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Sample records for acidic protein gfap

  1. Expression patterns of glial fibrillary acidic protein (GFAP)-delta in epilepsy-associated lesional pathologies

    NARCIS (Netherlands)

    L. Martinian; K. Boer; J. Middeldorp; E.M. Hol; S.M. Sisodiya; W. Squier; E.M.A. Aronica; M. Thom

    2009-01-01

    Aims: Glial fibrillary acidic protein (GFAP)-delta is a novel isoform that differs in its C-terminal sequence from other GFAP isoforms. Previous studies suggest restriction of expression to the subpial layer, subventricular zone and the subgranular zone astrocytes, with an absence in pathological co

  2. Regulatory mechanisms for 3'-end alternative splicing and polyadenylation of the Glial Fibrillary Acidic Protein, GFAP, transcript

    DEFF Research Database (Denmark)

    Blechingberg, Jenny; Lykke-Andersen, Søren; Jensen, Torben Heick

    2007-01-01

    molecular mechanisms participating in alternative GFAP expression. Usage of a polyadenylation signal within the alternatively spliced exon 7a is essential to generate the GFAP kappa and GFAP kappa transcripts. The GFAP kappa mRNA is distinct from GFAP epsilon mRNA given that it also includes intron 7a......The glial fibrillary acidic protein, GFAP, forms the intermediate cytoskeleton in cells of the glial lineage. Besides the common GFAP alpha transcript, the GFAP epsilon and GFAP kappa transcripts are generated by alternative mRNA 3'-end processing. Here we use a GFAP minigene to characterize...... (PTB) protein enhanced both exon 7a polyadenylation and exon 7a splicing. Finally, increasing transcription by the VP16 trans-activator did not affect the frequency of use of the exon 7a polyadenylation signal whereas the exon 7a splicing frequency was decreased. Our data suggest a model...

  3. Blood levels of glial fibrillary acidic protein (GFAP in patients with neurological diseases.

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    Christoph A Mayer

    Full Text Available BACKGROUND AND PURPOSE: The brain-specific astroglial protein GFAP is a blood biomarker candidate indicative of intracerebral hemorrhage in patients with symptoms suspicious of acute stroke. Comparably little, however, is known about GFAP release in other neurological disorders. In order to identify potential "specificity gaps" of a future GFAP test used to diagnose intracerebral hemorrhage, we measured GFAP in the blood of a large and rather unselected collective of patients with neurological diseases. METHODS: Within a one-year period, we randomly selected in-patients of our university hospital for study inclusion. Patients with ischemic stroke, transient ischemic attack and intracerebral hemorrhage were excluded. Primary endpoint was the ICD-10 coded diagnosis reached at discharge. During hospital stay, blood was collected, and GFAP plasma levels were determined using an advanced prototype immunoassay at Roche Diagnostics. RESULTS: A total of 331 patients were included, covering a broad spectrum of neurological diseases. GFAP levels were low in the vast majority of patients, with 98.5% of cases lying below the cut-off that was previously defined for the differentiation of intracerebral hemorrhage and ischemic stroke. No diagnosis or group of diagnoses was identified that showed consistently increased GFAP values. No association with age and sex was found. CONCLUSION: Most acute and chronic neurological diseases, including typical stroke mimics, are not associated with detectable GFAP levels in the bloodstream. Our findings underline the hypothesis that rapid astroglial destruction as in acute intracerebral hemorrhage is mandatory for GFAP increase. A future GFAP blood test applied to identify patients with intracerebral hemorrhage is likely to have a high specificity.

  4. Glial Fibrillary Acidic Protein Filaments Can Tolerate the Incorporation of Assembly-compromised GFAP-δ, but with Consequences for Filament Organization and αB-Crystallin Association

    Science.gov (United States)

    Perng, Ming-Der; Wen, Shu-Fang; Gibbon, Terry; Middeldorp, Jinte; Sluijs, Jacqueline; Hol, Elly M.

    2008-01-01

    The glial fibrillary acidic protein (GFAP) gene is alternatively spliced to give GFAP-α, the most abundant isoform, and seven other differentially expressed transcripts including GFAP-δ. GFAP-δ has an altered C-terminal domain that renders it incapable of self-assembly in vitro. When titrated with GFAP-α, assembly was restored providing GFAP-δ levels were kept low (∼10%). In a range of immortalized and transformed astrocyte derived cell lines and human spinal cord, we show that GFAP-δ is naturally part of the endogenous intermediate filaments, although levels were low (∼10%). This suggests that GFAP filaments can naturally accommodate a small proportion of assembly-compromised partners. Indeed, two other assembly-compromised GFAP constructs, namely enhanced green fluorescent protein (eGFP)-tagged GFAP and the Alexander disease–causing GFAP mutant, R416W GFAP both showed similar in vitro assembly characteristics to GFAP-δ and could also be incorporated into endogenous filament networks in transfected cells, providing expression levels were kept low. Another common feature was the increased association of αB-crystallin with the intermediate filament fraction of transfected cells. These studies suggest that the major physiological role of the assembly-compromised GFAP-δ splice variant is as a modulator of the GFAP filament surface, effecting changes in both protein– and filament–filament associations as well as Jnk phosphorylation. PMID:18685083

  5. Glial fibrillary acidic protein (GFAP) and the astrocyte intermediate filament system in diseases of the central nervous system.

    Science.gov (United States)

    Hol, Elly M; Pekny, Milos

    2015-02-01

    Glial fibrillary acidic protein (GFAP) is the hallmark intermediate filament (IF; also known as nanofilament) protein in astrocytes, a main type of glial cells in the central nervous system (CNS). Astrocytes have a range of control and homeostatic functions in health and disease. Astrocytes assume a reactive phenotype in acute CNS trauma, ischemia, and in neurodegenerative diseases. This coincides with an upregulation and rearrangement of the IFs, which form a highly complex system composed of GFAP (10 isoforms), vimentin, synemin, and nestin. We begin to unravel the function of the IF system of astrocytes and in this review we discuss its role as an important crisis-command center coordinating cell responses in situations connected to cellular stress, which is a central component of many neurological diseases. Copyright © 2015 Elsevier Ltd. All rights reserved.

  6. Alternative mRNA Splicing from the Glial Fibrillary Acidic Protein (GFAP) Gene Generates Isoforms with Distinct Subcellular mRNA Localization Patterns in Astrocytes

    DEFF Research Database (Denmark)

    Thomsen, Rune; Daugaard, Tina Fuglsang; Holm, Ida E;

    2013-01-01

    The intermediate filament network of astrocytes includes Glial fibrillary acidic protein (Gfap) as a major component. Gfap mRNA is alternatively spliced resulting in generation of different protein isoforms where Gfapa is the most predominant isoform. The Gfapd isoform is expressed in proliferating......RNA localization patterns were dependent on the different 39-exon sequences included in Gfapd and Gfapa mRNA. The presented results show that alternative Gfap mRNA splicing results in isoform-specific mRNA localization patterns with resulting different local mRNA concentration ratios which have potential...

  7. Measurement of the glial fibrillary acidic protein and its breakdown products GFAP-BDP biomarker for the detection of traumatic brain injury compared to computed tomography and magnetic resonance imaging.

    Science.gov (United States)

    McMahon, Paul J; Panczykowski, David M; Yue, John K; Puccio, Ava M; Inoue, Tomoo; Sorani, Marco D; Lingsma, Hester F; Maas, Andrew I R; Valadka, Alex B; Yuh, Esther L; Mukherjee, Pratik; Manley, Geoffrey T; Okonkwo, David O

    2015-04-15

    Glial fibrillary acidic protein and its breakdown products (GFAP-BDP) are brain-specific proteins released into serum as part of the pathophysiological response after traumatic brain injury (TBI). We performed a multi-center trial to validate and characterize the use of GFAP-BDP levels in the diagnosis of intracranial injury in a broad population of patients with a positive clinical screen for head injury. This multi-center, prospective, cohort study included patients 16-93 years of age presenting to three level 1 trauma centers with suspected TBI (loss of consciousness, post-trauma amnesia, and so on). Serum GFAP-BDP levels were drawn within 24 h and analyzed, in a blinded fashion, using sandwich enzyme-linked immunosorbent assay. The ability of GFAP-BDP to predict intracranial injury on admission computed tomography (CT) as well as delayed magnetic resonance imaging was analyzed by multiple regression and assessed by the area under the receiver operating characteristic curve (AUC). Utility of GFAP-BDP to predict injury and reduce unnecessary CT scans was assessed utilizing decision curve analysis. A total of 215 patients were included, of which 83% suffered mild TBI, 4% moderate, and 12% severe; mean age was 42.1±18 years. Evidence of intracranial injury was present in 51% of the sample (median Rotterdam Score, 2; interquartile range, 2). GFAP-BDP demonstrated very good predictive ability (AUC=0.87) and demonstrated significant discrimination of injury severity (odds ratio, 1.45; 95% confidence interval, 1.29-1.64). Use of GFAP-BDP yielded a net benefit above clinical screening alone and a net reduction in unnecessary scans by 12-30%. Used in conjunction with other clinical information, rapid measurement of GFAP-BDP is useful in establishing or excluding the diagnosis of radiographically apparent intracranial injury throughout the spectrum of TBI. As an adjunct to current screening practices, GFAP-BDP may help avoid unnecessary CT scans without sacrificing

  8. 应用CATS法分离和鉴定猪GFAP基因的研究%Isolation and Characterization of the Porcine Glial Fibrillary Acidic Protein ( GFAP) Gene by CATS

    Institute of Scientific and Technical Information of China (English)

    余梅; 李奎; 赵书红; 刘榜; 熊统安

    2001-01-01

    Primers for the glial fibrillary acidic protein (GFAP) gene weredesigned from a human cDNA sequence aligned with the mouse GFAP gene on the principle of comparative anchor tagged sequence (CATS). The 412bp PCR product isolated from Chinese Erhualian pig genome was characterized as the porcine GFAP gene by comparing the sequence with the GenBank database. The chromosomal location of the GFAP gene is on pig Chr: 12(p11-(2/3)p13) using pig×rodent somatic cell hybrid panel.%根据比较锚定序列示踪(CATS)法,选择人和小鼠胶质细胞原纤维酸性蛋白(GFAP)基因的同源区域设计引物,用PCR方法从二花脸猪基因组中分离到412bp的基因片段,经与基因资料库中已知功能基因的同源性比较,该片段可鉴定为猪的GFAP基因,利用猪-啮齿类体细胞杂种克隆板将CFAP基因定位于猪12号染色体12p11-(2/3)p13区域。

  9. Astrócitos imunorreativos à proteína glial fibrilar ácida (GFAP em sistema nervoso central de equinos normais e de equinos com leucoencefalomalácia Glial fibrillary acidic protein (GFAP immunoreactive astrocytes in the Central Nervous System of normal horses and horses with leukoencephalomalacia

    Directory of Open Access Journals (Sweden)

    Karen Regina Lemos

    1999-07-01

    Full Text Available A proteína glial fibrilar ácida (GFAP, subunidade dos filamentos intermediários do citoesqueleto celular, está presente no citoplasma de astrócitos. Técnicas imunohistoquímicas com anticorpos primários anti-GFAP são geralmente empregadas para identificar astrócitos no sistema nervoso, permitindo verificar também sua hipertrofia. Vários estudos mostram a distribuição, a morfologia e a citoarquitetura de astrócitos em várias regiões do SNC do homem e de animais de laboratório. No entanto, em animais domésticos e, especialmente em equinos, poucas informações estão disponíveis. No presente trabalho, verificou-se a densidade e a morfologia de astrócitos imunorreativos à GFAP na substância branca da córtex cerebral de equinos com leucoencefalomalácia (LEM comparando-se esses aspectos com o de equinos normais. Animais com LEM apresentaram hipertrofia de astrócitos em áreas próximas às lesões, representada pelo aumento do corpo celular, do núcleo e dos prolongamentos citoplasmáticos. O número de astrócitos apresentou-se reduzido e a imunorreatividade foi mais acentuada. Nos animais normais, verificou-se distribuição constante de astrócitos imunorreagentes com características de fibrosos. Alterações vasculares nos animais com LEM, como por exemplo degeneração de endotélio vascular, também foram observadas, podendo estar associadas às alterações astrocíticas.The glial fibrillary acidic protein (GFAP, subunit of the intermediary filaments of the cellular cytoskeleton, exists in the cytoplasm of astrocytes. Immunohistochemistry utilizing primary antibodies anti-GFAP is generally chosen to identify astrocytes in the central nervous system (CNS, allowing also to verify their hypertrophy. Several studies show the distribution, morphology and cytoarchitecture of the astrocytes in several areas of the CNS of humans and laboratory animals. However, in domestic animals, especially in horses, little information is

