together with a very efficient synthesis of [3H]-1 is described. The radiosynthesis employs in situ generated lithium trimethoxyborotritide. Screening of 1 against different CNS targets establishes a high selectivity and we demonstrate in vivo brain penetration. In vitro characterization of [3H]-1 binding shows high specificity to the high-affin...

  5. Mutational analysis of the high-affinity zinc binding site validates a refined human dopamine transporter homology model.

    Directory of Open Access Journals (Sweden)

    Thomas Stockner

    Full Text Available The high-resolution crystal structure of the leucine transporter (LeuT is frequently used as a template for homology models of the dopamine transporter (DAT. Although similar in structure, DAT differs considerably from LeuT in a number of ways: (i when compared to LeuT, DAT has very long intracellular amino and carboxyl termini; (ii LeuT and DAT share a rather low overall sequence identity (22% and (iii the extracellular loop 2 (EL2 of DAT is substantially longer than that of LeuT. Extracellular zinc binds to DAT and restricts the transporter's movement through the conformational cycle, thereby resulting in a decrease in substrate uptake. Residue H293 in EL2 praticipates in zinc binding and must be modelled correctly to allow for a full understanding of its effects. We exploited the high-affinity zinc binding site endogenously present in DAT to create a model of the complete transmemberane domain of DAT. The zinc binding site provided a DAT-specific molecular ruler for calibration of the model. Our DAT model places EL2 at the transporter lipid interface in the vicinity of the zinc binding site. Based on the model, D206 was predicted to represent a fourth co-ordinating residue, in addition to the three previously described zinc binding residues H193, H375 and E396. This prediction was confirmed by mutagenesis: substitution of D206 by lysine and cysteine affected the inhibitory potency of zinc and the maximum inhibition exerted by zinc, respectively. Conversely, the structural changes observed in the model allowed for rationalizing the zinc-dependent regulation of DAT: upon binding, zinc stabilizes the outward-facing state, because its first coordination shell can only be completed in this conformation. Thus, the model provides a validated solution to the long extracellular loop and may be useful to address other aspects of the transport cycle.

  6. Mutational analysis of the high-affinity zinc binding site validates a refined human dopamine transporter homology model.

    Science.gov (United States)

    Stockner, Thomas; Montgomery, Therese R; Kudlacek, Oliver; Weissensteiner, Rene; Ecker, Gerhard F; Freissmuth, Michael; Sitte, Harald H

    2013-01-01

    The high-resolution crystal structure of the leucine transporter (LeuT) is frequently used as a template for homology models of the dopamine transporter (DAT). Although similar in structure, DAT differs considerably from LeuT in a number of ways: (i) when compared to LeuT, DAT has very long intracellular amino and carboxyl termini; (ii) LeuT and DAT share a rather low overall sequence identity (22%) and (iii) the extracellular loop 2 (EL2) of DAT is substantially longer than that of LeuT. Extracellular zinc binds to DAT and restricts the transporter's movement through the conformational cycle, thereby resulting in a decrease in substrate uptake. Residue H293 in EL2 praticipates in zinc binding and must be modelled correctly to allow for a full understanding of its effects. We exploited the high-affinity zinc binding site endogenously present in DAT to create a model of the complete transmemberane domain of DAT. The zinc binding site provided a DAT-specific molecular ruler for calibration of the model. Our DAT model places EL2 at the transporter lipid interface in the vicinity of the zinc binding site. Based on the model, D206 was predicted to represent a fourth co-ordinating residue, in addition to the three previously described zinc binding residues H193, H375 and E396. This prediction was confirmed by mutagenesis: substitution of D206 by lysine and cysteine affected the inhibitory potency of zinc and the maximum inhibition exerted by zinc, respectively. Conversely, the structural changes observed in the model allowed for rationalizing the zinc-dependent regulation of DAT: upon binding, zinc stabilizes the outward-facing state, because its first coordination shell can only be completed in this conformation. Thus, the model provides a validated solution to the long extracellular loop and may be useful to address other aspects of the transport cycle.

  7. A nitrogen-dependent switch in the high affinity ammonium transport in Medicago truncatula.

    Science.gov (United States)

    Straub, Daniel; Ludewig, Uwe; Neuhäuser, Benjamin

    2014-11-01

    Ammonium transporters (AMTs) are crucial for the high affinity primary uptake and translocation of ammonium in plants. In the model legume Medicago truncatula, the genomic set of AMT-type ammonium transporters comprises eight members. Only four genes were abundantly expressed in young seedlings, both in roots and shoots. While the expression of all AMTs in the shoot was not affected by the nitrogen availability, the dominating MtAMT1;1 gene was repressed by nitrogen in roots, despite that cellular nitrogen concentrations were far above deficiency levels. A contrasting de-repression by nitrogen was observed for MtAMT1;4 and MtAMT2;1, which were both expressed at intermediate level. Weak expression was found for MtAMT1;2 and MtAMT2;3, while the other AMTs were not detected in young seedlings. When expressed from their endogenous promoters, translational fusion proteins of MtAMT1;1 and MtAMT2;1 with green fluorescent protein were co-localized in the plasma membrane of rhizodermal cells, but also detected in cortical root layers. Both transporter proteins similarly functionally complemented a yeast strain that is deficient in high affinity ammonium transport, both at acidic and neutral pH. The uptake into yeast mediated by these transporters saturated with Km AMT1;1 = 89 µM and Km AMT2;1 = 123 µM, respectively. When expressed in oocytes, MtAMT1;1 mediated much larger (15)N-ammonium uptake than MtAMT2;1, but NH4 (+) currents were only recorded for MtAMT1;1. These currents saturated with a voltage-dependent Km = 90 µM at -80 mV. The cellular localization and regulation of the AMTs suggests that MtAMT1;1 encodes the major high affinity ammonium transporter gene in low nitrogen grown young M. truncatula roots and despite the similar localization and substrate affinity, MtAMT2;1 appears functionally distinct and more important at higher nitrogen supply.

  8. Putative M2 muscarinic receptors of rat heart have high affinity for organophosphorus anticholinesterases

    Energy Technology Data Exchange (ETDEWEB)

    Silveira, C.L.; Eldefrawi, A.T.; Eldefrawi, M.E. (Univ. of Maryland, Baltimore (USA))

    1990-05-01

    The M2 subtype of muscarinic receptor is predominant in heart, and such receptors were reported to be located in muscles as well as in presynaptic cholinergic and adrenergic nerve terminals. Muscarinic receptors of rat heart were identified by the high affinity binding of the agonist (+)-(3H)cis-methyldioxolane ((3H)CD), which has been used to label a high affinity population of M2 receptors. A single population of sites was detected and (3H)CD binding was sensitive to the M2 antagonist himbacine but much less so to pirenzepine, the M1 antagonist. These cardiac receptors had different sensitivities to NiCl2 and N-ethylmaleimide from brain muscarinic receptors, that were also labeled with (3H)CD and considered to be of the M2 subtype. Up to 70% of the (3H)CD-labeled cardiac receptors had high affinities for several organophosphate (OP) anticholinesterases. (3H)CD binding was inhibited by the nerve agents soman, VX, sarin, and tabun, with K0.5 values of 0.8, 2, 20, and 50 nM, respectively. It was also inhibited by echothiophate and paraoxon with K0.5 values of 100 and 300 nM, respectively. The apparent competitive nature of inhibition of (3H)CD binding by both sarin and paraoxon suggests that the OPs bind to the acetylcholine binding site of the muscarinic receptor. Other OP insecticides had lower potencies, inhibiting less than 50% of 5 nM (3H)CD binding by 1 microM of EPN, coumaphos, dioxathion, dichlorvos, or chlorpyriphos. There was poor correlation between the potencies of the OPs in reversibly inhibiting (3H)CD binding, and their anticholinesterase activities and toxicities. Acetylcholinesterases are the primary targets for these OP compounds because of the irreversible nature of their inhibition, which results in building of acetylcholine concentrations that activate muscarinic and nicotinic receptors and desensitize them, thereby inhibiting respiration.

  9. Selection and design of high affinity DNA ligands for mutant single-chain derivatives of the bacteriophage 434 repressor

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Single-chain repressor RRTRES is a derivative of bacteriophage 434 repressor, which contains covalently dimerized DNA-binding domains (amino acids 1-69) of the phage 434 repressor. In this single-chain molecule, the wild type domain R is connected to the mutant domain RTRES by a recombinant linker in a head-to-tail arrangement. The DNA-contacting amino acids of RTRES at the -1, 1, 2, and 5 positions of the a3 helix are T, R, E, S respectively. By using a randomized DNA pool containing the central sequence -CATACAAGAAAGNNNNNNTTT-, a cyclic, in vitro DNA-binding site selection was performed. The selected population was cloned and the individual members were characterized by determining their binding affinities to RRTRES. The results showed that the optimal operators contained the TTAC or TTCC sequences in the underlined positions as above, and that the Kd values were in the 1×10-12 mol/L-1×10-11mol/L concentration range. Since the affinity of the natural 434 repressor to its natural operator sites is in the 1×10-9 mol/L range, the observed binding affinity increase is remarkable. It was also found that binding affinity was strongly affected by the flanking bases of the optimal tetramer binding sites, especially by the base at the 5′ position. We constructed a new homodimeric single-chain repressor RTRESRTRES and its DNA-binding specificity was tested by using a series of new operators designed according to the recog-nition properties previously determined for the RTRES domain. These operators containing the con-sensus sequence GTAAGAAARNTTACN or GGAAGAAARNTTCCN (R is A or G) were recognized by RTRESRTRES specifically, and with high binding affinity. Thus, by using a combination of random selection and rational design principles, we have discovered novel, high affinity protein-DNA inter-actions with new specificity. This method can potentially be used to obtain new binding specificity for other DNA-binding proteins.

  10. Centrifuge-induced hypergravity and glutamate efflux by reversal of high-affinity, sodium-dependent transporters from rat brain synaptosomes.

    Science.gov (United States)

    Borisova, T.; Himmelreich, N.

    Glutamate uptake by high affinity sodium-dependent glutamate transporters is essential for termination of the synaptic transmission. Glutamate transporters may also contribute to an increase in extracellular glutamate. Glutamate efflux can occur by reversal of the sodium-dependent glutamate transporters during ATP depletion and dissipation of the sodium gradient across the cell membrane. Depolarization-induced calcium independent release of neurotransmitter from synaptosomal cytosolic pool is Na+-dependent and due to reverse of the neurotransmitter transporters also. We used monovalent organic cations N-methyl-D-glucamine (NMDG) to replace extracellular sodium, suggesting that the reducing of Na+ elucidate further the mechanism underlying Ca2+-independent glutamate release. A reduction in extracellular sodium would facilitate reversal of sodium-dependent transporters with extrusion of glutamate. We have compared the basal release of glutamate in Ca2+-free Na+-supplemented and NMDG-supplemented medium in control and after exposure of animals to long-arm centrifuge-induced hypergravity (ten G, during one hour). Replacement of sodium by NMDG enhanced basal level of neurotransmitter. The value of basal level increased to 110± 4% and 140± 2% in the medium with NMDG in comparison with Na+ under the control and hypergravity conditions, respectively. It is likely to reflect the enhancement of the neurotransmitter level in cytosolic pool. Thermodynamic considerations show that the extracellular level of a amino acid neurotransmitter, such as glutamate, that can be generated by transporter reversal are directly proportional to the intracellular concentration of the intracellular concentration of amino acid. KCl-stimulated glutamate release from cytosolic pool increased not statistically after hypergravity loading. We examined the effects of transporter inhibitors DL-threo-beta-benzyloxyaspartate ( DL-TBOA) on the release to elucidate whether reverse transport via the

  11. Expression of the Arabidopsis high-affinity hexose transporter STP13 correlates with programmed cell death.

    Science.gov (United States)

    Norholm, Morten H H; Nour-Eldin, Hussam H; Brodersen, Peter; Mundy, John; Halkier, Barbara A

    2006-04-17

    We report the biochemical characterization in Xenopus oocytes of the Arabidopsis thaliana membrane protein, STP13, as a high affinity, hexose-specific H(+)-symporter. Studies with kinase activators suggest that it is negatively regulated by phosphorylation. STP13 promoter GFP reporter lines show GFP expression only in the vascular tissue in emerging petals under non-stressed conditions. Quantitative PCR and the pSTP13-GFP plants show induction of STP13 in programmed cell death (PCD) obtained by treatments with the fungal toxin fumonisin B1 and the pathogen Pseudomonas syringae. A role for STP13 in PCD is supported by microarray data from e.g. plants undergoing senescence and a strong correlation between STP13 transcripts and the PCD phenotype in different accelerated cell death (acd11) mutants.