  10. Oxytocin Rapidly Changes Astrocytic GFAP Plasticity by Differentially Modulating the Expressions of pERK 1/2 and Protein Kinase A

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    Ping Wang

    2017-08-01

    Full Text Available The importance of astrocytes to normal brain functions and neurological diseases has been extensively recognized; however, cellular mechanisms underlying functional and structural plasticities of astrocytes remain poorly understood. Oxytocin (OT is a neuropeptide that can rapidly change astrocytic plasticity in association with lactation, as indicated in the expression of glial fibrillary acidic protein (GFAP in the supraoptic nucleus (SON. Here, we used OT-evoked changes in GFAP expression in astrocytes of male rat SON as a model to explore the cellular mechanisms underlying GFAP plasticity. The results showed that OT significantly reduced the expression of GFAP filaments and proteins in SON astrocytes in brain slices. In lysates of the SON, OT receptors (OTRs were co-immunoprecipitated with GFAP; vasopressin (VP, a neuropeptide structurally similar to OT, did not significantly change GFAP protein level; OT-evoked depolarization of astrocyte membrane potential was sensitive to a selective OTR antagonist (OTRA but not to tetanus toxin, a blocker of synaptic transmission. The effects of OT on GFAP expression and on astrocyte uptake of Bauer-Peptide, an astrocyte-specific dye, were mimicked by isoproterenol (IPT; β-adrenoceptor agonist, U0126 or PD98059, inhibitors of extracellular signal-regulated protein kinase (ERK 1/2 kinase and blocked by the OTRA or KT5720, a protein kinase A (PKA inhibitor. The effect of OT on GFAP expressions and its association with these kinases were simulated by mSIRK, an activator of Gβγ subunits. Finally, suckling increased astrocytic expression of the catalytic subunit of PKA (cPKA at astrocytic processes while increasing the molecular associations of GFAP with cPKA and phosphorylated ERK (pERK 1/2. Upon the occurrence of the milk-ejection reflex, spatial co-localization of the cPKA with GFAP filaments further increased, which was accompanied with increased molecular association of GFAP with pERK 1/2 but not with

  11. Histone acetylation in astrocytes suppresses GFAP and stimulates a reorganization of the intermediate filament network

    NARCIS (Netherlands)

    Kanski, Regina; Sneeboer, Marjolein A M; van Bodegraven, Emma J; Sluijs, Jacqueline A; Kropff, Wietske; Vermunt, Marit W.; Creyghton, Menno P; De Filippis, Lidia; Vescovi, Angelo; Aronica, Eleonora; van Tijn, P.; van Strien, Miriam E; Hol, Elly M

    2014-01-01

    Glial fibrillary acidic protein (GFAP) is the main intermediate filament in astrocytes and is regulated by epigenetic mechanisms during development. We demonstrate that histone acetylation also controls GFAP expression in mature astrocytes. Inhibition of histone deacetylases (HDACs) with trichostati

  12. Silencing GFAP isoforms in astrocytoma cells disturbs laminin-dependent motility and cell adhesion

    NARCIS (Netherlands)

    Moeton, Martina; Kanski, Regina; Stassen, Oscar M J A; Sluijs, Jacqueline A; Geerts, Dirk; van Tijn, P.; Wiche, Gerhard; van Strien, Miriam E; Hol, Elly M

    2014-01-01

    Glial fibrillary acidic protein (GFAP) is an intermediate filament protein expressed in astrocytes and neural stem cells. The GFAP gene is alternatively spliced, and expression of GFAP is highly regulated during development, on brain damage, and in neurodegenerative diseases. GFAPα is the canonical

  13. Enteric GFAP expression and phosphorylation in Parkinson's disease

    NARCIS (Netherlands)

    Clairembault, Thomas; Kamphuis, W.; Leclair-Visonneau, Laurène; Rolli-Derkinderen, Malvyne; Coron, Emmanuel; Neunlist, Michel; Hol, Elly M; Derkinderen, Pascal

    2014-01-01

    Enteric glial cells (EGCs) are in many respects similar to astrocytes of the central nervous system and express similar proteins including glial fibrillary acidic protein (GFAP). Changes in GFAP expression and/or phosphorylation have been reported during brain damage or central nervous system degene

  14. LONGMU BUPLEURUM DECOCTION ON STEREOTYPED BEHAVIOR AND EXPRESSION OF GLIAL FIBRILLARY ACIDIC PROTEIN(GFAP)IN RAT MODEL OF SCHIZOPHRENIA%柴胡龙牡汤对精神分裂症大鼠模型刻板行为和胶质细胞纤维酸性蛋白(GFAP)表达的影响

    Institute of Scientific and Technical Information of China (English)

    李东阳; 周秀芳; 孙晓东; 陈涛; 鲁娜

    2011-01-01

    [目的]观察柴胡龙牡汤对精神分裂症大鼠模型刻板行为和GFAP表达的影响.[方法]将44只大鼠分为4组,空白组腹腔注射生理盐水.剩余3组均用氯胺酮造模(腹腔注射氯胺),其中氯氮平组从d4开始加用氮氮平灌胃,柴胡龙牡汤组从d4开始加用柴胡龙牡汤灌胃.[结果]模型组刻板行为较空白组明显增多;氯氮平组和柴胡龙牡汤组刻板行为均较模型组明显减少;柴胡龙牡汤组刻板行为较氯氮平组增多.模型组GFAP表达较空白组明显增多;氯氮平组GFAP表达较模型组明显减少;柴胡龙牡汤组GFAP表达较模型组明显减少;柴胡龙牡汤组GFAP表达较氯氮平组增多.[结论]柴胡龙牡汤对精神分裂症大鼠模型前额叶GFAP和刻板行为有一定的影响.%[Objective] To observe Longmu Bupleurum Decoction on stereotyped behavior and CFAP expression in rat model of schizophrenia. [Methods] 44 rats were divided into 4 groups, control group injected with normal saline.The remaining three groups were made using ketamine model (intraperitoneal injection of ketamine), in which clozapine group was added intragastric administration of clozapine from the fourth day, Chai Hu Tang Longmu d starting was added intragastric administration of Bupleurum Longmu from the fourth day. [Results] The stereotyped behavior in model group increased significantly comparing with control group; the stereotyped behaviors in clozapine group and Bupleurum Decoction group Longmu were significantly reduced comparing with model group; the stereotypic behavior in Bupleurum Longmu Decoction increased more comparing with that in clozapine group.GFAP expression in model group increased significantly comparing with control group; CFAP expression in clozapine group was significantly reduced comparingwith the model; CFAP expression in Bupleurum Decoction Longmu was significantly reduced comparing with the model; CFAP expression in Bupleurum Longmu Decoction increased more

  15. Glial fibrillary acidic protein isoform expression in plaque related astrogliosis in Alzheimer's disease

    NARCIS (Netherlands)

    Kamphuis, W.; Middeldorp, Jinte; Kooijman, Lieneke; Sluijs, Jacqueline A; Kooi, Evert-Jan; Moeton, Martina; Freriks, Michel; Mizee, Mark R; Hol, Elly M

    2014-01-01

    In Alzheimer's disease (AD), amyloid plaques are surrounded by reactive astrocytes with an increased expression of intermediate filaments including glial fibrillary acidic protein (GFAP). Different GFAP isoforms have been identified that are differentially expressed by specific subpopulations of ast

  16. GFAP expression as an indicator of disease severity in mouse models of Alexander disease

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    Albee Messing

    2013-03-01

    Full Text Available AxD (Alexander disease is a rare disorder caused by heterozygous mutations in GFAP (glial fibrillary acidic protein resulting in accumulation of the GFAP protein and elevation of Gfap mRNA. To test whether GFAP itself can serve as a biomarker of disease status or progression, we investigated two independent measures of GFAP expression in AxD mouse models, one using a genetic reporter of promoter activity and the other quantifying GFAP protein directly in a manner that could also be employed in human studies. Using a transgenic reporter line that expresses firefly luciferase under the control of the murine Gfap promoter (Gfap-luc, we found that luciferase activity reflected the regional CNS (central nervous system variability of Gfap mRNA in Gfap+/+ mice, and increased in mice containing a point mutation in Gfap that mimics a common human mutation in AxD (R239H in the human sequence, and R236H in the murine sequence. In a second set of studies, we quantified GFAP protein in CSF (cerebrospinal fluid taken from three different AxD mouse models and littermate controls. GFAP levels in CSF were increased in all three AxD models, in a manner corresponding to the concentrations of GFAP in brain. These studies demonstrate that transactivation of the Gfap promoter is an early and sustained indicator of the disease process in the mouse. Furthermore, GFAP in CSF serves as a potential biomarker that is comparable between mouse models and human patients.

  17. Serum GFAP levels in optic neuropathies.

    NARCIS (Netherlands)

    Storoni, M.; Verbeek, M.M.; Illes, Z.; Marignier, R.; Teunissen, C.E.; Grabowska, M.; Confavreux, C.; Plant, G.T.; Petzold, A.

    2012-01-01

    BACKGROUND: Complement mediated autoimmunity against aquaporin-4 results in astrocytic damage in neuromyelitis optica (NMO). There is evidence for increased CSF glial fibrillary acidic protein (GFAP) and S100B levels in acute NMO. Here we tested whether the CSF finding also holds true for the diagno

  18. Screening for GFAP rearrangements in a cohort of Alexander disease and undetermined leukoencephalopathy patients.

    Science.gov (United States)

    Ferreira, Marie-Céleste; Dorboz, Imen; Rodriguez, Diana; Boespflug Tanguy, Odile

    2015-09-01

    Alexander disease (AxD), a fatal degenerative leukoencephalopathy, is caused by de novo heterozygous missense mutations in the Glial Fibrillary Acidic Protein (GFAP) gene. The pathological hallmark of the disease is the presence of Rosenthal fibers, cytoplasmic inclusions in astrocytes, composed mainly of GFAP, αB-crystallin and HSP27. To date, several patients with a typical presentation of the disease or displaying characteristic Rosenthal fibers in brain material have been reported with no GFAP mutation. Recently, several studies have demonstrated a correlation between Rosenthal fiber formation and wild-type GFAP overexpression, despite the absence of mutations. We tested the hypothesis that a GFAP gene rearrangement could modulate AxD severity or promote GFAP overexpression and aggregation, resulting in leukoencephalopathy. A QMPSF assay was validated for 11 exonic fragments: 3 in control genes (CFTR, DSCR1, F9) and 8 corresponding to GFAP exons. A total of 97 patients suspected of AxD were analyzed: 28 with a GFAP mutation; 69 with clinical and magnetic resonance imaging criteria compatible with the disease. Neither duplications nor deletions of GFAP were found, suggesting that GFAP coding-region rearrangements may not be involved in AxD or Alexander-related leukoencephalopathies. In addition, 80 patients with undetermined leukodystrophies, and negative for PLP1, GJA12, Sox10 and MCT8 mutations and PLP1 and Lamin B1 rearrangements, were tested. These patients were also negative for GFAP rearrangements. Other hypotheses should be investigated for a molecular diagnosis in patients with undetermined leukoencephalopathy: mutations in GFAP isoforms, splicing sites or regulatory regions, or defaults in genes encoding molecular partners of GFAP. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

  19. Silencing GFAP isoforms in astrocytoma cells disturbs laminin-dependent motility and cell adhesion.

    Science.gov (United States)

    Moeton, Martina; Kanski, Regina; Stassen, Oscar M J A; Sluijs, Jacqueline A; Geerts, Dirk; van Tijn, Paula; Wiche, Gerhard; van Strien, Miriam E; Hol, Elly M

    2014-07-01

    Glial fibrillary acidic protein (GFAP) is an intermediate filament protein expressed in astrocytes and neural stem cells. The GFAP gene is alternatively spliced, and expression of GFAP is highly regulated during development, on brain damage, and in neurodegenerative diseases. GFAPα is the canonical splice variant and is expressed in all GFAP-positive cells. In the human brain, the alternatively spliced transcript GFAPδ marks specialized astrocyte populations, such as subpial astrocytes and the neurogenic astrocytes in the human subventricular zone. We here show that shifting the GFAP isoform ratio in favor of GFAPδ in astrocytoma cells, by selectively silencing the canonical isoform GFAPα with short hairpin RNAs, induced a change in integrins, a decrease in plectin, and an increase in expression of the extracellular matrix component laminin. Together, this did not affect cell proliferation but resulted in a significantly decreased motility of astrocytoma cells. In contrast, a down-regulation of all GFAP isoforms led to less cell spreading, increased integrin expression, and a >100-fold difference in the adhesion of astrocytoma cells to laminin. In summary, isoform-specific silencing of GFAP revealed distinct roles of a specialized GFAP network in regulating the interaction of astrocytoma cells with the extracellular matrix through laminin.-Moeton, M., Kanski, R., Stassen, O. M. J. A., Sluijs, J. A., Geerts, D., van Tijn, P., Wiche, G., van Strien, M. E., Hol, E. M. Silencing GFAP isoforms in astrocytoma cells disturbs laminin dependent motility and cell adhesion.

  20. From stem cell to astrocyte: Decoding the regulation of GFAP

    NARCIS (Netherlands)

    R. Kanski

    2014-01-01

    The research presented in this thesis focuses on glial fibrillary acidic protein (GFAP), the main intermediate filament (IF) in astrocytes and astrocyte subpopulations such as neural stem cells (NSCs). In neurodegenerative diseases or upon brain damage, astrocytes respond to an injury with an upregu

  1. How Relevant Are GFAP Autoantibodies in Autism and Tourette Syndrome?

    Science.gov (United States)

    Kirkman, Nikki J.; Libbey, Jane E.; Sweeten, Thayne L.; Coon, Hilary H.; Miller, Judith N.; Stevenson, Edward K.; Lainhart, Janet E.; McMahon, William M.; Fujinami, Robert S.

    2008-01-01

    Controversy exists over the role of autoantibodies to central nervous system antigens in autism and Tourette Syndrome. We investigated plasma autoantibody titers to glial fibrillary acidic protein (GFAP) in children with classic onset (33) and regressive onset (26) autism, controls (25, healthy age- and gender-matched) and individuals with…

  2. GFAP isoforms in adult mouse brain with a focus on neurogenic astrocytes and reactive astrogliosis in mouse models of Alzheimer disease.

    Directory of Open Access Journals (Sweden)

    Willem Kamphuis

    Full Text Available Glial fibrillary acidic protein (GFAP is the main astrocytic intermediate filament (IF. GFAP splice isoforms show differential expression patterns in the human brain. GFAPδ is preferentially expressed by neurogenic astrocytes in the subventricular zone (SVZ, whereas GFAP(+1 is found in a subset of astrocytes throughout the brain. In addition, the expression of these isoforms in human brain material of epilepsy, Alzheimer and glioma patients has been reported. Here, for the first time, we present a comprehensive study of GFAP isoform expression in both wild-type and Alzheimer Disease (AD mouse models. In cortex, cerebellum, and striatum of wild-type mice, transcripts for Gfap-α, Gfap-β, Gfap-γ, Gfap-δ, Gfap-κ, and a newly identified isoform Gfap-ζ, were detected. Their relative expression levels were similar in all regions studied. GFAPα showed a widespread expression whilst GFAPδ distribution was prominent in the SVZ, rostral migratory stream (RMS, neurogenic astrocytes of the subgranular zone (SGZ, and subpial astrocytes. In contrast to the human SVZ, we could not establish an unambiguous GFAPδ localization in proliferating cells of the mouse SVZ. In APPswePS1dE9 and 3xTgAD mice, plaque-associated reactive astrocytes had increased transcript levels of all detectable GFAP isoforms and low levels of a new GFAP isoform, Gfap-ΔEx7. Reactive astrocytes in AD mice showed enhanced GFAPα and GFAPδ immunolabeling, less frequently increased vimentin and nestin, but no GFAPκ or GFAP(+1 staining. In conclusion, GFAPδ protein is present in SVZ, RMS, and neurogenic astrocytes of the SGZ, but also outside neurogenic niches. Furthermore, differential GFAP isoform expression is not linked with aging or reactive gliosis. This evidence points to the conclusion that differential regulation of GFAP isoforms is not involved in the reorganization of the IF network in reactive gliosis or in neurogenesis in the mouse brain.

  3. Lithium Decreases Glial Fibrillary Acidic Protein in a Mouse Model of Alexander Disease.

    Science.gov (United States)

    LaPash Daniels, Christine M; Paffenroth, Elizabeth; Austin, Elizabeth V; Glebov, Konstantin; Lewis, Diana; Walter, Jochen; Messing, Albee

    2015-01-01

    Alexander disease is a fatal neurodegenerative disease caused by mutations in the astrocyte intermediate filament glial fibrillary acidic protein (GFAP). The disease is characterized by elevated levels of GFAP and the formation of protein aggregates, known as Rosenthal fibers, within astrocytes. Lithium has previously been shown to decrease protein aggregates by increasing the autophagy pathway for protein degradation. In addition, lithium has also been reported to decrease activation of the transcription factor STAT3, which is a regulator of GFAP transcription and astrogliogenesis. Here we tested whether lithium treatment would decrease levels of GFAP in a mouse model of Alexander disease. Mice with the Gfap-R236H point mutation were fed lithium food pellets for 4 to 8 weeks. Four weeks of treatment with LiCl at 0.5% in food pellets decreased GFAP protein and transcripts in several brain regions, although with mild side effects and some mortality. Extending the duration of treatment to 8 weeks resulted in higher mortality, and again with a decrease in GFAP in the surviving animals. Indicators of autophagy, such as LC3, were not increased, suggesting that lithium may decrease levels of GFAP through other pathways. Lithium reduced the levels of phosphorylated STAT3, suggesting this as one pathway mediating the effects on GFAP. In conclusion, lithium has the potential to decrease GFAP levels in Alexander disease, but with a narrow therapeutic window separating efficacy and toxicity.

  4. Evaluating the potential of the GFAP-KLH immune-tolerizing vaccine for type 1 diabetes in mice.

    Science.gov (United States)

    Pang, Zhengda; Higuchi, Masayoshi; Koriyama, Hiroshi; Yoshida, Shota; Kurinami, Hitomi; Shimamura, Munehisa; Takami, Yoichi; Rakugi, Hiromi; Morishita, Ryuichi; Nakagami, Hironori

    2017-01-01

    Glial fibrillary acidic protein (GFAP), expressed in peri-islet Schwann cells, is a novel target for the treatment of type 1 diabetes mellitus (T1DM). We designed a GFAP immune-tolerizing vaccine that successfully suppresses hyperglycemia and enhances C peptide secretion. The GFAP vaccine significantly prevented T cell infiltration into pancreatic islets. Moreover, after GFAP vaccination, naïve T-cell differentiation shifted from a cytotoxic Th1- to a Th2-biased humoral response. These results indicate that as a novel target, GFAP reliably predicts the development of T1DM, and that the GFAP vaccine successfully delays the progression of T1DM by regulating T-cell differentiation.

  5. Histone acetylation in astrocytes suppresses GFAP and stimulates a reorganization of the intermediate filament network.

    Science.gov (United States)

    Kanski, Regina; Sneeboer, Marjolein A M; van Bodegraven, Emma J; Sluijs, Jacqueline A; Kropff, Wietske; Vermunt, Marit W; Creyghton, Menno P; De Filippis, Lidia; Vescovi, Angelo; Aronica, Eleonora; van Tijn, Paula; van Strien, Miriam E; Hol, Elly M

    2014-10-15

    Glial fibrillary acidic protein (GFAP) is the main intermediate filament in astrocytes and is regulated by epigenetic mechanisms during development. We demonstrate that histone acetylation also controls GFAP expression in mature astrocytes. Inhibition of histone deacetylases (HDACs) with trichostatin A or sodium butyrate reduced GFAP expression in primary human astrocytes and astrocytoma cells. Because splicing occurs co-transcriptionally, we investigated whether histone acetylation changes the ratio between the canonical isoform GFAPα and the alternative GFAPδ splice variant. We observed that decreased transcription of GFAP enhanced alternative isoform expression, as HDAC inhibition increased the GFAPδ∶GFAPα ratio. Expression of GFAPδ was dependent on the presence and binding of splicing factors of the SR protein family. Inhibition of HDAC activity also resulted in aggregation of the GFAP network, reminiscent of our previous findings of a GFAPδ-induced network collapse. Taken together, our data demonstrate that HDAC inhibition results in changes in transcription, splicing and organization of GFAP. These data imply that a tight regulation of histone acetylation in astrocytes is essential, because dysregulation of gene expression causes the aggregation of GFAP, a hallmark of human diseases like Alexander's disease.

  6. Utilization of glial fibrillary acidic protein and galectin-3 in the ...

    African Journals Online (AJOL)

    Results: A total of 90 participants, forty patients with ischemic infarction who ... Conclusion: The levels of Galectin-3 and GFAP were increased in acute ischemic stroke patients. Keywords: Galectin-3, glial fibrillary acidic protein, ischemic stroke ...

  7. Time-lapse imaging reveals symmetric neurogenic cell division of GFAP-expressing progenitors for expansion of postnatal dentate granule neurons.

    Directory of Open Access Journals (Sweden)

    Takashi Namba

    Full Text Available Granule cells in the hippocampus, a region critical for memory and learning, are generated mainly during the early postnatal period but neurogenesis continues in adulthood. Postnatal neuronal production is carried out by primary progenitors that express glial fibrillary acidic protein (GFAP and they are assumed to function as stem cells. A central question regarding postnatal dentate neurogenesis is how astrocyte-like progenitors produce neurons. To reveal cell division patterns and the process of neuronal differentiation of astrocyte-like neural progenitors, we performed time-lapse imaging in cultured hippocampal slices from early postnatal transgenic mice with mouse GFAP promoter-controlled enhanced green fluorescent protein (mGFAP-eGFP Tg mice in combination with a retrovirus carrying a red fluorescent protein gene. Our results showed that the majority of GFAP-eGFP+ progenitor cells that express GFAP, Sox2 and nestin divided symmetrically to produce pairs of GFAP+ cells (45% or pairs of neuron-committed cells (45%, whereas a minority divided asymmetrically to generate GFAP+ cells and neuron-committed cells (10%. The present results suggest that a substantial number of GFAP-expressing progenitors functions as transient amplifying progenitors, at least in an early postnatal dentate gyrus, although a small population appears to be stem cell-like progenitors. From the present data, we discuss possible cell division patterns of adult GFAP+ progenitors.

  8. Time-lapse imaging reveals symmetric neurogenic cell division of GFAP-expressing progenitors for expansion of postnatal dentate granule neurons.

    Science.gov (United States)

    Namba, Takashi; Mochizuki, Hideki; Suzuki, Ryusuke; Onodera, Masafumi; Yamaguchi, Masahiro; Namiki, Hideo; Shioda, Seiji; Seki, Tatsunori

    2011-01-01

    Granule cells in the hippocampus, a region critical for memory and learning, are generated mainly during the early postnatal period but neurogenesis continues in adulthood. Postnatal neuronal production is carried out by primary progenitors that express glial fibrillary acidic protein (GFAP) and they are assumed to function as stem cells. A central question regarding postnatal dentate neurogenesis is how astrocyte-like progenitors produce neurons. To reveal cell division patterns and the process of neuronal differentiation of astrocyte-like neural progenitors, we performed time-lapse imaging in cultured hippocampal slices from early postnatal transgenic mice with mouse GFAP promoter-controlled enhanced green fluorescent protein (mGFAP-eGFP Tg mice) in combination with a retrovirus carrying a red fluorescent protein gene. Our results showed that the majority of GFAP-eGFP+ progenitor cells that express GFAP, Sox2 and nestin divided symmetrically to produce pairs of GFAP+ cells (45%) or pairs of neuron-committed cells (45%), whereas a minority divided asymmetrically to generate GFAP+ cells and neuron-committed cells (10%). The present results suggest that a substantial number of GFAP-expressing progenitors functions as transient amplifying progenitors, at least in an early postnatal dentate gyrus, although a small population appears to be stem cell-like progenitors. From the present data, we discuss possible cell division patterns of adult GFAP+ progenitors.