  12. Neutrophil recruitment limited by high-affinity bent β2 integrin binding ligand in cis.

    Science.gov (United States)

    Fan, Zhichao; McArdle, Sara; Marki, Alex; Mikulski, Zbigniew; Gutierrez, Edgar; Engelhardt, Britta; Deutsch, Urban; Ginsberg, Mark; Groisman, Alex; Ley, Klaus

    2016-08-31

    Neutrophils are essential for innate immunity and inflammation and many neutrophil functions are β2 integrin-dependent. Integrins can extend (E(+)) and acquire a high-affinity conformation with an 'open' headpiece (H(+)). The canonical switchblade model of integrin activation proposes that the E(+) conformation precedes H(+), and the two are believed to be structurally linked. Here we show, using high-resolution quantitative dynamic footprinting (qDF) microscopy combined with a homogenous conformation-reporter binding assay in a microfluidic device, that a substantial fraction of β2 integrins on human neutrophils acquire an unexpected E(-)H(+) conformation. E(-)H(+) β2 integrins bind intercellular adhesion molecules (ICAMs) in cis, which inhibits leukocyte adhesion in vitro and in vivo. This endogenous anti-inflammatory mechanism inhibits neutrophil aggregation, accumulation and inflammation.

  13. Selection of high affine peptide ligands for detection of Clostridium Tyrobutyricum spores.

    Science.gov (United States)

    Lavilla, María; De Luis, Ruth; Pérez, María Dolores; Calvo, Miguel; Sánchez, Lourdes

    2009-11-01

    Clostridium tyrobutyricum is the main agent responsible for "late blowing" in cheese, which causes severe economic losses. Nowadays, the reference method for its detection is the Most-Probable-Number (MPN); however, it is time consuming and non-specific. Thus, in order to check milk contamination with spores of C. tyrobutyricum, a more specific and rapid method would be required. The objective of this work was to obtain a ligand to establish the basis to develop a biomagnetic separation method for detection of C. tyrobutyricum spores. This study describes the selection of thirteen highly affine peptides to C. tyrobutyricum spores from a phage-display peptide library. In order to test the ability of the peptides attached to a solid support to bind the spores, the most frequent peptide was synthesised and used to coat paramagnetic beads.

  14. Practical strategies for the evaluation of high-affinity protein/nucleic acid interactions.

    Science.gov (United States)

    Altschuler, Sarah E; Lewis, Karen A; Wuttke, Deborah S

    2013-01-01

    The quantitative evaluation of binding interactions between proteins and nucleic acids is highly sensitive to a variety of experimental conditions. Optimization of these conditions is critical for obtaining high quality, reproducible data, particularly in the context of very high affinity interactions. Here, we discuss the practical considerations involved in optimizing the apparent binding constant of an interaction as measured by two common quantitative assays, electrophoretic mobility shift assay and double-filter binding when measuring extremely tight protein/nucleic acid interactions with sub-nanomolar binding affinities. We include specific examples from two telomere end-binding protein systems, Schizo -saccharomyces pombe Pot1 and Saccharomyces cerevisiae Cdc13, to demonstrate potential experimental pitfalls and some useful strategies for optimization.

  15. Practical strategies for the evaluation of high-affinity protein/nucleic acid interactions

    Directory of Open Access Journals (Sweden)

    Sarah E. Altschuler

    2013-09-01

    Full Text Available The quantitative evaluation of binding interactions between proteins and nucleic acids is highly sensitive to a variety of experimental conditions. Optimization of these conditions is critical for obtaining high quality, reproducible data, particularly in the context of very high affinity interactions. Here, we discuss the practical considerations involved in optimizing the apparent binding constant of an interaction as measured by two common quantitative assays, electrophoretic mobility shift assay and double-filter binding when measuring extremely tight protein/nucleic acid interactions with sub-nanomolar binding affinities. We include specific examples from two telomere end-binding protein systems, Schizosaccharomyces pombe Pot1 and Saccharomyces cerevisiae Cdc13, to demonstrate potential experimental pitfalls and some useful strategies for optimization.

  16. An Arabidopsis thaliana high-affinity molybdate transporter required for efficient uptake of molybdate from soil.

    Science.gov (United States)

    Tomatsu, Hajime; Takano, Junpei; Takahashi, Hideki; Watanabe-Takahashi, Akiko; Shibagaki, Nakako; Fujiwara, Toru

    2007-11-20

    Molybdenum (Mo) is a trace element essential for living organisms, however no molybdate transporter has been identified in eukaryotes. Here, we report the identification of a molybdate transporter, MOT1, from Arabidopsis thaliana. MOT1 is expressed in both roots and shoots, and the MOT1 protein is localized, in part, to plasma membranes and to vesicles. MOT1 is required for efficient uptake and translocation of molybdate and for normal growth under conditions of limited molybdate supply. Kinetics studies in yeast revealed that the K(m) value of MOT1 for molybdate is approximately 20 nM. Furthermore, Mo uptake by MOT1 in yeast was not affected by coexistent sulfate, and MOT1 did not complement a sulfate transporter-deficient yeast mutant strain. These data confirmed that MOT1 is specific for molybdate and that the high affinity of MOT1 allows plants to obtain scarce Mo from soil.

  17. Experimental conditions can obscure the second high-affinity site in LeuT.

    Science.gov (United States)

    Quick, Matthias; Shi, Lei; Zehnpfennig, Britta; Weinstein, Harel; Javitch, Jonathan A

    2012-01-15

    Neurotransmitter:Na(+) symporters (NSSs), the targets of antidepressants and psychostimulants, recapture neurotransmitters from the synapse in a Na(+)-dependent symport mechanism. The crystal structure of the NSS homolog LeuT from Aquifex aeolicus revealed one leucine substrate in an occluded, centrally located (S1) binding site next to two Na(+) ions. Computational studies combined with binding and flux experiments identified a second substrate (S2) site and a molecular mechanism of Na(+)-substrate symport that depends upon the allosteric interaction of substrate molecules in the two high-affinity sites. Here we show that the S2 site, which has not yet been identified by crystallographic approaches, can be blocked during preparation of detergent-solubilized LeuT, thereby obscuring its crucial role in Na(+)-coupled symport. This finding points to the need for caution in selecting experimental environments in which the properties and mechanistic features of membrane proteins can be delineated.

  18. Cytisine derivatives as high affinity nAChR ligands: synthesis and comparative molecular field analysis.

    Science.gov (United States)

    Nicolotti, O; Canu Boido, C; Sparatore, F; Carotti, A

    2002-06-01

    A number of new N-substituted cytisine derivatives were prepared and tested, along with similar compounds already described by us and others, as high affinity neuronal acetylcholine receptor ligands. Structure-affinity relationships were discussed in the light of our recently proposed pharmacophore model for nicotinic receptor agonists. The most significant physicochemical interactions modulating the receptor-ligand binding were detected at the three dimensional (3D) level by means of comparative molecular field analysis (CoMFA). The best predictive PLS model was a single-field steric model showing good statistical figures: n = 17, Q2 = 0.717, s(ev) = 0.566, r2 = 0.942, s = 0.275.

  19. A linker peptide with high affinity towards silica-containing materials.

    Science.gov (United States)

    Sunna, Anwar; Chi, Fei; Bergquist, Peter L

    2013-06-25

    A peptide sequence with affinity to silica-containing materials was fused to a truncated form of Streptococcus strain G148 Protein G. The resulting recombinant Linker-Protein G (LPG) was produced in Escherichia coli and purified to apparent homogeneity. It displayed high affinity towards two natural clinoptilolite zeolites. The LPG also displayed high binding affinity towards commercial-grade synthetic zeolite, silica and silica-containing materials. A commercial sample of the truncated Protein G and a basic protein, both without the linker, did not bind to natural or synthetic zeolites or silica. We conclude that the zeolite-binding affinity is mediated by the linker peptide sequence. As a consequence, these data may imply that the binding affinity is directed to the SiO2 component rather than to the atomic orientation on the zeolite crystal surface as previously assumed.

  20. Structural insights into a high affinity nanobody:antigen complex by homology modelling

    DEFF Research Database (Denmark)

    Skottrup, Peter Durand

    2017-01-01

    B binding were identified and used as input to the docking. Furthermore, residues likely involved in the RgpB epitope was identified based upon RgpB:RgpA alignment and analysis of residue surface accessibility. CDR residues and putitative RgpB epitope residues were used as input to an information-driven...... flexible docking approach using the HADDOCK server. Analysis of the VHH7:RgpB model demonstrated that the epitope was found in the immunoglobulin-like domain and residue pairs located at the molecular paratope:epitope interface important for complex stability was identified. Collectively, the VHH7 homology...... model and VHH7:RgpB docking supplies knowledge of the residues involved in the high affinity interaction. This information could prove valuable in the design of an antibody-drug conjugate for specific RgpB targeting....

  1. A complex water network contributes to high-affinity binding in an antibody–antigen interface

    Directory of Open Access Journals (Sweden)

    S.F. Marino

    2016-03-01

    Full Text Available This data article presents an analysis of structural water molecules in the high affinity interaction between a potent tumor growth inhibiting antibody (fragment, J22.9-xi, and the tumor marker antigen CD269 (B cell maturation antigen, BCMA. The 1.89 Å X-ray crystal structure shows exquisite details of the binding interface between the two molecules, which comprises relatively few, mostly hydrophobic, direct contacts but many indirect interactions over solvent waters. These are partly or wholly buried in, and therefore part of, the interface. A partial description of the structure is included in an article on the tumor inhibiting effects of the antibody: “Potent anti-tumor response by targeting B cell maturation antigen (BCMA in a mouse model of multiple myeloma”, Mol. Oncol. 9 (7 (2015 pp. 1348–58.

  2. Expression of a Hybrid Human Superoxide Dismutase with a High Affinity for Heparin

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    A designed heparin-affinity of human Cu, Zn-SOD is described. The natural leader peptide of P.leiognathi Cu, Zn-SOD and a heparin-binding peptide containing a stretch of 7 Arg were fused to the N-terminal and the C-terminal of human Cu, Zn-SOD respectively. The resulted hybrid enzyme had not only a normal SOD activity but also a high affinity for heparin eluted on the heparin-Sepharose column at 0.4 mol/L NaCl. Some properties, such as the optimum pH, the thermostability and the half-life in the circulation of rats, were also analyzed.

  3. Effects of anticonvulsants in vivo on high affinity choline uptake in vitro in mouse hippocampal synaptosomes.

    Science.gov (United States)

    Miller, J. A.; Richter, J. A.