  9. Existence of vimentin and GFAP protein expressions as a result of 2-Methoxyethanol administration in cerebral cortex tissue of Swiss Webster mice (Mus musculus): an immunohistochemical analysis.

    Science.gov (United States)

    Irnidayanti, Yulia

    2014-07-01

    Une of the plastic-based materials widely used in the plastics industry in various countries is ester phthalate. This compound will be oxidized in the body into 2-methoxyethanol (2-ME). The effect of 2-ME on human health and environment depends on the number, duration and the frequency of exposure. Recently, the incidence of brain damage tends to increase. In the last decade, it has been widely reported the negative effects of chemical pollutants to the environment. The aim of this study were to know the existence of the expression of Vimentin and GFAP proteins caused by 2-ME on the histological structure of the cerebral cortex of mice fetal during the prenatal period on gestation day 14 (GD 14) and day 18 (GD 18). The 2-ME compound was injected intraperitoneally with a dose of 7.5 mmol kg(-1) of body weight at GD-10. The result showed that there is a change in existence Vimentin protein in the cerebral cortex fetal of treated mice at GD 14, which is very conspicuous. Meanwhile, a change in existence of GFAP protein in cerebral cortex fetal of treated mice at GD 14, have relatively no difference from controls and no impact on histological structure changes of the cerebral corteks at GD 14. The change in existence of Vimentin protein in the cerebral cortex fetal of treated mice at GD 14 have an impact on histological structure of the cerebral cortex of mice treated at GD 18. It is believed that the impact is due to the effects of 2-methoxyethanol.

  10. Glial fibrillary acidic protein isoform expression in plaque related astrogliosis in Alzheimer's disease.

    Science.gov (United States)

    Kamphuis, Willem; Middeldorp, Jinte; Kooijman, Lieneke; Sluijs, Jacqueline A; Kooi, Evert-Jan; Moeton, Martina; Freriks, Michel; Mizee, Mark R; Hol, Elly M

    2014-03-01

    In Alzheimer's disease (AD), amyloid plaques are surrounded by reactive astrocytes with an increased expression of intermediate filaments including glial fibrillary acidic protein (GFAP). Different GFAP isoforms have been identified that are differentially expressed by specific subpopulations of astrocytes and that impose different properties to the intermediate filament network. We studied transcript levels and protein expression patterns of all known GFAP isoforms in human hippocampal AD tissue at different stages of the disease. Ten different transcripts for GFAP isoforms were detected at different abundancies. Transcript levels of most isoforms increased with AD progression. GFAPδ-immunopositive astrocytes were observed in subgranular zone, hilus, and stratum-lacunosum-moleculare. GFAPδ-positive cells also stained for GFAPα. In AD donors, astrocytes near plaques displayed increased staining of both GFAPα and GFAPδ. The reading-frame-shifted isoform, GFAP(+1), staining was confined to a subset of astrocytes with long processes, and their number increased in the course of AD. In conclusion, the various GFAP isoforms show differential transcript levels and are upregulated in a concerted manner in AD. The GFAP(+1) isoform defines a unique subset of astrocytes, with numbers increasing with AD progression. These data indicate the need for future exploration of underlying mechanisms concerning the functions of GFAPδ and GFAP(+1) isoforms in astrocytes and their possible role in AD pathology. Copyright © 2014 Elsevier Inc. All rights reserved.

  11. Serum Glial Fibrillary Acidic Protein as a Specific Marker for Necrotizing Meningoencephalitis in Pug Dogs

    Science.gov (United States)

    MIYAKE, Hizuru; INOUE, Akiko; TANAKA, Miho; MATSUKI, Naoaki

    2013-01-01

    ABSTRACT To evaluate the ability of serum glial fibrillary acidic protein (GFAP) concentration as a diagnostic marker for canine central nervous system (CNS) disorders, sera from dogs with various CNS (n=47) and non-CNS (n=56) disorders were measured for GFAP by using an ELISA kit. Healthy Beagles (n=15) and Pug dogs (n=12) were also examined as controls. Interestingly, only Pug dogs with necrotizing meningoencephalitis (NME) showed elevated serum GFAP concentrations (<0.01 to 1.14 ng/ml), while other breeds of dogs with NME did not. Among the Pug dogs with NME, serum GFAP concentrations did not correlate with their clinical features, such as ages or survival times. Our data indicate the usefulness of serum GFAP as a novel marker for Pug dogs with NME. PMID:23856761

  12. GFAP and vimentin deficiency alters gene expression in astrocytes and microglia in wild-type mice and changes the transcriptional response of reactive glia in mouse model for Alzheimer's disease

    NARCIS (Netherlands)

    W. Kamphuis; L. Kooijman; M. Orre; O. Stassen; M. Pekny; E.M. Hol

    2015-01-01

    Reactive astrocytes with an increased expression of intermediate filament (IF) proteins Glial Fibrillary Acidic Protein (GFAP) and Vimentin (VIM) surround amyloid plaques in Alzheimer's disease (AD). The functional consequences of this upregulation are unclear. To identify molecular pathways coupled

  13. GFAP and S100B are biomarkers of traumatic brain injury : an observational cohort study

    NARCIS (Netherlands)

    Vos, P E; Jacobs, B; Andriessen, T M J C; Lamers, K J B; Borm, G F; Beems, T; Edwards, M; Rosmalen, C F; Vissers, J L M

    2010-01-01

    BACKGROUND: Biomarker levels in blood after traumatic brain injury (TBI) may offer diagnostic and prognostic tools in addition to clinical indices. This study aims to validate glial fibrillary acidic protein (GFAP) and S100B concentrations in blood as outcome predictors of TBI using cutoff levels of

  14. Intermediate filaments of zebrafish retinal and optic nerve astrocytes and Müller glia: differential distribution of cytokeratin and GFAP

    Directory of Open Access Journals (Sweden)

    Mosier Amanda L

    2010-03-01

    Full Text Available Abstract Background Optic nerve regeneration (ONR following injury is a model for central nervous system regeneration. In zebrafish, ONR is rapid - neurites cross the lesion and enter the optic tectum within 7 days; in mammals regeneration does not take place unless astrocytic reactivity is suppressed. Glial fibrillary acidic protein (GFAP is used as a marker for retinal and optic nerve astrocytes in both fish and mammals, even though it has long been known that astrocytes of optic nerves in many fish, including zebrafish, express cytokeratins and not GFAP. We used immunofluorescence to localize GFAP and cytokeratin in wild-type zebrafish and transgenic zebrafish expressing green fluorescent protein (GFP under control of a GFAP promoter to determine the pattern of expression of intermediate filaments in retina and optic nerve. Findings GFAP labeling and GFAP gene expression as indicated by GFP fluorescence was found only in the Müller glial cells of the retina. Within Müller cells, GFP fluorescence filled the entire cell while GFAP labelling was more restricted in distribution. No GFAP expression was observed in optic nerves. Cytokeratin labeling of astrocytes was observed throughout the optic nerve and less intensely in cells in the retinal inner plexiform layer. The retinal inner limiting membrane was strongly labeled by anti-cytokeratin. Conclusions Studies of astrocyte function during ONR in zebrafish cannot solely rely on GFAP as an astrocyte marker or indicator of reactivity. Future studies of ONR in zebrafish should include evaluation of changes in cytokeratin expression and localization in the optic nerve.

  15. Genetic background affects human glial fibrillary acidic protein promoter activity.

    Directory of Open Access Journals (Sweden)

    Xianshu Bai

    Full Text Available The human glial fibrillary acidic protein (hGFAP promoter has been used to generate numerous transgenic mouse lines, which has facilitated the analysis of astrocyte function in health and disease. Here, we evaluated the expression levels of various hGFAP transgenes at different ages in the two most commonly used inbred mouse strains, FVB/N (FVB and C57BL/6N (B6N. In general, transgenic mice maintained on the B6N background displayed weaker transgene expression compared with transgenic FVB mice. Higher level of transgene expression in B6N mice could be regained by crossbreeding to FVB wild type mice. However, the endogenous murine GFAP expression was equivalent in both strains. In addition, we found that endogenous GFAP expression was increased in transgenic mice in comparison to wild type mice. The activities of the hGFAP transgenes were not age-dependently regulated. Our data highlight the importance of proper expression analysis when non-homologous recombination transgenesis is used.

  16. The determination of glial fibrillary acidic protein for the diagnosis and histogenetic study of central nervous system tumors: a study of 152 cases.

    Directory of Open Access Journals (Sweden)

    Hamaya,Kazuo

    1985-12-01

    Full Text Available Glial fibrillary acidic protein (GFAP was purified from human spinal cord and cerebral white matter. GFAP was localized by an immuno-peroxidase method in normal adult and fetal human brains, rat brains, and 152 central nervous system (CNS tumors. GFAP was found in reactive and normal astrocytes, immature cells of fetal brain at the 18th to 21st gestational weeks, and normal rat astrocytes. This GFAP staining was quite specific for glial tumors, including astrocytomas, glioblastomas, astroblastomas, and ependymomas. GFAP-positive cells were also found in oligodendrogliomas and choroid plexus papillomas, and they were interpreted as being astroglial or ependymal differentiations. Stromal cells in cerebellar hemangioblastomas were negative. However, engulfed astrocytes were found at the periphery of such tumors and often adjacent to the proliferate blood vessels. In meningiomas, neurinomas, metastatic carcinomas, pituitary adenomas and other non-glial tumors, GFAP-positive cells were not identified.

  17. Increased GFAP and S100beta but not NSE serum levels after subarachnoid haemorrhage are associated with clinical severity.

    NARCIS (Netherlands)

    Vos, P.E.; Gils, M. van; Beems, T.; Zimmerman, C.; Verbeek, M.M.

    2006-01-01

    Assessment of initial disease severity after subarachnoid haemorrhage (SAH) remains difficult. The objective of the study is to identify biochemical markers of brain damage in peripheral blood after SAH. Hospital admission S100beta, glial fibrillary acidic protein (GFAP) and neuron-specific enolase

  18. Effect of Ferulic Acid on Learning-memory and Expression of GFAP in the Hippocampus Tissues of Alzheimer' s Disease-like Model Mice%阿魏酸对拟痴呆小鼠学习记忆和海马胶质 纤维酸性蛋白表达的影响

    Institute of Scientific and Technical Information of China (English)

    金蓓蓓; 陈勤; 陈庆林

    2011-01-01

    目的:探讨阿魏酸对阿尔茨海默病(AD)模型小鼠神经行为学和海马胶质纤维酸性蛋白(GFAP)表达的影响,分析阿魏酸对小鼠脑的保护作用.方法:海马CA1区注射微量红藻氨酸(KA)建立痴呆模型,然后对痴呆小鼠用不同剂量的阿魏酸(FA)灌胃治疗.Morris水迷宫实验观察小鼠行为学变化,免疫组织化学方法观察GFAP的表达.结果:与假手术组相比,模型组学习记忆能力明显降低(P<0.01)GFAP阳性细胞表达明显增多(P<0.01);与模型组相比,阿魏酸治疗组学习记忆能力均明显提高(P<O.01),GFAP阳性细胞表达均明显减少(P<0.01).结论:用不同剂量的阿魏酸治疗拟AD小鼠后,小鼠学习记忆能力得到明显改善,GFAP表达得到明显抑制,起到保护脑的作用.%Objective: To investigate the effects of ferulic acid on neurological behavior and the expression of glial fibril-lary acidic protein(GFAP) of hippocampus tissues in Alzheimer' s disease-like model mice, and to analyze the protective effects of ferulic acid on the brain. Methods: kainic acid ( KA) was injected into hippocampus CA1 region of mice and to establish Alzheimer' s disease-like model, then the drug group with different doses of ferulic acid gavage lasted a month. The learning and memory abilities of the mice were assessed through Morris water maze behavioral test, and GFAP were observed by immunohistochemistry respectively. Results: Compared with the normal group, the abilities of learning and memory of the mice in the model group significantly decreased (P < 0.01 ) and the expression of GFAP in the CA1 region were increased(P<0.01). Compared with the model group, the abilities of learning and memory of the mice in the ferulic acid group improved ( P < 0. 01 ) and the expressions of GFAP in the CA1 region decreased ( P < 0.01). Conclusion: The different doses of ferulic acid dealing with the Alzheimer disease-like model mice can improve the abilities of learning

  19. Levels and Age Dependency of Neurofilament Light and Glial Fibrillary Acidic Protein in Healthy Individuals and Their Relation to the Brain Parenchymal Fraction.

    Science.gov (United States)

    Vågberg, Mattias; Norgren, Niklas; Dring, Ann; Lindqvist, Thomas; Birgander, Richard; Zetterberg, Henrik; Svenningsson, Anders

    2015-01-01

    Neurofilament light (NFL) and Glial Fibrillary Acidic Protein (GFAP) are integral parts of the axonal and astrocytal cytoskeletons respectively and are released into the cerebrospinal fluid (CSF) in cases of cellular damage. In order to interpret the levels of these biomarkers in disease states, knowledge on normal levels in the healthy is required. Another biomarker for neurodegeneration is brain atrophy, commonly measured as brain parenchymal fraction (BPF) using magnetic resonance imaging (MRI). Potential correlations between levels of NFL, GFAP and BPF in healthy individuals have not been investigated. To present levels of NFL and GFAP in healthy individuals stratified for age, and investigate the correlation between them as well as their correlation with BPF. The CSF was analysed in 53 healthy volunteers aged 21 to 70 (1 sample missing for GFAP analysis) and 48 of the volunteers underwent determination of BPF using MRI. Mean (±SD) NFL was 355 ng/L (±214), mean GFAP was 421 ng/L (±129) and mean BPF was 0.867 (±0.035). All three biomarkers correlated with age. NFL also correlated with both GFAP and BPF. When controlled for age, only the correlation between NFL and GFAP retained statistical significance. This study presents data on age-stratified levels of NFL and GFAP in the CSF of healthy individuals. There is a correlation between levels of NFL and GFAP and both increase with age. A correlation between NFL and BPF was also found, but did not retain statistical significance if controlled for age.

  20. Withaferin A Targets Intermediate Filaments Glial Fibrillary Acidic Protein and Vimentin in a Model of Retinal Gliosis*

    OpenAIRE

    2010-01-01

    Gliosis is a biological process that occurs during injury repair in the central nervous system and is characterized by the overexpression of the intermediate filaments (IFs) glial fibrillary acidic protein (GFAP) and vimentin. A common thread in many retinal diseases is reactive Müller cell gliosis, an untreatable condition that leads to tissue scarring and even blindness. Here, we demonstrate that the vimentin-targeting small molecule withaferin A (WFA) is a novel chemical probe of GFAP. Usi...

  1. Time course degeneration and expression of glial fibrillary acidic protein in mer-knockout mice

    Institute of Scientific and Technical Information of China (English)

    LIANG Xiao-ying; WANG Huai-zhou; WANG Ning-li

    2010-01-01

    Background Muller cells in the mammalian retina normally express low levels of glial fibrillary acidic protein (GFAP); however, its expression is upregulated in response to the loss of retinal neurons. The change in expression of GFAP is one of the earliest indicators of retinal damage and is correlated with the time course of disease. The aim of this study was to investigate the time course of degeneration and the expression of GFAP in the retina of mer knockout mice. Methods A total of 30 mer knockout mice, aged from 15-20 days to 1 year and 32 age-matched wild type mice as controls were tested. Immunohistochemistry was used to show the expression of GFAP in the central and peripheral retina of mer knockout and control mice at postnatal age of 15 days (P15d), 20 days (P20d), 4 weeks (P4w), 6 weeks (P6w), 8 weeks (P8w), 3 months (P3m), 6 months (P6m) and 1 years (P1y).Results The expression of GFAP in the central and peripheral retina of wild type mice was limited to the retinal ganglion cell and nerve fiber layers. In the central retina of mer knockout mice, GFAP expression was upregulated at P4w and GFAP immunolabelling penetrates across the entire thickness of the retina at P8w; whereas in the peripheral retina, the GFAP expression was upregulated at P20d and GFAP immunolabelling penetrates the entire retina after P4w. Conclusions Increased expression of GFAP in Muller cells of mer knockout mice occur at P20d in the peripheral retina and P4w in the central retina. GFAP expression in Muller cells appears to be a secondary response to the loss of retinal neurons. Increased expression of GFAP may occur prior to any detectable morphological changes in the retina. This study suggests that the loss of retinal neurons may begin in the early stages of retinitis pigmentosa, prior to the discovery of any morphological changes in the retina.

  2. GFAP mutations, age at onset, and clinical subtypes in Alexander disease.

    Science.gov (United States)

    Prust, M; Wang, J; Morizono, H; Messing, A; Brenner, M; Gordon, E; Hartka, T; Sokohl, A; Schiffmann, R; Gordish-Dressman, H; Albin, R; Amartino, H; Brockman, K; Dinopoulos, A; Dotti, M T; Fain, D; Fernandez, R; Ferreira, J; Fleming, J; Gill, D; Griebel, M; Heilstedt, H; Kaplan, P; Lewis, D; Nakagawa, M; Pedersen, R; Reddy, A; Sawaishi, Y; Schneider, M; Sherr, E; Takiyama, Y; Wakabayashi, K; Gorospe, J R; Vanderver, A

    2011-09-27

    To characterize Alexander disease (AxD) phenotypes and determine correlations with age at onset (AAO) and genetic mutation. AxD is an astrogliopathy usually characterized on MRI by leukodystrophy and caused by glial fibrillary acidic protein (GFAP) mutations. We present 30 new cases of AxD and reviewed 185 previously reported cases. We conducted Wilcoxon rank sum tests to identify variables scaling with AAO, survival analysis to identify predictors of mortality, and χ(2) tests to assess the effects of common GFAP mutations. Finally, we performed latent class analysis (LCA) to statistically define AxD subtypes. LCA identified 2 classes of AxD. Type I is characterized by early onset, seizures, macrocephaly, motor delay, encephalopathy, failure to thrive, paroxysmal deterioration, and typical MRI features. Type II is characterized by later onset, autonomic dysfunction, ocular movement abnormalities, bulbar symptoms, and atypical MRI features. Survival analysis predicted a nearly 2-fold increase in mortality among patients with type I AxD relative to those with type II. R79 and R239 GFAP mutations were most common (16.6% and 20.3% of all cases, respectively). These common mutations predicted distinct clinical outcomes, with R239 predicting the most aggressive course. AAO and the GFAP mutation site are important clinical predictors in AxD, with clear correlations to defined patterns of phenotypic expression. We propose revised AxD subtypes, type I and type II, based on analysis of statistically defined patient groups.

  3. Autoantibody to glial fibrillary acidic protein in the sera of cattle with bovine spongiform encephalopathy.

    Science.gov (United States)

    Nomura, Sachiko; Miyasho, Taku; Maeda, Naoyuki; Doh-ura, Katsumi; Yokota, Hiroshi

    2009-08-01

    It is desirable to make the diagnosis in live cattle with bovine spongiform encephalopathy (BSE), and thus surrogate markers for the disease have been eagerly sought. Serum proteins from BSE cattle were analyzed by 2-D Western blotting and TOF-MS. Autoantibodies against proteins in cytoskeletal fractions prepared from normal bovine brains were found in the sera of BSE cattle. The protein recognized was identified to be glial fibrillary acidic protein (GFAP), which is expressed mainly in astrocytes in the brain. The antigen protein, GFAP, was also found in the sera of BSE cattle. The percentages of both positive sera in the autoantibody and GFAP were 44.0% for the BSE cattle, 0% for the healthy cattle, and 5.0% for the clinically suspected BSE-negative cattle. A significant relationship between the presence of GFAP and the expression of its autoantibody in the serum was recognized in the BSE cattle. These findings suggest a leakage of GFAP into the peripheral blood during neurodegeneration associated with BSE, accompanied by the autoantibody production, and might be useful in understanding the pathogenesis and in developing a serological diagnosis of BSE in live cattle.

  4. Anorexia Reduces GFAP+ Cell Density in the Rat Hippocampus

    Directory of Open Access Journals (Sweden)

    Daniel Reyes-Haro

    2016-01-01

    Full Text Available Anorexia nervosa is an eating disorder observed primarily in young women. The neurobiology of the disorder is unknown but recently magnetic resonance imaging showed a volume reduction of the hippocampus in anorexic patients. Dehydration-induced anorexia (DIA is a murine model that mimics core features of this disorder, including severe weight loss due to voluntary reduction in food intake. The energy supply to the brain is mediated by astrocytes, but whether their density is compromised by anorexia is unknown. Thus, the aim of this study was to estimate GFAP+ cell density in the main regions of the hippocampus (CA1, CA2, CA3, and dentate gyrus in the DIA model. Our results showed that GFAP+ cell density was significantly reduced (~20% in all regions of the hippocampus, except in CA1. Interestingly, DIA significantly reduced the GFAP+ cells/nuclei ratio in CA2 (−23% and dentate gyrus (−48%. The reduction of GFAP+ cell density was in agreement with a lower expression of GFAP protein. Additionally, anorexia increased the expression of the intermediate filaments vimentin and nestin. Accordingly, anorexia increased the number of reactive astrocytes in CA2 and dentate gyrus more than twofold. We conclude that anorexia reduces the hippocampal GFAP+ cell density and increases vimentin and nestin expression.

  5. Anorexia Reduces GFAP+ Cell Density in the Rat Hippocampus

    Science.gov (United States)

    Labrada-Moncada, Francisco Emmanuel; Varman, Durairaj Ragu; Krüger, Janina; Morales, Teresa; Miledi, Ricardo; Martínez-Torres, Ataúlfo

    2016-01-01

    Anorexia nervosa is an eating disorder observed primarily in young women. The neurobiology of the disorder is unknown but recently magnetic resonance imaging showed a volume reduction of the hippocampus in anorexic patients. Dehydration-induced anorexia (DIA) is a murine model that mimics core features of this disorder, including severe weight loss due to voluntary reduction in food intake. The energy supply to the brain is mediated by astrocytes, but whether their density is compromised by anorexia is unknown. Thus, the aim of this study was to estimate GFAP+ cell density in the main regions of the hippocampus (CA1, CA2, CA3, and dentate gyrus) in the DIA model. Our results showed that GFAP+ cell density was significantly reduced (~20%) in all regions of the hippocampus, except in CA1. Interestingly, DIA significantly reduced the GFAP+ cells/nuclei ratio in CA2 (−23%) and dentate gyrus (−48%). The reduction of GFAP+ cell density was in agreement with a lower expression of GFAP protein. Additionally, anorexia increased the expression of the intermediate filaments vimentin and nestin. Accordingly, anorexia increased the number of reactive astrocytes in CA2 and dentate gyrus more than twofold. We conclude that anorexia reduces the hippocampal GFAP+ cell density and increases vimentin and nestin expression. PMID:27579183

  6. GFAP expression in astrocytes of suprachiasmatic nucleus and medial preoptic area are differentially affected by malnutrition during rat brain development.