    1985-01-01

    The effects of several anticonvulsant drugs on sodium-dependent high affinity choline uptake (HACU) in mouse hippocampal synaptosomes was investigated. HACU was measured in vitro after in vivo administration of the drug to mice. HACU was inhibited by drugs which have in common the ability to facilitate gamma-aminobutyric acid (GABA) transmission, pentobarbitone, phenobarbitone, barbitone, diazepam, chloridiazepoxide, and valproic acid. Dose-response relationships were determined for these drugs and the drugs' potencies at inhibiting HACU correlated well with their anticonvulsant potencies. Clonazepam, ethosuximide, carbamazepine, and barbituric acid had no effect on HACU in the doses used while phenytoin and trimethadione stimulated HACU. These results suggest that certain anticonvulsants may elicit a part of their anticonvulsant activity by modulating cholinergic neurones. This effect may be mediated through a GABA mechanism. PMID:3978310

  4. Kinetics and autoradiography of high affinity uptake of serotonin by primary astrocyte cultures

    Energy Technology Data Exchange (ETDEWEB)

    Katz, D.M.; Kimelberg, H.K.

    1985-07-01

    Primary astrocyte cultures prepared from the cerebral cortices of neonatal rats showed significant accumulation of serotonin (5-hydroxytryptamine; (/sup 3/H)-5-HT). At concentrations in the range of 0.01 to 0.7 microM (/sup 3/H)-5-HT, this uptake was 50 to 85% Na+ dependent and gave a Km of 0.40 +/- 0.11 microM (/sup 3/H)-5-HT and a Vmax of 6.42 +/- 0.85 (+/- SEM) pmol of (/sup 3/H)-5-HT/mg of protein/4 min for the Na+-dependent component. In the absence of Na+ the uptake was nonsaturable. Omission of the monoamine oxidase inhibitor pargyline markedly reduced the Na+-dependent component of (/sup 3/H)-5-HT uptake but had a negligible effect on the Na+-independent component. This suggest significant oxidative deamination of serotonin after it has been taken up by the high affinity system, followed by release of its metabolite. The authors estimated that this system enabled the cells to concentrate (/sup 3/H)-5-HT up to 44-fold at an external (/sup 3/H)-5-HT concentration of 10(-7) M. Inhibition of (/sup 3/H)-5-HT uptake by a number of clinically effective antidepressants was also consistent with a specific high affinity uptake mechanism for 5-HT, the order of effectiveness of inhibition being chlorimipramine greater than fluoxetine greater than imipramine = amitriptyline greater than desmethylimipramine greater than iprindole greater than mianserin. Uptake of (/sup 3/H)-5-HT was dependent on the presence of Cl- as well as Na+ in the medium, and the effect of omission of both ions was nonadditive. Varying the concentration of K+ in the media from 1 to 50 mM had a limited effect on (/sup 3/H)-5-HT uptake.

  5. The integration of genomic and structural information in the development of high affinity plasmepsin inhibitors.

    Science.gov (United States)

    Nezami, Azin; Freire, Ernesto

    2002-12-04

    The plasmepsins are key enzymes in the life cycle of the Plasmodium parasites responsible for malaria. Since plasmepsin inhibition leads to parasite death, these enzymes have been acknowledged to be important targets for the development of new antimalarial drugs. The development of effective plasmepsin inhibitors, however, is compounded by their genomic diversity which gives rise not to a unique target for drug development but to a family of closely related targets. Successful drugs will have to inhibit not one but several related enzymes with high affinity. Structure-based drug design against heterogeneous targets requires a departure from the classic 'lock-and-key' paradigm that leads to the development of conformationally constrained molecules aimed at a single target. Drug molecules designed along those principles are usually rigid and unable to adapt to target variations arising from naturally occurring genetic polymorphisms or drug-induced resistant mutations. Heterogeneous targets need adaptive drug molecules, characterised by the presence of flexible elements at specific locations that sustain a viable binding affinity against existing or expected polymorphisms. Adaptive ligands have characteristic thermodynamic signatures that distinguish them from their rigid counterparts. This realisation has led to the development of rigorous thermodynamic design guidelines that take advantage of correlations between the structure of lead compounds and the enthalpic and entropic components of the binding affinity. In this paper, we discuss the application of the thermodynamic approach to the development of high affinity (K(i) - pM) plasmepsin inhibitors. In particular, a family of allophenylnorstatine-based compounds is evaluated for their potential to inhibit a wide spectrum of plasmepsins.

  6. Selection of DNA aptamers against epidermal growth factor receptor with high affinity and specificity

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Deng-Liang [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Department of Neurosurgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China); Song, Yan-Ling; Zhu, Zhi; Li, Xi-Lan; Zou, Yuan [State Key Laboratory for Physical Chemistry of Solid Surfaces, Key Laboratory for Chemical Biology of Fujian Province, Key Laboratory of Analytical Chemistry, and Department of Chemical Biology, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen 361005 (China); Yang, Hai-Tao; Wang, Jiang-Jie [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Yao, Pei-Sen [Department of Neurosurgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China); Pan, Ru-Jun [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Yang, Chaoyong James, E-mail: cyyang@xmu.edu.cn [State Key Laboratory for Physical Chemistry of Solid Surfaces, Key Laboratory for Chemical Biology of Fujian Province, Key Laboratory of Analytical Chemistry, and Department of Chemical Biology, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen 361005 (China); Kang, De-Zhi, E-mail: kdzy99988@163.com [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Department of Neurosurgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China)

    2014-10-31

    Highlights: • This is the first report of DNA aptamer against EGFR in vitro. • Aptamer can bind targets with high affinity and selectivity. • DNA aptamers are more stable, cheap and efficient than RNA aptamers. • Our selected DNA aptamer against EGFR has high affinity with K{sub d} 56 ± 7.3 nM. • Our selected DNA aptamer against EGFR has high selectivity. - Abstract: Epidermal growth factor receptor (EGFR/HER1/c-ErbB1), is overexpressed in many solid cancers, such as epidermoid carcinomas, malignant gliomas, etc. EGFR plays roles in proliferation, invasion, angiogenesis and metastasis of malignant cancer cells and is the ideal antigen for clinical applications in cancer detection, imaging and therapy. Aptamers, the output of the systematic evolution of ligands by exponential enrichment (SELEX), are DNA/RNA oligonucleotides which can bind protein and other substances with specificity. RNA aptamers are undesirable due to their instability and high cost of production. Conversely, DNA aptamers have aroused researcher’s attention because they are easily synthesized, stable, selective, have high binding affinity and are cost-effective to produce. In this study, we have successfully identified DNA aptamers with high binding affinity and selectivity to EGFR. The aptamer named TuTu22 with K{sub d} 56 ± 7.3 nM was chosen from the identified DNA aptamers for further study. Flow cytometry analysis results indicated that the TuTu22 aptamer was able to specifically recognize a variety of cancer cells expressing EGFR but did not bind to the EGFR-negative cells. With all of the aforementioned advantages, the DNA aptamers reported here against cancer biomarker EGFR will facilitate the development of novel targeted cancer detection, imaging and therapy.

  7. Bacteriophage lambda terminase: alterations of the high-affinity ATPase affect viral DNA packaging.

    Science.gov (United States)

    Dhar, Alok; Feiss, Michael

    2005-03-18

    DNA packaging by large DNA viruses such as the tailed bacteriophages and the herpesviruses involves DNA translocation into a preformed protein shell, called the prohead. Translocation is driven by an ATP hydrolysis-powered DNA packaging motor. The bacteriophages encode a heterodimeric viral DNA packaging protein, called terminase. The terminases have an ATPase center located in the N terminus of the large subunit implicated in DNA translocation. In previous work with phage lambda, lethal mutations that changed ATP-reactive residues 46 and 84 of gpA, the large terminase subunit, were studied. These mutant enzymes retained the terminase endonuclease and helicase activities, but had severe defects in virion assembly, and lacked the terminase high-affinity ATPase activity. Surprisingly, in the work described here, we found that enzymes with the conservative gpA changes Y46F and Y46A had only mild packaging defects. These mild defects contrast with their profound virion assembly defects. Thus, these mutant enzymes have, in addition to the mild DNA packaging defects, a severe post-DNA packaging defect. In contrast, the gpA K84A enzyme had similar virion assembly and DNA packaging defects. The DNA packaging energy budget, i.e. DNA packaged/ATP hydrolyzed, was unchanged for the mutant enzymes, indicating that DNA translocation is tightly coupled to ATP hydrolysis. A model is proposed in which gpA residues 46 and 84 are important for terminase's high-affinity ATPase activity. Assembly of the translocation complex remodels this ATPase so that residues 46 and 84 are not crucial for the activated translocation ATPase. Changing gpA residues 46 and 84 primarily affects assembly, rather than the activity, of the translocation complex.

  8. Neurotensin decreases high affinity [3H]-ouabain binding to cerebral cortex membranes.

    Science.gov (United States)

    Rosin, Carina; Ordieres, María Graciela López; Arnaiz, Georgina Rodríguez de Lores

    2011-12-10

    Previous work from this laboratory showed the ability of neurotensin to inhibit synaptosomal membrane Na(+), K(+)-ATPase activity, the effect being blocked by SR 48692, a non-peptidic antagonist for high affinity neurotensin receptor (NTS1) [López Ordieres and Rodríguez de Lores Arnaiz 2000; 2001]. To further study neurotensin interaction with Na(+), K(+)-ATPase, peptide effect on high affinity [(3)H]-ouabain binding was studied in cerebral cortex membranes. It was observed that neurotensin modified binding in a dose-dependent manner, leading to 80% decrease with 1 × 10(-4)M concentration. On the other hand, the single addition of 1 × 10(-6)M, 1 × 10(-5)M and 1 × 10(-4)M SR 48692 (Sanofi-Aventis, U.S., Inc.) decreased [(3)H]-ouabain binding (in %) to 87 ± 16; 74 ± 16 and 34 ± 17, respectively. Simultaneous addition of neurotensin and SR 48692 led to additive or synergic effects. Partial NTS2 agonist levocabastine inhibited [(3)H]-ouabain binding likewise. Saturation assays followed by Scatchard analyses showed that neurotensin increased K(d) value whereas failed to modify B(max) value, indicating a competitive type interaction of the peptide at Na(+), K(+)-ATPase ouabain site. At variance, SR 48692 decreased B(max) value whereas it did not modify K(d) value. [(3)H]-ouabain binding was also studied in cerebral cortex membranes obtained from rats injected i. p. 30 min earlier with 100 μg and 250 μg/kg SR 48692. It was observed that the 250 μg/kg SR 48692 dose led to 19% decrease in basal [(3)H]-ouabain binding. After SR 48692 treatments, addition of 1 × 10(-6)M led to additive or synergic effect. Results suggested that [(3)H]-ouabain binding inhibition by neurotensin hardly involves NTS1 receptor.

  9. New Insights in Nutritional Management and Amino Acid Supplementation in Urea Cycle Disorders

    OpenAIRE

    Scaglia, Fernando

    2010-01-01

    Sodium phenylbutyrate is used in the pharmacological treatment of urea cycle disorders to create alternative pathways for nitrogen excretion. The primary metabolite, phenylacetate, conjugates glutamine in the liver and kidney to form phenylacetylglutamine that is readily excreted in the urine. Patients with urea cycle disorders taking sodium phenylbutyrate have a selective reduction in the plasma concentrations of branched chain amino acids despite adequate dietary protein intake. Moreover, t...