    Science.gov (United States)

    Mendonça, João Esmeraldo Frota; Vilela, Maria Cristina Ramos; Bittencourt, Heitor; Lapa, Raíssa Maria; Oliveira, Francisco Gilberto; Alessio, Maria Luíza Martins; Guedes, Rubem Carlos Araújo; De Oliveira Costa, Miriam Stela Maris; Da Costa, Belmira Lara da Silveira Andrade

    2004-08-01

    The aim of the present study was investigate, in young rats, the effects of malnutrition on astrocyte distribution of two hypothalamic regions, the circadian pacemaker suprachiasmatic nucleus (SCN) and the medial preoptic area (MPA). Control rats were born from mothers fed on commercial diet since gestation and malnourished rats from mothers fed on multideficient diet, from the beginning of gestation (GLA group) or from the onset of lactation (LA group). After weaning, pups received ad libitum the same diet as their mothers, and were maintained under a 12/12 h light/dark cycle. The animals were analyzed either at 30-33, or 60-63 days of life. Brain coronal sections (50 microm) were processed to visualize glial fibrillary acidic protein (GFAP) immunoreactivity. Compared to control rats, both malnourished groups of 30 and 60 days exhibited a reduced number of GFAP-immunoreactive astrocytes in the SCN. The total GFAP-immunoreactive area in the SCN of the GLA group differed from the control group at both age ranges analyzed. The GFAP expression as measured by the relative optical density (ROD) exhibited a 50-60% reduction in the MPA in both malnourished groups, compared to controls. The results suggest that malnutrition early in life leads to alterations in gliogenesis or glial cell proliferation in both nuclei, being these alterations greater in the MPA. Compensatory plasticity mechanisms in the GFAP-expression seem to be developed in the astrocyte differentiation process in the SCN, especially when the malnutrition is installed from the lactation.

  7. GFAP and vimentin deficiency alters gene expression in astrocytes and microglia in wild-type mice and changes the transcriptional response of reactive glia in mouse model for Alzheimer's disease.

    Science.gov (United States)

    Kamphuis, Willem; Kooijman, Lieneke; Orre, Marie; Stassen, Oscar; Pekny, Milos; Hol, Elly M

    2015-06-01

    Reactive astrocytes with an increased expression of intermediate filament (IF) proteins Glial Fibrillary Acidic Protein (GFAP) and Vimentin (VIM) surround amyloid plaques in Alzheimer's disease (AD). The functional consequences of this upregulation are unclear. To identify molecular pathways coupled to IF regulation in reactive astrocytes, and to study the interaction with microglia, we examined WT and APPswe/PS1dE9 (AD) mice lacking either GFAP, or both VIM and GFAP, and determined the transcriptome of cortical astrocytes and microglia from 15- to 18-month-old mice. Genes involved in lysosomal degradation (including several cathepsins) and in inflammatory response (including Cxcl5, Tlr6, Tnf, Il1b) exhibited a higher AD-induced increase when GFAP, or VIM and GFAP, were absent. The expression of Aqp4 and Gja1 displayed the same pattern. The downregulation of neuronal support genes in astrocytes from AD mice was absent in GFAP/VIM null mice. In contrast, the absence of IFs did not affect the transcriptional alterations induced by AD in microglia, nor was the cortical plaque load altered. Visualizing astrocyte morphology in GFAP-eGFP mice showed no clear structural differences in GFAP/VIM null mice, but did show diminished interaction of astrocyte processes with plaques. Microglial proliferation increased similarly in all AD groups. In conclusion, absence of GFAP, or both GFAP and VIM, alters AD-induced changes in gene expression profile of astrocytes, showing a compensation of the decrease of neuronal support genes and a trend for a slightly higher inflammatory expression profile. However, this has no consequences for the development of plaque load, microglial proliferation, or microglial activation. © 2015 Wiley Periodicals, Inc.

  8. DISTRIBUTION OF GLIAL FIBRILLARY ACIDIC PROTEIN IN DIFFERENT PARTS OF THE RAT BRAIN UNDER CADMIUM EXPOSURE.

    Science.gov (United States)

    Kovalchuk, Yu P; Prischepa, I V; Si, U; Nedzvetsky, V S; Kot, Y G; Persky, E E; Ushakova, G A

    2015-01-01

    The chronic effects of low doses of cadmium on the distribution of soluble and filament forms of glial fibrillary acidic protein (GFAP) and their polypeptide fragments in different parts of the rat brain were investigated. Obtained results showed dose-dependent effect of cadmium on the soluble form of GFAP and more pronounced effect on the filament form and composition of the polypeptide fragments of the protein in the rat brain. Prolonged intoxication by cadmium ions in a dose of 1.0 μg/kg of body weight induced a significant decrease in soluble GFAP and an increase in the filament form in the rat brain, pointing to the development of reactive astrogliosis and the risk of neurodegeneration.

  9. Changes in the distribution of the neuron-specific B-50, neurofilament protein and glial fibrillary acidic proteins following an unilateral mesencephalic lesion in the rat

    NARCIS (Netherlands)

    Gispen, W.H.; Oestreicher, A.B.; Devay, P.; Isaacson, R.L.

    1988-01-01

    Following a unilateral electrolytic lesion in the ventral rat mesencephalon, changes in the immunocytochemical distribution of the neuron-specific B-50, neurofilament (NF) protein and glial fibrillary acidic (GFAP) proteins were studied around the lesion after 0, 3, 10 and 28 days. At all recovery t

  10. Hippocampal kindling alters the concentration of glial fibrillary acidic protein and other marker proteins in rat brain

    DEFF Research Database (Denmark)

    Hansen, A; Jørgensen, Ole Steen; Bolwig, T G;

    1990-01-01

    The effect of hippocampal kindling on neuronal and glial marker proteins was studied in the rat by immunochemical methods. In hippocampus, pyriform cortex and amygdala there was an increase in glial fibrillary acidic protein (GFAP), indicating reactive gliosis, and an increase in the glycolytic...... enzyme NSE, suggesting increased anaerobic metabolism. Neuronal cell adhesion molecule (NCAM) decreased in pyriform cortex and amygdala of kindled rats, indicating neuronal degeneration....

  11. Intermediate filaments of zebrafish retinal and optic nerve astrocytes and Müller glia: differential distribution of cytokeratin and GFAP

    OpenAIRE

    Mosier Amanda L; Koke Joseph R; García Dana M

    2010-01-01

    Abstract Background Optic nerve regeneration (ONR) following injury is a model for central nervous system regeneration. In zebrafish, ONR is rapid - neurites cross the lesion and enter the optic tectum within 7 days; in mammals regeneration does not take place unless astrocytic reactivity is suppressed. Glial fibrillary acidic protein (GFAP) is used as a marker for retinal and optic nerve astrocytes in both fish and mammals, even though it has long been known that astrocytes of optic nerves i...

  12. Phenotypes of articular disc cells in the rat temporomandibular joint as demonstrated by immunohistochemistry for nestin and GFAP.

    Science.gov (United States)

    Miyako, Hitoshi; Suzuki, Akiko; Nozawa-Inoue, Kayoko; Magara, Jin; Kawano, Yoshiro; Ono, Kazuhiro; Maeda, Takeyasu

    2011-10-01

    The articular disc is a dense collagenous tissue containing disc cells that are phenotypically described as chondrocyte-like cells or fibrochondrocytes. Despite the possible existence of these phenotypes in systemic joints, little is known about the detailed classification of the articular disc cells in the temporomandibular joint. In this immunocytochemical study we examined the localization and distribution patterns of nestin and glial fibrillary acidic protein (GFAP) in the articular disc of the rat temporomandibular joint at postnatal day 1, and weeks 1, 2, 4 and 8, based on the status of tooth eruption and occlusion. Nestin and GFAP are intermediate filament proteins whose expression patterns are closely related to cell differentiation and cell migration. Both types of immunopositive cell greatly increased postnatally to a stable level after postnatal week 4, but they showed different distribution patterns and cell morphologies. Nestin-reactive disc cells, which were characterized by a meagre cytoplasm and thin cytoplasmic processes, were scattered in the articular disc, whereas GFAP-positive cells, characterized by broader processes, existed exclusively in the deeper area. In mature discs, the major proportion of articular disc cells exhibited GFAP immunoreactivity. Furthermore, a double-immunostaining demonstrated that the nestin-negative cells, consisting of GFAP-positive and -negative cells, exhibited immunoreactions for heat shock protein 25. These findings indicate that the articular disc cells comprise at least three types in the rat temporomandibular joint and suggest that their expressions closely relate to mechanical loading forces within the joint, including occlusal force, as observed through postnatal development.

  13. Intranasal cotinine improves memory, and reduces depressive-like behavior, and GFAP+ cells loss induced by restraint stress in mice.

    Science.gov (United States)

    Perez-Urrutia, Nelson; Mendoza, Cristhian; Alvarez-Ricartes, Nathalie; Oliveros-Matus, Patricia; Echeverria, Florencia; Grizzell, J Alex; Barreto, George E; Iarkov, Alexandre; Echeverria, Valentina

    2017-09-01

    Posttraumatic stress disorder (PTSD), chronic psychological stress, and major depressive disorder have been found to be associated with a significant decrease in glial fibrillary acidic protein (GFAP) immunoreactivity in the hippocampus of rodents. Cotinine is an alkaloid that prevents memory impairment, depressive-like behavior and synaptic loss when co-administered during restraint stress, a model of PTSD and stress-induced depression, in mice. Here, we investigated the effects of post-treatment with intranasal cotinine on depressive- and anxiety-like behaviors, visual recognition memory as well as the number and morphology of GFAP+ immunoreactive cells, in the hippocampus and frontal cortex of mice subjected to prolonged restraint stress. The results revealed that in addition to the mood and cognitive impairments, restraint stress induced a significant decrease in the number and arborization of GFAP+ cells in the brain of mice. Intranasal cotinine prevented these stress-derived symptoms and the morphological abnormalities GFAP+ cells in both of these brain regions which are critical to resilience to stress. The significance of these findings for the therapy of PTSD and depression is discussed. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Inhalation exposure to white spirit causes region-dependent alterations in the levels of glial fibrillary acidic protein

    DEFF Research Database (Denmark)

    Lam, Henrik Rye; Ladefoged, Ole; østergaard, G.

    2000-01-01

    Enhanced expression of glial fibrillary acidic protein (GFAP) is known to be associated with toxicant-induced gliosis, a homotypic response of the central nervous system to neural injury. A variety of neurochemical and neurophysiological effects have been observed in experimental animals exposed ...

  15. Serum levels of GFAP and EGFR in primary and recurrent high-grade gliomas: correlation to tumor volume, molecular markers, and progression-free survival.

    Science.gov (United States)

    Kiviniemi, Aida; Gardberg, Maria; Frantzén, Janek; Parkkola, Riitta; Vuorinen, Ville; Pesola, Marko; Minn, Heikki

    2015-09-01

    Our aim was to study the association of two potential serum biomarkers glial fibrillary acidic protein (GFAP) and epidermal growth factor receptor (EGFR) with prognostic markers such as IDH1 mutation, tumor burden, and survival in patients with high-grade gliomas (HGG). Additionally, our objective was to evaluate the potential of serum EGFR as a surrogate marker for EGFR status in the tumor. Pre-operative serum samples were prospectively collected from patients with primary (n = 17) or recurrent (n = 10) HGG. Serum GFAP and EGFR levels were determined by ELISA and studied for correlation with molecular markers including EGFR amplification, tumor volume in contrast-enhanced T1-weighted MRI, and progression-free survival (PFS). Pre-operative serum GFAP level of ≥0.014 ng/ml was 86 % sensitive and 85 % specific for the diagnosis of glioblastoma. High GFAP was related to the lack of IDH1 mutation (P = 0.016), high Ki67 proliferation index (P < 0.001), and poor PFS (HR 5.9, CI 1.2-29.9, P = 0.032). Serum GFAP correlated with enhancing tumor volume in primary (r = 0.64 P = 0.005), but also in recurrent HGGs (r = 0.76 P = 0.011). In contrast, serum EGFR levels did not differ between HGG patients and 13 healthy controls, and were not related to EGFR status in the tumor. We conclude that high serum GFAP associates with IDH1 mutation-negative HGG, and poor PFS. Correlation with tumor burden in recurrent HGG implicates the potential of serum GFAP for detection of tumor recurrence. Our results suggest that circulating EGFR is not derived from glioma cells and cannot be used as a marker for EGFR status in the tumor.

  16. Effects of endurance exercise on expressions of glial fibrillary acidic protein and myelin basic protein in developing rats with maternal infection-induced cerebral palsy.