  10. Amino Acid Metabolism of Thermoanaerobacter Strain AK90: The Role of Electron-Scavenging Systems in End Product Formation

    Directory of Open Access Journals (Sweden)

    Sean Michael Scully

    2015-01-01

    Full Text Available The catabolism of the 20 amino acids by Thermoanaerobacter strain AK90 (KR007667 was investigated under three different conditions: as single amino acids without an electron-scavenging system, in the presence of thiosulfate, and in coculture with a hydrogenotrophic methanogen. The strain degraded only serine without an alternative electron acceptor but degraded 11 amino acids (alanine, cysteine, isoleucine, leucine, lysine, methionine, phenylalanine, serine, threonine, tyrosine, and valine under both of the electron-scavenging systems investigated. Acetate was the dominant end product from alanine, cysteine, lysine, serine, and threonine under electron-scavenging conditions. The branched-chain amino acids, isoleucine, leucine, and valine, were degraded to their corresponding fatty acids under methanogenic conditions and to a mixture of their corresponding fatty acids and alcohols in the presence of thiosulfate. The partial pressure of hydrogen seems to be of importance for the branched-chain alcohol formation. This was suggested by low but detectable hydrogen concentrations at the end of cultivation on the branched-chain amino acid in the presence of thiosulfate but not when cocultured with the methanogen. A more detailed examination of the role of thiosulfate as an electron acceptor was performed with Thermoanaerobacter ethanolicus (DSM 2246 and Thermoanaerobacter brockii (DSM 1457.

  11. Gene Structure and Expression of the High-affinity Nitrate Transport System in Rice Roots

    Institute of Scientific and Technical Information of China (English)

    Chao Cai; Jun-Yi Wang; Yong-Guan Zhu; Qi-Rong Shen; Bin Li; Yi-Ping Tong; Zhen-Sheng Li

    2008-01-01

    Rice has a preference for uptake of ammonium over nitrate and can use ammonium-N efficiently. Consequently, transporters mediating ammonium uptake have been extensively studied, but nitrate transporters have been largely ignored. Recently,some reports have shown that rice also has high capacity to acquire nitrate from growth medium, so understanding the nitrate transport system in rice roots is very important for improving N use efficiency in rice. The present study identified four putative NRT2 and two putative NAR2 genes that encode components of the high-affinity nitrate transport system (HATS) in the rice (Oryza sativa L. subsp, japonica cv. Nipponbare) genome. OsNRT2.1 and OsNRT2.2 share an identical coding region sequence, and their deduced proteins are closely related to those from monocotyledonous plants. The two NAR2 proteins are closely related to those from mono-cotyledonous plants as well. However, OsNRT2.3 and OsNRT2.4 are more closely related to Arabidopsis NRT2 proteins. Relative quantitative reverse tranecdption-polymerase chain reaction analysis showed that all of the six genes were rapidly upregulated and then downregulated in the roots of N-starved rice plants after they were re-supplied with 0.2 mM nitrate, but the response to nitrate differed among gene members.The results from phylogenetic tree, gene structure and expression analysis implied the divergent roles for the individual members of the rice NRT2 and NAR2 families. High-affinity nitrate influx rates associated with nitrate induction in rice roots were investigated and were found to be regulated by external pH. Compared with the nitrate influx rates at pH 6.5, alkaline pH (pH 8.0) inhibited nitrate Influx, and acidic pH (pH 5.0) enhanced the nitrate influx In I h nitrate induced roots, but did not significantly affect that in 4 to 8 h nitrate induced roots.

  12. Is there in vivo evidence for amino acid shuttles carrying ammonia from neurons to astrocytes?

    Science.gov (United States)

    Rothman, Douglas L; De Feyter, Henk M; Maciejewski, Paul K; Behar, Kevin L

    2012-11-01

    The high in vivo flux of the glutamate/glutamine cycle puts a strong demand on the return of ammonia released by phosphate activated glutaminase from the neurons to the astrocytes in order to maintain nitrogen balance. In this paper we review several amino acid shuttles that have been proposed for balancing the nitrogen flows between neurons and astrocytes in the glutamate/glutamine cycle. All of these cycles depend on the directionality of glutamate dehydrogenase, catalyzing reductive glutamate synthesis (forward reaction) in the neuron in order to capture the ammonia released by phosphate activated glutaminase, while catalyzing oxidative deamination of glutamate (reverse reaction) in the astrocytes to release ammonia for glutamine synthesis. Reanalysis of results from in vivo experiments using (13)N and (15)N labeled ammonia and (15)N leucine in rats suggests that the maximum flux of the alanine/lactate or branched chain amino acid/branched chain amino acid transaminase shuttles between neurons and astrocytes are approximately 3-5 times lower than would be required to account for the ammonia transfer from neurons to astrocytes needed for glutamine synthesis (amide nitrogen) to sustain the glutamate/glutamine cycle. However, in the rat brain both the total ammonia fixation rate by glutamate dehydrogenase and the total branched chain amino acid transaminase activity are sufficient to support a branched chain amino acid/branched chain keto acid shuttle, as proposed by Hutson and coworkers, which would support the de novo synthesis of glutamine in the astrocyte to replace the ~20 % of neurotransmitter glutamate that is oxidized. A higher fraction of the nitrogen needs of total glutamate neurotransmitter cycling could be supported by hybrid cycles in which glutamate and tricarboxylic acid cycle intermediates act as a nitrogen shuttle. A limitation of all in vivo studies in animals conducted to date is that none have shown transfer of nitrogen for glutamine amide

  13. New Synthesis and Tritium Labeling of a Selective Ligand for Studying High-affinity γ-Hydroxybutyrate (GHB) Binding Sites

    Science.gov (United States)

    Vogensen, Stine B.; Marek, Aleš; Bay, Tina; Wellendorph, Petrine; Kehler, Jan; Bundgaard, Christoffer; Frølund, Bente; Pedersen, Martin H.F.; Clausen, Rasmus P.

    2013-01-01

    3-Hydroxycyclopent-1-enecarboxylic acid (HOCPCA, 1) is a potent ligand for the high-affinity GHB binding sites in the CNS. An improved synthesis of 1 together with a very efficient synthesis of [3H]-1 is described. The radiosynthesis employs in situ generated lithium trimethoxyborotritide. Screening of 1 against different CNS targets establishes a high selectivity and we demonstrate in vivo brain penetration. In vitro characterization of [3H]-1 binding shows high specificity to the high-affinity GHB binding sites. PMID:24053696

  14. New synthesis and tritium labeling of a selective ligand for studying high-affinity γ-hydroxybutyrate (GHB) binding sites.

    Science.gov (United States)

    Vogensen, Stine B; Marek, Aleš; Bay, Tina; Wellendorph, Petrine; Kehler, Jan; Bundgaard, Christoffer; Frølund, Bente; Pedersen, Martin H F; Clausen, Rasmus P

    2013-10-24

    3-Hydroxycyclopent-1-enecarboxylic acid (HOCPCA, 1) is a potent ligand for the high-affinity GHB binding sites in the CNS. An improved synthesis of 1 together with a very efficient synthesis of [(3)H]-1 is described. The radiosynthesis employs in situ generated lithium trimethoxyborotritide. Screening of 1 against different CNS targets establishes a high selectivity, and we demonstrate in vivo brain penetration. In vitro characterization of [(3)H]-1 binding shows high specificity to the high-affinity GHB binding sites.

  15. New Synthesis and Tritium Labeling of a Selective Ligand for Studying High-Affinity γ-Hydroxybutyrate (GHB) Binding Sites

    DEFF Research Database (Denmark)

    Vogensen, Stine B.; Marek, Ales; Bay, Tina

    2013-01-01

    3-Hydroxycyclopent-1-enecarboxylic acid (HOCPCA, 1) is a potent ligand for the high-affinity GHB binding sites in the CNS. An improved synthesis of 1 together with a very efficient synthesis of [3H]-1 is described. The radiosynthesis employs in situ generated lithium trimethoxyborotritide....... Screening of 1 against different CNS targets establishes a high selectivity, and we demonstrate in vivo brain penetration. In vitro characterization of [3H]-1 binding shows high specificity to the high-affinity GHB binding sites....

  16. Prevalence and mutation analysis of short/branched chain acyl-CoA dehydrogenase deficiency (SBCADD) detected on newborn screening in Wisconsin.

    Science.gov (United States)

    Van Calcar, Sandra C; Baker, Mei W; Williams, Phillip; Jones, Susan A; Xiong, Blia; Thao, Mai Choua; Lee, Sheng; Yang, Mai Khou; Rice, Greg M; Rhead, William; Vockley, Jerry; Hoffman, Gary; Durkin, Maureen S

    2013-01-01

    Short/branched chain acyl-CoA dehydrogenase deficiency (SBCADD), also called 2-methylbutyryl CoA dehydrogenase deficiency (2-MBCDD), is a disorder of l-isoleucine metabolism of uncertain clinical significance. SBCADD is inadvertently detected on expanded newborn screening by elevated 2-methylbutyrylcarnitine (C5), which has the same mass to charge (m/s) on tandem mass spectrometry (MS/MS) as isovalerylcarnitine (C5), an analyte that is elevated in isovaleric acidemia (IVA), a disorder in leucine metabolism. SBCADD cases identified in the Hmong-American population have been found in association with the c.1165 A>G mutation in the ACADSB gene. The purposes of this study were to: (a) estimate the prevalence of SBCADD and carrier frequency of the c.1165 A>G mutation in the Hmong ethnic group; (b) determine whether the c.1165 A>G mutation is common to all Hmong newborns screening positive for SBCADD; and (c) evaluate C5 acylcarnitine cut-off values to detect and distinguish between SBCADD and IVA diagnoses. During the first 10years of expanded newborn screening using MS/MS in Wisconsin (2001-2011), 97 infants had elevated C5 values (≥0.44μmol/L), of whom five were Caucasian infants confirmed to have IVA. Of the remaining 92 confirmed SBCADD cases, 90 were of Hmong descent. Mutation analysis was completed on an anonymous, random sample of newborn screening cards (n=1139) from Hmong infants. Fifteen infants, including nine who had screened positive for SBCADD based on a C5 acylcarnitine concentration ≥0.44μmol/L, were homozygous for the c.1165 A>G mutation. This corresponds to a prevalence in this ethnic group of being homozygous for the mutation of 1.3% (95% confidence interval 0.8-2.2%) and of being heterozygous for the mutation of 21.8% (95% confidence interval 19.4-24.3%), which is consistent with the Hardy-Weinberg equilibrium. Detection of homozygous individuals who were not identified on newborn screening suggests that the C5 screening cut-off would need to

  17. Rapid and efficient cancer cell killing mediated by high-affinity death receptor homotrimerizing TRAIL variants.

    Science.gov (United States)

    Reis, C R; van der Sloot, A M; Natoni, A; Szegezdi, E; Setroikromo, R; Meijer, M; Sjollema, K; Stricher, F; Cool, R H; Samali, A; Serrano, L; Quax, W J

    2010-10-21

    The tumour necrosis factor family member TNF-related apoptosis-inducing ligand (TRAIL) selectively induces apoptosis in a variety of cancer cells through the activation of death receptors 4 (DR4) and 5 (DR5) and is considered a promising anticancer therapeutic agent. As apoptosis seems to occur primarily via only one of the two death receptors in many cancer cells, the introduction of DR selectivity is thought to create more potent TRAIL agonists with superior therapeutic properties. By use of a computer-aided structure-based design followed by rational combination of mutations, we obtained variants that signal exclusively via DR4. Besides an enhanced selectivity, these TRAIL-DR4 agonists show superior affinity to DR4, and a high apoptosis-inducing activity against several TRAIL-sensitive and -resistant cancer cell lines in vitro. Intriguingly, combined treatment of the DR4-selective variant and a DR5-selective TRAIL variant in cancer cell lines signalling by both death receptors leads to a significant increase in activity when compared with wild-type rhTRAIL or each single rhTRAIL variant. Our results suggest that TRAIL induced apoptosis via high-affinity and rapid-selective homotrimerization of each DR represent an important step towards an efficient cancer treatment.