    Science.gov (United States)

    Kim, Kijeong; Shin, Mal-Soon; Cho, Han-Sam; Kim, Young-Pyo

    2014-02-01

    Periventricular leukomalacia (PVL) is a common white matter lesion affecting the neonatal brain. PVL is closely associated with cerebral palsy (CP) and characterized by increase in the number of astrocytes, which can be detected by positivity for glial fibrillary acidic protein (GFAP). Change in myelin basic protein (MBP) is an early sign of white matter abnormality. Maternal or placental infection can damage the neonatal brain. In the present study, we investigated the effects of treadmill walking exercise on GFAP and MBP expressions in rats with maternal lipopolysaccharide (LPS)-induced PVL. Immunohistochemistry was performed for the detection of GFAP and MBP. The present results showed that intracervical maternal LPS injection during pregnancy increased GFAP expression in the striatum and decreased MBP expression in the corpus callosum of rats. The results also showed that treadmill walking exercise suppressed GFAP expression and enhanced MBP expression in the brains of rats with maternal LPS-induced PVL. The present study revealed that treadmill walking exercise is effective for the suppressing astrogliosis and hypomyelination associated with PVL. Here in this study, we showed that treadmill walking exercise may be effective therapeutic strategy for alleviating the detrimental effects of CP.

  17. GFAP and Fos immunoreactivity in lumbo-sacral spinal cord and medulla oblongata after chronic colonic inflammation in rats

    Institute of Scientific and Technical Information of China (English)

    Yi-Ning Sun; Jin-Yan Luo; Zhi-Ren Rao; Li Lan; Li Duan

    2005-01-01

    AIM:- To investigate the response of astrocytes and neurons in rat lumbo-sacral spinal cord and medulla oblongata induced by chronic colonic inflammation, and the relationship between them.METHODS: Thirty-three male Sprague-Dawley rats were randomly divided into two groups: experimental group (n = 17), colonic inflammation was induced by intra-luminal administration of trinitrobenzenesulfonic acid (TNBS);control group (n = 16), saline was administered intra-luminally.After 3, 7, 14, and 28 d of administration, the lumbo-sacral spinal cord and medulla oblongata were removed and processed for anti-glial fibrillary acidic protein (GFAP),Fos and GFAP/Fos immunohistochemistry.RESULTS: Activated astrocytes positive for GFAP were mainly distributed in the superficial laminae (laminae Ⅰ-Ⅱ)of dorsal horn, intermediolateral nucleus (laminae V),posterior commissural nucleus (laminae X) and anterolateral nucleus (laminae Ⅸ). Fos-IR (Fos-immunoreactive)neurons were mainly distributed in the deeper laminae of the spinal cord (laminae Ⅲ-Ⅳ, V-Ⅵ). In the medulla oblongata, both GFAP-IR astrocytes and Fos-IR neurons were mainly distributed in the medullary visceral zone (MVZ). The density of GFAP in the spinal cord of experimental rats was significantly higher after 3, 7, and 14 d of TNBS administration compared with the controls (50.4±16.8,29.2±6.5, 24.1±5.6, P<0.05). The density of GFAP in MVZ was significantly higher after 3 d of TNBS administration (34.3±2.5, P<0.05). After 28 d of TNBS administration,the density of GFAP in the spinal cord and MVZ decreased and became comparable to that of the controls (18.0±4.9,14.6±6.4, P>0.05).CONCLUSION: Astrocytes in spinal cord and medulla oblongata can be activated by colonic inflammation. The activated astrocytes are closely related to Fos-IR neurons.With the recovery of colonic inflammation, the activity of astrocytes in the spinal cord and medulla oblongata is reduced.

  18. Levels and Age Dependency of Neurofilament Light and Glial Fibrillary Acidic Protein in Healthy Individuals and Their Relation to the Brain Parenchymal Fraction.

    Directory of Open Access Journals (Sweden)

    Mattias Vågberg

    Full Text Available Neurofilament light (NFL and Glial Fibrillary Acidic Protein (GFAP are integral parts of the axonal and astrocytal cytoskeletons respectively and are released into the cerebrospinal fluid (CSF in cases of cellular damage. In order to interpret the levels of these biomarkers in disease states, knowledge on normal levels in the healthy is required. Another biomarker for neurodegeneration is brain atrophy, commonly measured as brain parenchymal fraction (BPF using magnetic resonance imaging (MRI. Potential correlations between levels of NFL, GFAP and BPF in healthy individuals have not been investigated.To present levels of NFL and GFAP in healthy individuals stratified for age, and investigate the correlation between them as well as their correlation with BPF.The CSF was analysed in 53 healthy volunteers aged 21 to 70 (1 sample missing for GFAP analysis and 48 of the volunteers underwent determination of BPF using MRI.Mean (±SD NFL was 355 ng/L (±214, mean GFAP was 421 ng/L (±129 and mean BPF was 0.867 (±0.035. All three biomarkers correlated with age. NFL also correlated with both GFAP and BPF. When controlled for age, only the correlation between NFL and GFAP retained statistical significance.This study presents data on age-stratified levels of NFL and GFAP in the CSF of healthy individuals. There is a correlation between levels of NFL and GFAP and both increase with age. A correlation between NFL and BPF was also found, but did not retain statistical significance if controlled for age.

  19. Scorpion Venom Heat-Resistant Peptide Attenuates Glial Fibrillary Acidic Protein Expression via c-Jun/AP-1.

    Science.gov (United States)

    Cao, Zhen; Wu, Xue-Fei; Peng, Yan; Zhang, Rui; Li, Na; Yang, Jin-Yi; Zhang, Shu-Qin; Zhang, Wan-Qin; Zhao, Jie; Li, Shao

    2015-11-01

    Scorpion venom has been used in the Orient to treat central nervous system diseases for many years, and the protein/peptide toxins in Buthus martensii Karsch (BmK) venom are believed to be the effective components. Scorpion venom heat-resistant peptide (SVHRP) is an active component of the scorpion venom extracted from BmK. In a previous study, we found that SVHRP could inhibit the formation of a glial scar, which is characterized by enhanced glial fibrillary acidic protein (GFAP) expression, in the epileptic hippocampus. However, the cellular and molecular mechanisms underlying this process remain to be clarified. The results of the present study indicate that endogenous GFAP expression in primary rat astrocytes was attenuated by SVHRP. We further demonstrate that the suppression of GFAP was primarily mediated by inhibiting both c-Jun expression and its binding with AP-1 DNA binding site and other factors at the GFAP promoter. These results support that SVHRP contributes to reducing GFAP at least in part by decreasing the activity of the transcription factor AP-1. In conclusion, the effects of SVHRP on astrocytes with respect to the c-Jun/AP-1 signaling pathway in vitro provide a practical basis for studying astrocyte activation and inhibition and a scientific basis for further studies of traditional medicine.

  20. UCH-L1 and GFAP Serum Levels in Neonates with Hypoxic-Ischemic Encephalopathy: A single center pilot study

    Directory of Open Access Journals (Sweden)

    Martha V. Douglas-Escobar

    2014-12-01

    Full Text Available Objective - We examined two potential biomarkers of brain damage in HIE neonates: glial fibrillary acidic protein (GFAP; a marker of gliosis and ubiquitin C-terminal hydrolase L1 (UCH-L1; a marker of neuronal injury. We hypothesized the biomarkers would be measurable in cord blood of healthy neonates and could serve as a normative reference for brain injury in HIE infants. Further, we hypothesized that serum samples of HIE neonates would have higher levels and would correlate with brain damage on MRI and later developmental outcomes.Study Design - Serum UCH - L1 and GFAP concentrations from HIE neonates(n = 16 were compared with controls(n = 11.Pearson correlation coefficients and a mixed model design examined the relationship between biomarker concentrations of HIE neonates and brain damage(MRI and developmental outcomes(Bayley - III.Result– Both biomarkers were detected in cord blood from control subjects.UCH - L1 concentrations were higher in HIE neonates(p < 0.001 and associated with cortical injury(p < 0.055 and later motor and cognitive developmental outcomes(p < 0.05.The temporal change in GFAP concentrations from birth to 96 hours of age predicted motor developmental outcomes(p < 0.05 and injury to the basal ganglia and white matter.Conclusion– UCH - L1 concentrations correlated with cortical injury and developmental delays and GFAP concentrations correlated with basal ganglia and white matter injury and motor delay in HIE affected patients.Researchers should continue to explore UCH - L1 and GFAP as promising serum biomarkers of brain damage and predictors of neurodevelopmental outcomes in neonates with HIE.

  1. Glial fibrillary acidic protein is a body fluid biomarker for glial pathology in human disease.

    Science.gov (United States)

    Petzold, Axel

    2015-03-10

    This review on the role of glial fibrillary acidic protein (GFAP) as a biomarker for astroglial pathology in neurological diseases provides background to protein synthesis, assembly, function and degeneration. Qualitative and quantitative analytical techniques for the investigation of human tissue and biological fluid samples are discussed including partial lack of parallelism and multiplexing capabilities. Pathological implications are reviewed in view of immunocytochemical, cell-culture and genetic findings. Particular emphasis is given to neurodegeneration related to autoimmune astrocytopathies and to genetic gain of function mutations. The current literature on body fluid levels of GFAP in human disease is summarised and illustrated by disease specific meta-analyses. In addition to the role of GFAP as a diagnostic biomarker for chronic disease, there are important data on the prognostic value for acute conditions. The published evidence permits to classify the dominant GFAP signatures in biological fluids. This classification may serve as a template for supporting diagnostic criteria of autoimmune astrocytopathies, monitoring disease progression in toxic gain of function mutations, clinical treatment trials (secondary outcome and toxicity biomarker) and provide prognostic information in neurocritical care if used within well defined time-frames.

  2. Glial fibrillary acidic protein as an early marker of hepatic stellate cell activation in chronic and posttransplant recurrent hepatitis C.

    Science.gov (United States)

    Carotti, Simone; Morini, Sergio; Corradini, Stefano Ginanni; Burza, Maria Antonella; Molinaro, Antonio; Carpino, Guido; Merli, Manuela; De Santis, Adriano; Muda, Andrea Onetti; Rossi, Massimo; Attili, Adolfo Francesco; Gaudio, Eugenio

    2008-06-01

    Activated alpha-smooth muscle actin (alpha-SMA)-positive hepatic stellate cells (HSCs) are pericytes responsible for fibrosis in chronic liver injury. The glial fibrillary acidic protein (GFAP), commonly expressed by astrocytes in the central nervous system, is expressed in vivo in the liver in a subpopulation of quiescent stellate cells. In the rat, increased GFAP expression in the acute response to injury and down-regulation in the chronic response have been observed, whereas reports concerning GFAP expression in human liver are still conflicting. We investigated the utility of GFAP compared to alpha-SMA as an immunohistochemical marker of early activated HSCs in chronic and posttransplant recurrent hepatitis C and correlated GFAP expression with vascular remodeling and fibrosis progression. With immunohistochemistry and a semiquantitative scoring system, the expression of GFAP and alpha-SMA in HSCs and the microvessel density were analyzed in biopsies from normal livers obtained from cadaveric donors [donor liver (DL); n = 21] and from livers from posttransplant hepatitis C virus recurrent hepatitis (HCV-PTR) patients (n = 19), hepatitis C virus chronic hepatitis (HCV-CH) patients, (n = 12), and hepatitis C virus cirrhosis (HCV-C) patients (n = 16). The percentage of alpha-SMA-positive HSCs was significantly higher in the HCV-PTR, HCV-CH, and HCV-C groups compared to the DL group (P HCV-PTR group compared to the DL, HCV-C (P HCV-CH (P HCV-CH group compared to the DL group (P HCV-PTR group, the percentage of GFAP-positive HSCs correlated with fibrosis progression (P HCV-CH and seems to predict fibrosis progression in HCV-PTR.