  18. Cyclic GMP-AMP Containing Mixed Phosphodiester Linkages Is An Endogenous High Affinity Ligand for STING

    Science.gov (United States)

    Zhang, Xu; Shi, Heping; Wu, Jiaxi; Zhang, Xuewu; Sun, Lijun; Chen, Chuo; Chen, Zhijian J.

    2013-01-01

    The presence of microbial or self DNA in the cytoplasm of mammalian cells is a danger signal detected by the DNA sensor cyclic-GMP-AMP (cGAMP) synthase (cGAS), which catalyzes the production of cGAMP that in turn serves as a second messenger to activate innate immune responses. Here we show that endogenous cGAMP in mammalian cells contains two distinct phosphodiester linkages, one between 2′-OH of GMP and 5′-phosphate of AMP, and the other between 3′-OH of AMP and 5′-phosphate of GMP. This molecule, termed 2′3′-cGAMP, is unique in that it binds to the adaptor protein STING with a much greater affinity than cGAMP molecules containing other combinations of phosphodiester linkages. The crystal structure of STING bound to 2′3′-cGAMP revealed the structural basis of this high-affinity binding and a ligand-induced conformational change in STING that may underlie its activation. PMID:23747010

  19. Early signs of pathological cognitive aging in mice lacking high-affinity nicotinic receptors.

    Directory of Open Access Journals (Sweden)

    Eleni eKonsolaki

    2016-04-01

    Full Text Available In order to address pathological cognitive decline effectively, it is critical to adopt early preventive measures in individuals considered at risk. It is therefore essential to develop approaches that identify such individuals before the onset of irreversible dementia. Α deficient cholinergic system has been consistently implicated as one of the main factors associated with a heightened vulnerability to the aging process. In the present study we used mice lacking high affinity nicotinic receptors (β2-/-, which have been proposed as an animal model of accelerated/premature cognitive aging. Our aim was to identify behavioural signs that could serve as indicators or predictors of impending cognitive decline. We used test batteries in order to assess cognitive functions and additional tasks to investigate spontaneous behaviours, such as species-specific activities and exploration/locomotion in a novel environment. Our data confirm and extend the hypothesis that β2-/- animals exhibit age-related cognitive impairments, manifested in both spatial learning and recognition memory tasks. In addition, we reveal deficits in spontaneous behaviour and habituation processes earlier in life. To our knowledge, this is the first study to perform an extensive behavioural examination of an animal model of premature cognitive aging, and our results suggest that β2-nAChR dependent cognitive deterioration progressively evolves from initial subtle behavioural changes to global dementia due to the combined effect of the neuropathology and aging.

  20. Glycation of the high affinity NGF-receptor and RAGE leads to reduced ligand affinity.

    Science.gov (United States)

    Bennmann, Dorit; Kannicht, Christoph; Fisseau, Claudine; Jacobs, Kathleen; Navarette-Santos, Alexander; Hofmann, Britt; Horstkorte, Rüdiger

    2015-09-01

    AGEs are posttranslational modifications generated by irreversible non-enzymatic crosslinking reactions between sugars and proteins - a reaction referred to as glycation. Glycation, a feature of ageing, can lead to non-degradable and less functional proteins and enzymes and can additionally induce inflammation and further pathophysiological processes such as neurodegeneration. In this study we investigated the influence of glycation on the high affinity NGF-receptor TrkA and the AGE-receptor RAGE. We quantified the binding affinity of the TrkA-receptor and RAGE to their ligands by surface plasmon resonance (SPR) and compared these to the binding affinity after glycation. At the same time, we established a glycation procedure using SPR. We found that glycation of TrkA reduced the affinity to NGF by a factor of three, which could be shown to lead to a reduction of NGF-dependent neurite outgrowth in PC12 cells. Glycation of RAGE reduced binding affinity of AGEs by 10-fold.

  1. High-affinity cannabinoid binding site in brain: A possible marijuana receptor

    Energy Technology Data Exchange (ETDEWEB)

    Nye, J.S.

    1988-01-01

    The mechanism by which delta{sup 9} tetrahydrocannabinol (delta{sup 9}THC), the major psychoactive component of marijuana or hashish, produces its potent psychological and physiological effects is unknown. To find receptor binding sites for THC, we designed a water-soluble analog for use as a radioligand. 5{prime}-Trimethylammonium-delta{sup 8}THC (TMA) is a positively charged analog of delta-{sup 8}THC modified on the 5{prime} carbon, a portion of the molecule not important for its psychoactivity. We have studied the binding of ({sup 3}H)-5{prime}-trimethylammonium-delta-{sup 8}THC (({sup 3}H)TMA) to rat neuronal membranes. ({sup 3}H)TMA binds saturably and reversibly to brain membranes with high affinity to apparently one class of sites. Highest binding site density occurs in brain, but several peripheral organs also display specific binding. Detergent solubilizes the sites without affecting their pharmacologial properties. Molecular sieve chromatography reveals a bimodal peak of ({sup 3}H)TMA binding activity of approximately 60,000 daltons apparent molecular weight.

  2. Affinity Crystallography: A New Approach to Extracting High-Affinity Enzyme Inhibitors from Natural Extracts.

    Science.gov (United States)

    Aguda, Adeleke H; Lavallee, Vincent; Cheng, Ping; Bott, Tina M; Meimetis, Labros G; Law, Simon; Nguyen, Nham T; Williams, David E; Kaleta, Jadwiga; Villanueva, Ivan; Davies, Julian; Andersen, Raymond J; Brayer, Gary D; Brömme, Dieter

    2016-08-26

    Natural products are an important source of novel drug scaffolds. The highly variable and unpredictable timelines associated with isolating novel compounds and elucidating their structures have led to the demise of exploring natural product extract libraries in drug discovery programs. Here we introduce affinity crystallography as a new methodology that significantly shortens the time of the hit to active structure cycle in bioactive natural product discovery research. This affinity crystallography approach is illustrated by using semipure fractions of an actinomycetes culture extract to isolate and identify a cathepsin K inhibitor and to compare the outcome with the traditional assay-guided purification/structural analysis approach. The traditional approach resulted in the identification of the known inhibitor antipain (1) and its new but lower potency dehydration product 2, while the affinity crystallography approach led to the identification of a new high-affinity inhibitor named lichostatinal (3). The structure and potency of lichostatinal (3) was verified by total synthesis and kinetic characterization. To the best of our knowledge, this is the first example of isolating and characterizing a potent enzyme inhibitor from a partially purified crude natural product extract using a protein crystallographic approach.

  3. High Affinity Antibodies against Influenza Characterize the Plasmablast Response in SLE Patients After Vaccination.

    Directory of Open Access Journals (Sweden)

    Kaval Kaur

    Full Text Available Breakdown of B cell tolerance is a cardinal feature of systemic lupus erythematosus (SLE. Increased numbers of autoreactive mature naïve B cells have been described in SLE patients and autoantibodies have been shown to arise from autoreactive and non-autoreactive precursors. How these defects, in the regulation of B cell tolerance and selection, influence germinal center (GC reactions that are directed towards foreign antigens has yet to be investigated. Here, we examined the characteristics of post-GC foreign antigen-specific B cells from SLE patients and healthy controls by analyzing monoclonal antibodies generated from plasmablasts induced specifically by influenza vaccination. We report that many of the SLE patients had anti-influenza antibodies with higher binding affinity and neutralization capacity than those from controls. Although overall frequencies of autoreactivity in the influenza-specific plasmablasts were similar for SLE patients and controls, the variable gene repertoire of influenza-specific plasmablasts from SLE patients was altered, with increased usage of JH6 and long heavy chain CDR3 segments. We found that high affinity anti-influenza antibodies generally characterize the plasmablast responses of SLE patients with low levels of autoreactivity; however, certain exceptions were noted. The high-avidity antibody responses in SLE patients may also be correlated with cytokines that are abnormally expressed in lupus. These findings provide insights into the effects of dysregulated immunity on the quality of antibody responses following influenza vaccination and further our understanding of the underlying abnormalities of lupus.

  4. High Affinity Antibodies against Influenza Characterize the Plasmablast Response in SLE Patients After Vaccination

    Science.gov (United States)

    Kaur, Kaval; Zheng, Nai-Ying; Smith, Kenneth; Huang, Min; Li, Lie; Pauli, Noel T.; Henry Dunand, Carole J.; Lee, Jane-Hwei; Morrissey, Michael; Wu, Yixuan; Joachims, Michelle L.; Munroe, Melissa E.; Lau, Denise; Qu, Xinyan; Krammer, Florian; Wrammert, Jens; Palese, Peter; Ahmed, Rafi; James, Judith A.; Wilson, Patrick C.

    2015-01-01

    Breakdown of B cell tolerance is a cardinal feature of systemic lupus erythematosus (SLE). Increased numbers of autoreactive mature naïve B cells have been described in SLE patients and autoantibodies have been shown to arise from autoreactive and non-autoreactive precursors. How these defects, in the regulation of B cell tolerance and selection, influence germinal center (GC) reactions that are directed towards foreign antigens has yet to be investigated. Here, we examined the characteristics of post-GC foreign antigen-specific B cells from SLE patients and healthy controls by analyzing monoclonal antibodies generated from plasmablasts induced specifically by influenza vaccination. We report that many of the SLE patients had anti-influenza antibodies with higher binding affinity and neutralization capacity than those from controls. Although overall frequencies of autoreactivity in the influenza-specific plasmablasts were similar for SLE patients and controls, the variable gene repertoire of influenza-specific plasmablasts from SLE patients was altered, with increased usage of JH6 and long heavy chain CDR3 segments. We found that high affinity anti-influenza antibodies generally characterize the plasmablast responses of SLE patients with low levels of autoreactivity; however, certain exceptions were noted. The high-avidity antibody responses in SLE patients may also be correlated with cytokines that are abnormally expressed in lupus. These findings provide insights into the effects of dysregulated immunity on the quality of antibody responses following influenza vaccination and further our understanding of the underlying abnormalities of lupus. PMID:25951191

  5. Lymphocyte crawling and transendothelial migration require chemokine triggering of high-affinity LFA-1 integrin.

    Science.gov (United States)

    Shulman, Ziv; Shinder, Vera; Klein, Eugenia; Grabovsky, Valentin; Yeger, Orna; Geron, Erez; Montresor, Alessio; Bolomini-Vittori, Matteo; Feigelson, Sara W; Kirchhausen, Tomas; Laudanna, Carlo; Shakhar, Guy; Alon, Ronen

    2009-03-20

    Endothelial chemokines are instrumental for integrin-mediated lymphocyte adhesion and transendothelial migration (TEM). By dissecting how chemokines trigger lymphocyte integrins to support shear-resistant motility on and across cytokine-stimulated endothelial barriers, we found a critical role for high-affinity (HA) LFA-1 integrin in lymphocyte crawling on activated endothelium. Endothelial-presented chemokines triggered HA-LFA-1 and adhesive filopodia at numerous submicron dots scattered underneath crawling lymphocytes. Shear forces applied to endothelial-bound lymphocytes dramatically enhanced filopodia density underneath crawling lymphocytes. A fraction of the adhesive filopodia invaded the endothelial cells prior to and during TEM and extended large subluminal leading edge containing dots of HA-LFA-1 occupied by subluminal ICAM-1. Memory T cells generated more frequent invasive filopodia and transmigrated more rapidly than their naive counterparts. We propose that shear forces exerted on HA-LFA-1 trigger adhesive and invasive filopodia at apical endothelial surfaces and thereby promote lymphocyte crawling and probing for TEM sites.

  6. High-affinity DNA base analogs as supramolecular, nanoscale promoters of macroscopic adhesion.

    Science.gov (United States)

    Anderson, Cyrus A; Jones, Amanda R; Briggs, Ellen M; Novitsky, Eric J; Kuykendall, Darrell W; Sottos, Nancy R; Zimmerman, Steven C

    2013-05-15

    Adhesion phenomena are essential to many biological processes and to synthetic adhesives and manufactured coatings and composites. Supramolecular interactions are often implicated in various adhesion mechanisms. Recently, supramolecular building blocks, such as synthetic DNA base-pair mimics, have drawn attention in the context of molecular recognition, self-assembly, and supramolecular polymers. These reversible, hydrogen-bonding interactions have been studied extensively for their adhesive capabilities at the nano- and microscale, however, much less is known about their utility for practical adhesion in macroscopic systems. Herein, we report the preparation and evaluation of supramolecular coupling agents based on high-affinity, high-fidelity quadruple hydrogen-bonding units (e.g., DAN·DeUG, Kassoc = 10(8) M(-1) in chloroform). Macroscopic adhesion between polystyrene films and glass surfaces modified with 2,7-diamidonaphthyridine (DAN) and ureido-7-deazaguanine (DeUG) units was evaluated by mechanical testing. Structure-property relationships indicate that the designed supramolecular interaction at the nanoscale plays a key role in the observed macroscopic adhesive response. Experiments probing reversible adhesion or self-healing properties of bulk samples indicate that significant recovery of initial strength can be realized after failure but that the designed noncovalent interaction does not lead to healing during the process of adhesion loss.

  7. Dynein and dynactin leverage their bivalent character to form a high-affinity interaction.

    Directory of Open Access Journals (Sweden)

    Amanda E Siglin

    Full Text Available Cytoplasmic dynein and dynactin participate in retrograde transport of organelles, checkpoint signaling and cell division. The principal subunits that mediate this interaction are the dynein intermediate chain (IC and the dynactin p150(Glued; however, the interface and mechanism that regulates this interaction remains poorly defined. Herein, we use multiple methods to show the N-terminus of mammalian dynein IC, residues 10-44, is sufficient for binding p150(Glued. Consistent with this mapping, monoclonal antibodies that antagonize the dynein-dynactin interaction also bind to this region of the IC. Furthermore, double and triple alanine point mutations spanning residues 6 to 19 in the yeast IC homolog, Pac11, produce significant defects in spindle positioning. Using the same methods we show residues 381 to 530 of p150(Glued form a minimal fragment that binds to the dynein IC. Sedimentation equilibrium experiments indicate that these individual fragments are predominantly monomeric, but admixtures of the IC and p150(Glued fragments produce a 2:2 complex. This tetrameric complex is sensitive to salt, temperature and pH, suggesting that the binding is dominated by electrostatic interactions. Finally, circular dichroism (CD experiments indicate that the N-terminus of the IC is disordered and becomes ordered upon binding p150(Glued. Taken together, the data indicate that the dynein-dynactin interaction proceeds through a disorder-to-order transition, leveraging its bivalent-bivalent character to form a high affinity, but readily reversible interaction.

  8. High affinity anchoring of the decoration protein pb10 onto the bacteriophage T5 capsid

    Science.gov (United States)

    Vernhes, Emeline; Renouard, Madalena; Gilquin, Bernard; Cuniasse, Philippe; Durand, Dominique; England, Patrick; Hoos, Sylviane; Huet, Alexis; Conway, James F.; Glukhov, Anatoly; Ksenzenko, Vladimir; Jacquet, Eric; Nhiri, Naïma; Zinn-Justin, Sophie; Boulanger, Pascale

    2017-01-01

    Bacteriophage capsids constitute icosahedral shells of exceptional stability that protect the viral genome. Many capsids display on their surface decoration proteins whose structure and function remain largely unknown. The decoration protein pb10 of phage T5 binds at the centre of the 120 hexamers formed by the major capsid protein. Here we determined the 3D structure of pb10 and investigated its capsid-binding properties using NMR, SAXS, cryoEM and SPR. Pb10 consists of an α-helical capsid-binding domain and an Ig-like domain exposed to the solvent. It binds to the T5 capsid with a remarkably high affinity and its binding kinetics is characterized by a very slow dissociation rate. We propose that the conformational exchange events observed in the capsid-binding domain enable rearrangements upon binding that contribute to the quasi-irreversibility of the pb10-capsid interaction. Moreover we show that pb10 binding is a highly cooperative process, which favours immediate rebinding of newly dissociated pb10 to the 120 hexamers of the capsid protein. In extreme conditions, pb10 protects the phage from releasing its genome. We conclude that pb10 may function to reinforce the capsid thus favouring phage survival in harsh environments. PMID:28165000

  9. [The high-affinity IgE receptor: lessons from structural analysis].

    Science.gov (United States)

    Blank, Ulrich; Jouvin, Marie-Hélène; Guérin-Marchand, Claudine; Kinet, Jean-Pierre

    2003-01-01

    The high affinity receptor for IgE, FcERI, is at the core of the allergic reaction. This receptor is expressed mainly on mast cells and basophils. Interaction of an allergen with its specific IgE bound to FcERI triggers cell activation, which induces the release of numerous mediators that are responsible for allergic manifestations. The recent increase in the prevalence of allergic diseases in developed countries has resulted in renewed efforts towards the development of new drugs. One of these is a humanised antibody directed against the IgE ligand. This antibody recognises specifically free but not FcERI-bound IgE thus preventing ligand binding and subsequent cell activation. This antibody has shown some efficacy in clinical trials involving patients with asthma and allergic rhinitis. The recent elucidation of the tridimensional structure of the complex between IgE and FcERI provides unexpected information regarding the mechanism of assembly of the complex, which now can be used to design small chemical compounds capable of specifically inhibiting this interaction.

  10. Specific high-affinity binding of fatty acids to epidermal cytosolic proteins

    Energy Technology Data Exchange (ETDEWEB)

    Raza, H.; Chung, W.L.; Mukhtar, H. (Department of Dermatology, University Hospitals of Cleveland, Case Western Reserve University, OH (USA))

    1991-08-01

    Cytosol from rat, mouse, and human skin or rat epidermis was incubated with (3H)arachidonic acid, (14C)retinoic acid, (14C)oleic acid, (3H)leukotriene A4, (3H)prostaglandin E2 (PGE2) or (3H) 15-hydroxyeicosatetraenoic acid (15-HETE), and protein-bound ligands were separated using Lipidex-1000 at 4C to assess the binding specificity. The binding of oleic acid and arachidonic acid with rat epidermal cytosol was rapid, saturable, and reversible. Binding of oleic acid was competed out with the simultaneous addition of other ligands and found to be in the following order: arachidonic acid greater than oleic acid greater than linoleic acid greater than lauric acid greater than leukotriene A4 greater than 15-HETE = PGE1 greater than PGE2 = PGF2. Scatchard analysis of the binding with arachidonic acid, oleic acid, and retinoic acid revealed high-affinity binding sites with the dissociation constant in the nM range. SDS-PAGE analysis of the oleic acid-bound epidermal cytosolic protein(s) revealed maximum binding at the 14.5 kDa region. The presence of the fatty acid-binding protein in epidermal cytosol and its binding to fatty acids and retinoic acid may be of significance both in the trafficking and the metabolism of fatty acids and retinoids across the skin.

  11. Conformation-Dependent High-Affinity Potent Ricin-Neutralizing Monoclonal Antibodies

    Directory of Open Access Journals (Sweden)

    Wei-Gang Hu

    2013-01-01

    Full Text Available Ricin is a potential biothreat agent with no approved antidote available for ricin poisoning. The aim of this study was to develop potent antibody-based antiricin antidotes. Four strong ricin resistant hybridoma clones secreting antiricin monoclonal antibodies (mAbs were developed. All four mAbs are bound to conformational epitopes of ricin toxin B (RTB with high affinity (KD values from 2.55 to 36.27 nM. RTB not only triggers cellular uptake of ricin, but also facilitates transport of the ricin toxin A (RTA from the endoplasmic reticulum to the cytosol, where RTA exerts its toxic activity. The four mAbs were found to have potent ricin-neutralizing capacities and synergistic effects among them as determined by an in vitro neutralization assay. In vivo protection assay demonstrated that all four mAbs had strong efficacy against ricin challenges. D9 was found to be exceptionally effective. Intraperitoneal (i.p. administration of D9, at a dose of 5 μg, 6 weeks before or 6 hours after an i.p. challenge with 5 × LD50 of ricin was able to protect or rescue 100% of the mice, indicating that mAb D9 is an excellent candidate to be developed as a potent antidote against ricin poisoning for both prophylactic and therapeutic purposes.

  12. Sequential administration of the high affinity CXCR4 antagonist BKT140 promotes megakaryopoiesis and platelet production.

    Science.gov (United States)

    Abraham, Michal; Weiss, Ido D; Wald, Hanna; Wald, Ori; Nagler, Arnon; Beider, Katia; Eizenberg, Orly; Peled, Amnon

    2013-10-01

    Platelets are the terminal differentiation product of megakaryocytes (MKs). Cytokines, such as thrombopoietin (TPO), are known to influence different steps in MK development; however, the complex differentiation and platelet localization processes are not fully understood. MKs express the receptor CXCR4 and have been shown to migrate in response to CXCL12 and to increase their platelet production. In this study, we studied the role of CXCR4 in platelet production with the high affinity CXCR4 antagonist, BKT140. Single and sequential administration of BKT140 significantly increased the number of MKs and haematopoietic progenitors (HPCs) within the bone marrow (BM). Increased megakaryopoiesis was associated with increased platelet production. Single and sequential administration of BKT140 also increased the number of HPCs in the blood. In a model of 5-fluorouracil-induced thrombocytopenia, BKT140 significantly reduced the severity and duration of thrombocytopenia and cytopenia when administered before and after chemotherapy. Our results demonstrated that the CXCR4 antagonist, BKT140, mediated unique beneficial effects by stimulating megakaryopoiesis and platelet production. These results provide evidence for the possible therapeutic use of BKT140 for modulating platelet numbers in thrombocytopenic conditions. © 2013 John Wiley & Sons Ltd.

  13. Fc-Binding Ligands of Immunoglobulin G: An Overview of High Affinity Proteins and Peptides

    Directory of Open Access Journals (Sweden)

    Weonu Choe

    2016-12-01

    Full Text Available The rapidly increasing application of antibodies has inspired the development of several novel methods to isolate and target antibodies using smart biomaterials that mimic the binding of Fc-receptors to antibodies. The Fc-binding domain of antibodies is the primary binding site for e.g., effector proteins and secondary antibodies, whereas antigens bind to the Fab region. Protein A, G, and L, surface proteins expressed by pathogenic bacteria, are well known to bind immunoglobulin and have been widely exploited in antibody purification strategies. Several difficulties are encountered when bacterial proteins are used in antibody research and application. One of the major obstacles hampering the use of bacterial proteins is sample contamination with trace amounts of these proteins, which can invoke an immune response in the host. Many research groups actively develop synthetic ligands that are able to selectively and strongly bind to antibodies. Among the reported ligands, peptides that bind to the Fc-domain of antibodies are attractive tools in antibody research. Besides their use as high affinity ligands in antibody purification chromatography, Fc-binding peptides are applied e.g., to localize antibodies on nanomaterials and to increase the half-life of proteins in serum. In this review, recent developments of Fc-binding peptides are presented and their binding characteristics and diverse applications are discussed.

  14. A high-affinity, radioiodinatable neuropeptide FF analogue incorporating a photolabile p-(4-hydroxybenzoyl)phenylalanine.

    Science.gov (United States)

    Bray, Lauriane; Moulédous, Lionel; Tafani, Jean A M; Germanier, Maryse; Zajac, Jean-Marie

    2014-05-15

    A new radioiodinated photoaffinity compound, [(125)I]YE(Bpa)WSLAAPQRFNH2, derived from a peptide present in the rat neuropeptide FF (NPFF) precursor was synthesized, and its binding characteristics were investigated on a neuroblastoma clone, SH-SY5Y, stably expressing rat NPFF2 receptors tagged with the T7 epitope. The binding of the probe was saturable and revealed a high-affinity interaction (KD=0.24nM) with a single class of binding sites. It was also able to affinity label NPFF2 receptor in a specific and efficient manner given that 38% of the bound radioligand at saturating concentration formed a wash-resistant binding after ultraviolet (UV) irradiation. Photoaffinity labeling with [(125)I]YE(Bpa)WSLAAPQRFamide showed two molecular forms of NPFF2 receptor with apparent molecular weights of 140 and 95kDa in a 2:1 ratio. The comparison of the results between photoaffinity labeling and Western blot analysis suggests that all receptor forms bind the probe irreversibly with the same efficiency. On membranes of mouse olfactory bulb, only the high molecular weight form of NPFF2 receptor is observed. [(125)I]YE(Bpa)WSLAAPQRFamide is an excellent radioiodinated peptidic ligand for direct and selective labeling of NPFF2 receptors in vitro.

  15. High affinity germinal center B cells are actively selected into the plasma cell compartment.

    Science.gov (United States)

    Phan, Tri Giang; Paus, Didrik; Chan, Tyani D; Turner, Marian L; Nutt, Stephen L; Basten, Antony; Brink, Robert

    2006-10-30

    A hallmark of T cell-dependent immune responses is the progressive increase in the ability of serum antibodies to bind antigen and provide immune protection. Affinity maturation of the antibody response is thought to be connected with the preferential survival of germinal centre (GC) B cells that have acquired increased affinity for antigen via somatic hypermutation of their immunoglobulin genes. However, the mechanisms that drive affinity maturation remain obscure because of the difficulty in tracking the affinity-based selection of GC B cells and their differentiation into plasma cells. We describe a powerful new model that allows these processes to be followed as they occur in vivo. In contrast to evidence from in vitro systems, responding GC B cells do not undergo plasma cell differentiation stochastically. Rather, only GC B cells that have acquired high affinity for the immunizing antigen form plasma cells. Affinity maturation is therefore driven by a tightly controlled mechanism that ensures only antibodies with the greatest possibility of neutralizing foreign antigen are produced. Because the body can sustain only limited numbers of plasma cells, this "quality control" over plasma cell differentiation is likely critical for establishing effective humoral immunity.

  16. Enhanced membrane pore formation through high-affinity targeted antimicrobial peptides.

    Directory of Open Access Journals (Sweden)

    Christopher J Arnusch

    Full Text Available Many cationic antimicrobial peptides (AMPs target the unique lipid composition of the prokaryotic cell membrane. However, the micromolar activities common for these peptides are considered weak in comparison to nisin, which follows a targeted, pore-forming mode of action. Here we show that AMPs can be modified with a high-affinity targeting module, which enables membrane permeabilization at low concentration. Magainin 2 and a truncated peptide analog were conjugated to vancomycin using click chemistry, and could be directed towards specific membrane embedded receptors both in model membrane systems and whole cells. Compared with untargeted vesicles, a gain in permeabilization efficacy of two orders of magnitude was reached with large unilamellar vesicles that included lipid II, the target of vancomycin. The truncated vancomycin-peptide conjugate showed an increased activity against vancomycin resistant Enterococci, whereas the full-length conjugate was more active against a targeted eukaryotic cell model: lipid II containing erythrocytes. This study highlights that AMPs can be made more selective and more potent against biological membranes that contain structures that can be targeted.

  17. Function and Regulation of the Plant COPT Family of High-Affinity Copper Transport Proteins

    Directory of Open Access Journals (Sweden)

    Sergi Puig

    2014-01-01

    Full Text Available Copper (Cu is an essential micronutrient for all eukaryotes because it participates as a redox active cofactor in multiple biological processes, including mitochondrial respiration, photosynthesis, oxidative stress protection, and iron (Fe transport. In eukaryotic cells, Cu transport toward the cytoplasm is mediated by the conserved CTR/COPT family of high-affinity Cu transport proteins. This outlook paper reviews the contribution of our research group to the characterization of the function played by the Arabidopsis thaliana COPT1–6 family of proteins in plant Cu homeostasis. Our studies indicate that the different tissue specificity, Cu-regulated expression, and subcellular localization dictate COPT-specialized contribution to plant Cu transport and distribution. By characterizing lack-of-function Arabidopsis mutant lines, we conclude that COPT1 mediates root Cu acquisition, COPT6 facilitates shoot Cu distribution, and COPT5 mobilizes Cu from storage organelles. Furthermore, our work with copt2 mutant and COPT-overexpressing plants has also uncovered Cu connections with Fe homeostasis and the circadian clock, respectively. Future studies on the interaction between COPT transporters and other components of the Cu homeostasis network will improve our knowledge of plant Cu acquisition, distribution, regulation, and utilization by Cu-proteins.

  18. Characterization of high affinity binding motifs for the discoidin domain receptor DDR2 in collagen.

    Science.gov (United States)

    Konitsiotis, Antonios D; Raynal, Nicolas; Bihan, Dominique; Hohenester, Erhard; Farndale, Richard W; Leitinger, Birgit

    2008-03-14

    The discoidin domain receptors, DDR1 and DDR2, are receptor tyrosine kinases that are activated by native triple-helical collagen. Here we have located three specific DDR2 binding sites by screening the entire triple-helical domain of collagen II, using the Collagen II Toolkit, a set of overlapping triple-helical peptides. The peptide sequence that bound DDR2 with highest affinity interestingly contained the sequence for the high affinity binding site for von Willebrand factor in collagen III. Focusing on this sequence, we used a set of truncated and alanine-substituted peptides to characterize the sequence GVMGFO (O is hydroxyproline) as the minimal collagen sequence required for DDR2 binding. Based on a recent NMR analysis of the DDR2 collagen binding domain, we generated a model of the DDR2-collagen interaction that explains why a triple-helical conformation is required for binding. Triple-helical peptides comprising the DDR2 binding motif not only inhibited DDR2 binding to collagen II but also activated DDR2 transmembrane signaling. Thus, DDR2 activation may be effected by single triple-helices rather than fibrillar collagen.

  19. PREPARATION AND CHARACTERISTICS OF ANIONIC POLYACRYLAMIDES CONTAINING DIRECT DYE WITH A HIGH AFFINITY FOR CELLULOSE

    Directory of Open Access Journals (Sweden)

    Shingo Yokota

    2009-05-01

    Full Text Available Direct dye with a high affinity for cellulose substrate was utilized as a cellulose anchor to promote retention of paper strengthening additives under various conditions associated with the wet end of a paper machine. Direct Red 28 (DR was covalently linked to anionic polyacrylamide (A-PAM via a condensation reaction using water-soluble carbodiimide. The DR-conjugated A-PAM (DR-A-PAM demonstrated good retention efficiency, resulting in strength enhancement of handsheets. Anionic trash showed no interference with the performance of DR-A-PAM in the wet end, while the additive performance was sensitive to calcium ions. Surface plasmon resonance analysis gave useful information on the cellulose-anchoring ability of DR-A-PAM. Dye molecules were irreversibly adsorbed onto the cellulose substrate under aqueous conditions, while A-PAM possessed no significant affinity for cellulose. These results suggest that anionic DR moieties in DR-A-PAM molecules served as a cellulose-anchor, possibly due to multiple CH-π interaction between hydrophobic face of cellulose substrate and π-conjugated system of dye molecules. Such a unique interaction of direct dye and cellulose provides a new insight into the wet end system, and does not depend on conventional electrostatic attraction.

  20. Age-related changes of muscle and plasma amino acids in healthy children.

    Science.gov (United States)

    Hammarqvist, Folke; Angsten, Gertrud; Meurling, Staffan; Andersson, Kerstin; Wernerman, Jan

    2010-07-01

    The aim of the study was to explore if changes in muscle and plasma amino acid concentrations developed during growth and differed from levels seen in adults. The gradient and concentrations of free amino acids in muscle and plasma were investigated in relation to age in metabolic healthy children. Plasma and specimens from the abdominal muscle were obtained during elective surgery. The children were grouped into three groups (group 1: amino acids analysed increased with age, namely taurine, aspartate, threonine, alanine, valine, isoleucine, leucine, histidine, as well as the total sums of branched chain amino acids (BCAA), basic amino acids (BAA) and total sum of amino acids (P amino acids correlated with age (P < 0.05). These results indicate that there is an age dependency of the amino acid pattern in skeletal muscle and plasma during growth.

  1. The physiological significance of HKT1, a Na{sup +} - coupled high affinity K{sup +} transporter in `Triticum aestivum`

    Energy Technology Data Exchange (ETDEWEB)

    Box, S.; Schachtman, D.P. [University of Adelaide, SA (Australia). Department of Botany

    1997-12-31

    Full text: Several mechanisms for high affinity K{sup +} uptake by higher plants have been proposed:-an ATP-energised K:+ pump, a K{sup +}/H{sup +} antiport and a H{sup +}coupled carrier. Recently, a Na{sup +}--coupled high affinity K{sup +} transporter, HKT1, was isolated from wheat roots. Whilst Na{sup +}K{sup +} symports have been described in charophyte algae, the cloning of HKT1 from wheat is the first, evidence that this type d transport mechanism may function in higher plants. Is the activity of HKT1 an important mechanism involved in K{sup +} acquisition by wheat? The aim of this study was to assess the physiological significance of Na{sup +}- coupled high affinity K{sup +} uptake in T. aestivum. To determine whether HKT1 plays a significant role in wheat growth, we measured the dry weights and ion content of plants grown in a range of [K{sup +}], with and without Na{sup +}. To directly assess the activity of Na{sup +}- coupled K{sup +} transport, {sup 86}Rb{sup +} and {sup 22}Na{sup +} flux analyses were performed on the elongation zones and whole roots of intact seedlings, expressing a high affinity K{sup +} uptake system. The results of these growth and tracer flux studies will be discussed in relation to the expression of the gene encoding HKT1 in T. aestivum

  2. Are basophil histamine release and high affinity IgE receptor expression involved in asymptomatic skin sensitization?

    DEFF Research Database (Denmark)

    Jensen, Bettina Margrethe; Assing, K; Jensen, Lone Hummelshøj;

    2006-01-01

    Immunoglobulin (Ig)E-sensitized persons with positive skin prick test, but no allergy symptoms, are classified as being asymptomatic skin sensitized (AS). The allergic type 1 disease is dependant on IgE binding to the high affinity IgE-receptor (FcepsilonRI) expressed on basophils and mast cells...

  3. Novel high-affinity and selective biaromatic 4-substituted ¿-hydroxybutyric acid (GHB) analogues as GHB ligands

    DEFF Research Database (Denmark)

    Høg, Signe; Wellendorph, Petrine; Nielsen, Birgitte;

    2008-01-01

    Gamma-hydroxybutyrate (GHB) is a metabolite of gamma-aminobutyric acid (GABA) and has been proposed to function as a neurotransmitter or neuromodulator. GHB is used in the treatment of narcolepsy and is a drug of abuse. GHB binds to both GABA(B) receptors and specific high-affinity GHB sites...

  4. A rhodamine-labeled citalopram analogue as a high-affinity fluorescent probe for the serotonin transporter

    DEFF Research Database (Denmark)

    Zhang, Peng; Jørgensen, Trine Nygaard; Løland, Claus Juul

    2013-01-01

    A novel fluorescent ligand was synthesized as a high-affinity, high specificity probe for visualizing the serotonin transporter (SERT). The rhodamine fluorophore was extended from an aniline substitution on the 5-position of the dihydroisobenzofuran ring of citalopram (2, 1-(3-(dimethylamino)prop...

  5. The AFT1 transcriptional factor is differentially required for expression of high-affinity iron uptake genes in Saccharomyces cerevisiae.

    Science.gov (United States)

    Casas, C; Aldea, M; Espinet, C; Gallego, C; Gil, R; Herrero, E

    1997-06-15

    High-affinity iron uptake in Saccharomyces cerevisiae involves the extracytoplasmic reduction of ferric ions by FRE1 and FRE2 reductases. Ferrous ions are then transported across the plasma membrane through the FET3 oxidase-FTR1 permease complex. Expression of the high-affinity iron uptake genes is induced upon iron deprivation. We demonstrate that AFT1 is differentially involved in such regulation. Aft1 protein is required for maintaining detectable non-induced level of FET3 expression and for induction of FRE2 in iron starvation conditions. On the contrary, FRE1 mRNA induction is normal in the absence of Aft1, although the existence of AFT1 point mutations causing constitutive expression of FRE1 (Yamaguchi-Iwai et al., EMBO J. 14: 1231-1239, 1995) indicates that Aft1 may also participate in FRE1 expression in a dispensable way. The alterations in the basal levels of expression of the high-affinity iron uptake genes may explain why the AFT1 mutant is unable to grow on respirable carbon sources. Overexpression of AFT1 leads to growth arrest of the G1 stage of the cell cycle. Aft1 is a transcriptional activator that would be part of the different transcriptional complexes interacting with the promoter of the high-affinity iron uptake genes. Aft1 displays phosphorylation modifications depending on the growth stage of the cells, and it might link induction of genes for iron uptake to other metabolically dominant requirement for cell growth.

  6. High-affinity accumulation of a maytansinoid in cells via weak tubulin interaction.

    Science.gov (United States)

    Goldmacher, Victor S; Audette, Charlene A; Guan, Yinghua; Sidhom, Eriene-Heidi; Shah, Jagesh V; Whiteman, Kathleen R; Kovtun, Yelena V

    2015-01-01

    The microtubule-targeting maytansinoids accumulate in cells and induce mitotic arrest at 250- to 1000-fold lower concentrations than those required for their association with tubulin or microtubules. To identify the mechanisms of this intracellular accumulation and exceptional cytotoxicity of maytansinoids we studied interaction of a highly cytotoxic maytansinoid, S-methyl DM1 and several other maytansinoids with cells. S-methyl DM1 accumulated inside the cells with a markedly higher apparent affinity than to tubulin or microtubules. The apparent affinities of maytansinoids correlated with their cytotoxicities. The number of intracellular binding sites for S-methyl DM1 in MCF7 cells was comparable to the number of tubulin molecules per cell (~ 4-6 × 10(7) copies). Efflux of 3[H]-S-methyl DM1 from cells was enhanced in the presence of an excess of non-labeled S-methyl DM1, indicating that re-binding of 3 [H]-S-methyl DM1 to intracellular binding sites contributed to its intracellular retention. Liposomes loaded with non-polymerized tubulin recapitulated the apparent high-affinity association of S-methyl DM1 to cells. We propose a model for the intracellular accumulation of maytansinoids in which molecules of the compounds diffuse into a cell and associate with tubulin. Affinities of maytansinoids for individual tubulin molecules are weak, but the high intracellular concentration of tubulin favors, after dissociation of a compound-tubulin complex, their re-binding to a tubulin molecule, or to a tip of a microtubule in the same cell, over their efflux. As a result, a significant fraction of microtubule tips is occupied with a maytansinoid when added to cells at sub-nanomolar concentrations, inducing mitotic arrest and cell death.

  7. Early Signs of Pathological Cognitive Aging in Mice Lacking High-Affinity Nicotinic Receptors.

    Science.gov (United States)

    Konsolaki, Eleni; Tsakanikas, Panagiotis; Polissidis, Alexia V; Stamatakis, Antonios; Skaliora, Irini

    2016-01-01

    In order to address pathological cognitive decline effectively, it is critical to adopt early preventive measures in individuals considered at risk. It is therefore essential to develop approaches that identify such individuals before the onset of irreversible dementia. A deficient cholinergic system has been consistently implicated as one of the main factors associated with a heightened vulnerability to the aging process. In the present study we used mice lacking high affinity nicotinic receptors (β2-/-), which have been proposed as an animal model of accelerated/premature cognitive aging. Our aim was to identify behavioral signs that could serve as indicators or predictors of impending cognitive decline. We used test batteries in order to assess cognitive functions and additional tasks to investigate spontaneous behaviors, such as species-specific activities and exploration/locomotion in a novel environment. Our data confirm the hypothesis that β2-/- animals exhibit age-related cognitive impairments in spatial learning. In addition, they document age-related deficits in other areas, such as recognition memory, burrowing and nesting building, thereby extending the validity of this animal model for the study of pathological aging. Finally, our data reveal deficits in spontaneous behavior and habituation processes that precede the onset of cognitive decline and could therefore be useful as a non-invasive behavioral screen for identifying animals at risk. To our knowledge, this is the first study to perform an extensive behavioral assessment of an animal model of premature cognitive aging, and our results suggest that β2-nAChR dependent cognitive deterioration progressively evolves from initial subtle behavioral changes to global dementia due to the combined effect of the neuropathology and aging.

  8. Devices and approaches for generating specific high-affinity nucleic acid aptamers

    Science.gov (United States)

    Szeto, Kylan; Craighead, Harold G.

    2014-09-01

    High-affinity and highly specific antibody proteins have played a critical role in biological imaging, medical diagnostics, and therapeutics. Recently, a new class of molecules called aptamers has emerged as an alternative to antibodies. Aptamers are short nucleic acid molecules that can be generated and synthesized in vitro to bind to virtually any target in a wide range of environments. They are, in principal, less expensive and more reproducible than antibodies, and their versatility creates possibilities for new technologies. Aptamers are generated using libraries of nucleic acid molecules with random sequences that are subjected to affinity selections for binding to specific target molecules. This is commonly done through a process called Systematic Evolution of Ligands by EXponential enrichment, in which target-bound nucleic acids are isolated from the pool, amplified to high copy numbers, and then reselected against the desired target. This iterative process is continued until the highest affinity nucleic acid sequences dominate the enriched pool. Traditional selections require a dozen or more laborious cycles to isolate strongly binding aptamers, which can take months to complete and consume large quantities of reagents. However, new devices and insights from engineering and the physical sciences have contributed to a reduction in the time and effort needed to generate aptamers. As the demand for these new molecules increases, more efficient and sensitive selection technologies will be needed. These new technologies will need to use smaller samples, exploit a wider range of chemistries and techniques for manipulating binding, and integrate and automate the selection steps. Here, we review new methods and technologies that are being developed towards this goal, and we discuss their roles in accelerating the availability of novel aptamers.

  9. Molecular basis for the high-affinity binding and stabilization of firefly luciferase by PTC124

    Energy Technology Data Exchange (ETDEWEB)

    Auld, Douglas S.; Lovell, Scott; Thorne, Natasha; Lea, Wendy A.; Maloney, David J.; Shen, Min; Rai, Ganesha; Battaile, Kevin P.; Thomas, Craig J.; Simeonov, Anton; Hanzlik, Robert P.; Inglese, James (NIH); (Kansas); (HWMRI)

    2010-04-07

    Firefly luciferase (FLuc), an ATP-dependent bioluminescent reporter enzyme, is broadly used in chemical biology and drug discovery assays. PTC124 Ataluren; (3-[5-(2-fluorophenyl)-1,2,4-oxadiazol-3-yl]benzoic acid) discovered in an FLuc-based assay targeting nonsense codon suppression, is an unusually potent FLuc-inhibitor. Paradoxically, PTC124 and related analogs increase cellular FLuc activity levels by posttranslational stabilization. In this study, we show that FLuc inhibition and stabilization is the result of an inhibitory product formed during the FLuc-catalyzed reaction between its natural substrate, ATP, and PTC124. A 2.0 {angstrom} cocrystal structure revealed the inhibitor to be the acyl-AMP mixed-anhydride adduct PTC124-AMP, which was subsequently synthesized and shown to be a high-affinity multisubstrate adduct inhibitor (MAI; KD = 120 pM) of FLuc. Biochemical assays, liquid chromatography/mass spectrometry, and near-attack conformer modeling demonstrate that formation of this novel MAI is absolutely dependent upon the precise positioning and reactivity of a key meta-carboxylate of PTC124 within the FLuc active site. We also demonstrate that the inhibitory activity of PTC124-AMP is relieved by free coenzyme A, a component present at high concentrations in luciferase detection reagents used for cell-based assays. This explains why PTC124 can appear to increase, instead of inhibit, FLuc activity in cell-based reporter gene assays. To our knowledge, this is an unusual example in which the 'off-target' effect of a small molecule is mediated by an MAI mechanism.

  10. Targeting protein-protein interactions with trimeric ligands: high affinity inhibitors of the MAGUK protein family.

    Directory of Open Access Journals (Sweden)

    Klaus B Nissen

    Full Text Available PDZ domains in general, and those of PSD-95 in particular, are emerging as promising drug targets for diseases such as ischemic stroke. We have previously shown that dimeric ligands that simultaneously target PDZ1 and PDZ2 of PSD-95 are highly potent inhibitors of PSD-95. However, PSD-95 and the related MAGUK proteins contain three consecutive PDZ domains, hence we envisioned that targeting all three PDZ domains simultaneously would lead to more potent and potentially more specific interactions with the MAGUK proteins. Here we describe the design, synthesis and characterization of a series of trimeric ligands targeting all three PDZ domains of PSD-95 and the related MAGUK proteins, PSD-93, SAP-97 and SAP-102. Using our dimeric ligands targeting the PDZ1-2 tandem as starting point, we designed novel trimeric ligands by introducing a PDZ3-binding peptide moiety via a cysteine-derivatized NPEG linker. The trimeric ligands generally displayed increased affinities compared to the dimeric ligands in fluorescence polarization binding experiments and optimized trimeric ligands showed low nanomolar inhibition towards the four MAGUK proteins, thus being the most potent inhibitors described. Kinetic experiments using stopped-flow spectrometry showed that the increase in affinity is caused by a decrease in the dissociation rate of the trimeric ligand as compared to the dimeric ligands, likely reflecting the lower probability of simultaneous dissociation of all three PDZ ligands. Thus, we have provided novel inhibitors of the MAGUK proteins with exceptionally high affinity, which can be used to further elucidate the therapeutic potential of these proteins.

  11. In vitro selection, characterization, and biosensing application of high-affinity cylindrospermopsin-targeting aptamers.

    Science.gov (United States)

    Elshafey, Reda; Siaj, Mohamed; Zourob, Mohammed

    2014-09-16

    Contamination of freshwater with cyanotoxin cylindrospermopsin (CYN) represents a significant global concern for public health. The sensitive detection of CYN