  3. Proliferation and differentiation of reactive nestin~+/GFAP~+ cells in an adult rat model of compression-induced spinal cord injury

    Institute of Scientific and Technical Information of China (English)

    Pinglin Yang; Xijing He; Haopeng Li; Binshang Lan; Guoyu Wang; Yiheng Liu

    2009-01-01

    BACKGROUND:Studies have demonstrated that astrocytes may possess similar properties to neural stem cells/neural precursor cells and have the potential to differentiate into neurons.OBJECTIVE:To observe neuroepithelial stem cell protein (nestin) and glial fibrillary acidic protein (GFAP) expression following spinal cord injury,and to explore whether nestin~+/GFAP~+ cells,which are detected at peak levels in gray and white matter around the ependymal region of the central canal in injured spinal cord,possess similar properties of neural stem cells.DESIGN,TIME AND SETTING:A randomized,controlled experiment.The study was performed at the Key Laboratory of Environment and Genes Related to Diseases (Xi'an Jiaotong University),Ministry of Education between January 2004 and December 2006.MATERIALS:Rabbit anti-rat nestin,β-tubulin Ⅲ,mouse anti-rat GFAP,galactocerebroside (GaLC) antibodies were utilized,as well as flow cytometry.METHODS:A total of 60 male,Sprague Dawiey rats,aged 8 weeks,were randomly assigned to control (n=12) and model (n=48) groups.The spinal cord injury model was established in the model group by aneurysm clip compression,while the control animals were not treated.The gray and white matter around the ependymal region of the central canal exhibited peak expression of nestin~+/GFAP~+ cells.These cells were harvested and prepared into single cell suspension,followed by primary and passage cultures.The cells were incubated with serum-containing neural stem cell complete medium.MAIN OUTCOME MEASURES:Nestin and GFAP expression in injured spinal cord was determined using immunohistochemistry and double-labeled immunofluorescence at 1,3,5,7,14,28,and 56 days post-injury.In addition,cell proliferation and differentiation were detected using immunofluorescence cytochemistry and flow cytometry.RESULTS:Compared with the control group,the model group exhibited significantly increased nestin and GFAP expression (P<0.05),which reached peak levels between 3 and 7

  4. Glial fibrillary acidic protein is elevated in the lysosomal storage disease classical late-infantile neuronal ceroid lipofuscinosis, but is not a component of the storage material.

    Science.gov (United States)

    Xu, Su; Sleat, David E; Jadot, Michel; Lobel, Peter

    2010-05-27

    Classical late-infantile neuronal ceroid lipofuscinosis (LINCL) is a fatal neurodegenerative disease of children caused by mutations in TPP1, the gene encoding the lysosomal protease tripeptidyl peptidase 1. LINCL is characterized by lysosomal accumulation of storage material of which only a single protein component, subunit c of mitochondrial ATP synthase, has been well established to date. Identification of other protein constituents of the storage material could provide useful insights into the pathophysiology of disease and the natural substrates for TPP1. We have therefore initiated a proteomic analysis of storage material in brain from a LINCL mouse model. One protein, GFAP (glial fibrillary acidic protein), was found to be elevated in the LINCL mice compared with normal controls in both isolated storage bodies and a lysosome-enriched subcellular fraction that contains storage material. To determine whether GFAP accumulates within the lysosome in LINCL, we examined its intracellular distribution using subcellular fractionation and morphological methods. These experiments demonstrate that GFAP is not a component of the storage material in LINCL, suggesting that reports of GFAP storage in other NCLs may need to be re-examined. A number of other proteins were elevated in the storage material and/or lysosome-enriched fraction from the LINCL mice, but it remains unclear whether these proteins are true constituents of the storage material or, like GFAP, whether they associate with this material upon purification.

  5. Human influenza viral infection in utero alters glial fibrillary acidic protein immunoreactivity in the developing brains of neonatal mice.

    Science.gov (United States)

    Fatemi, S H; Emamian, E S; Sidwell, R W; Kist, D A; Stary, J M; Earle, J A; Thuras, P

    2002-01-01

    Epidemiological reports describe a strong association between prenatal human influenza viral infection and later development of schizophrenia. Postmodern human brain studies, however, indicate a lack of gliosis in schizophrenic brains presumably secondary to absence of glial cells during the second trimester viral infection in utero. We hypothesized that human influenza infection in day 9 pregnant mice would alter the expression of glial fibrillary acidic protein (GFAP, an important marker of gliosis, neuron migration, and reactive injury) in developing brains of postnatal days 0, 14 and 35 mice. Determination of cellular GFAP immunoreactivity (IR) expressed as cell density in cortex and hippocampus of control and experimental brains showed increases in GFAP-positive density in exposed cortical (P = 0.03 day 14 vs control) and hippocampal cells (P = 0.035 day 14, P = 0.034 day 35). Similarly, ependymal cell layer GFAP-IR cell counts showed increases with increasing brain age from day 0, to days 14 and 35 in infected groups (P = 0.037, day 14) vs controls. The GFAP-positive cells in prenatally exposed brains showed 'hypertrophy' and more stellate morphology. These results implicate a significant role of prenatal human influenza viral infection on subsequent gliosis, which persists throughout brain development in mice from birth to adolescence.

  6. Glial Fibrillary Acidic Protein is not an early marker of injury in perinatal asphyxia and hypoxic ischaemic encephalopathy

    Directory of Open Access Journals (Sweden)

    Ann-Marie eLooney

    2015-12-01

    Full Text Available Brain specific glial fibrillary acidic protein (GFAP has been suggested as a potential biomarker for hypoxic ischaemic encephalopathy (HIE in newborns (1, 2. Previous studies have shown increased levels in postnatal blood samples. However its ability to guide therapeutic intervention in HIE is unknown. Therapeutic hypothermia for HIE must be initiated within 6 hours of birth, therefore a clinically useful marker of injury would have to be available immediately following delivery. The goal of our study was to examine the ability of GFAP to predict grade of encephalopathy and neurological outcome when measured in umbilical cord blood. Infants with suspected perinatal asphyxia (PA and HIE were enrolled in a single, tertiary maternity hospital, where umbilical cord blood (UCB was drawn, processed and bio-banked at birth. Expression levels of GFAP were measured by ELISA. In total 169 infants (83 controls, 56 PA, 30 HIE were included in the study. GFAP levels were not increased in UCB of case infants (PA/HIE when compared to healthy controls or when divided into specific grades of HIE. Additionally, no correlation was found between UCB levels of GFAP and outcome at 36 months.

  7. GFAP isoforms control intermediate filament network dynamics, cell morphology, and focal adhesions.

    Science.gov (United States)

    Moeton, Martina; Stassen, Oscar M J A; Sluijs, Jacqueline A; van der Meer, Vincent W N; Kluivers, Liselot J; van Hoorn, Hedde; Schmidt, Thomas; Reits, Eric A J; van Strien, Miriam E; Hol, Elly M

    2016-11-01

    Glial fibrillary acidic protein (GFAP) is the characteristic intermediate filament (IF) protein in astrocytes. Expression of its main isoforms, GFAPα and GFAPδ, varies in astrocytes and astrocytoma implying a potential regulatory role in astrocyte physiology and pathology. An IF-network is a dynamic structure and has been functionally linked to cell motility, proliferation, and morphology. There is a constant exchange of IF-proteins with the network. To study differences in the dynamic properties of GFAPα and GFAPδ, we performed fluorescence recovery after photobleaching experiments on astrocytoma cells with fluorescently tagged GFAPs. Here, we show for the first time that the exchange of GFP-GFAPδ was significantly slower than the exchange of GFP-GFAPα with the IF-network. Furthermore, a collapsed IF-network, induced by GFAPδ expression, led to a further decrease in fluorescence recovery of both GFP-GFAPα and GFP-GFAPδ. This altered IF-network also changed cell morphology and the focal adhesion size, but did not alter cell migration or proliferation. Our study provides further insight into the modulation of the dynamic properties and functional consequences of the IF-network composition.

  8. GFAP expression and social deficits in transgenic mice overexpressing human sAPPα

    Science.gov (United States)

    Bailey, Antoinette R; Hou, Huayan; Song, Min; Obregon, Demian F; Portis, Samantha; Barger, Steven; Shytle, Doug; Stock, Saundra; Mori, Takashi; Sanberg, Paul G; Murphy, Tanya; Tan, Jun

    2013-01-01

    Autistic individuals display impaired social interactions and language, and restricted, stereotyped behaviors. Elevated levels of secreted amyloid precursor protein-alpha (sAPPα), the product of α-secretase cleavage of APP, are found in the plasma of some individuals with autism. The sAPPα protein is neurotrophic and neuroprotective and recently showed a correlation to glial differentiation in human neural stem cells (NSCs) via the IL-6 pathway. Considering evidence of gliosis in postmortem autistic brains, we hypothesized that subsets of patients with autism would exhibit elevations in CNS sAPPα and mice generated to mimic this observation would display markers suggestive of gliosis and autism-like behavior. Elevations in sAPPα levels were observed in brains of autistic patients compared to controls. Transgenic mice engineered to overexpress human sAPPα (TgsAPPα mice) displayed hypoactivity, impaired sociability, increased brain glial fibrillary acidic protein (GFAP) expression, and altered Notch1 and IL-6 levels. NSCs isolated from TgsAPPα mice, and those derived from wild-type mice treated with sAPPα, displayed suppressed β-tubulin III and elevated GFAP expression. These results suggest that elevations in brain sAPPα levels are observed in subsets of individuals with autism and TgsAPPα mice display signs suggestive of gliosis and behavioral impairment. PMID:23840007

  9. Assessment of Serum UCH-L1 and GFAP in Acute Stroke Patients

    Science.gov (United States)

    Ren, Changhong; Kobeissy, Firas; Alawieh, Ali; Li, Na; Li, Ning; Zibara, Kazem; Zoltewicz, Susie; Guingab-Cagmat, Joy; Larner, Stephen F.; Ding, Yuchuan; Hayes, Ronald L.; Ji, Xunming; Mondello, Stefania

    2016-01-01

    A rapid and reliable diagnostic test to distinguish ischemic from hemorrhagic stroke in patients presenting with stroke-like symptoms is essential to optimize management and triage for thrombolytic therapy. The present study measured serum concentrations of ubiquitin C-terminal hydrolase (UCH-L1) and glial fibrillary astrocytic protein (GFAP) in acute stroke patients and healthy controls and investigated their relation to stroke severity and patient characteristics. We also assessed the diagnostic performance of these markers for the differentiation of intracerebral hemorrhage (ICH) from ischemic stroke (IS). Both UCH-L1 and GFAP concentrations were significantly greater in ICH patients than in controls (p < 0.0001). However, exclusively GFAP differed in ICH compared with IS (p < 0.0001). GFAP yielded an AUC of 0.86 for differentiating between ICH and IS within 4.5hrs of symptom onset with a sensitivity of 61% and a specificity of 96% using a cut-off of 0.34ng/ml. Higher GFAP levels were associated with stroke severity and history of prior stroke. Our results demonstrate that blood UCH-L1 and GFAP are increased early after stroke and distinct biomarker-specific release profiles are associated with stroke characteristics and type. We also confirmed the potential of GFAP as a tool for early rule-in of ICH, while UCH-L1 was not clinically useful. PMID:27074724

  10. GFAP-BDP as an acute diagnostic marker in traumatic brain injury: results from the prospective transforming research and clinical knowledge in traumatic brain injury study.

    Science.gov (United States)

    Okonkwo, David O; Yue, John K; Puccio, Ava M; Panczykowski, David M; Inoue, Tomoo; McMahon, Paul J; Sorani, Marco D; Yuh, Esther L; Lingsma, Hester F; Maas, Andrew I R; Valadka, Alex B; Manley, Geoffrey T

    2013-09-01

    Reliable diagnosis of traumatic brain injury (TBI) is a major public health need. Glial fibrillary acidic protein (GFAP) is expressed in the central nervous system, and breakdown products (GFAP-BDP) are released following parenchymal brain injury. Here, we evaluate the diagnostic accuracy of elevated levels of plasma GFAP-BDP in TBI. Participants were identified as part of the prospective Transforming Research And Clinical Knowledge in Traumatic Brain Injury (TRACK-TBI) Study. Acute plasma samples (<24 h post-injury) were collected from patients presenting with brain injury who had CT imaging. The ability of GFAP-BDP level to discriminate patients with demonstrable traumatic lesions on CT, and with failure to return to pre-injury baseline at 6 months, was evaluated by the area under the receiver operating characteristic curve (AUC). Of the 215 patients included for analysis, 83% had mild, 4% had moderate, and 13% had severe TBI; 54% had acute traumatic lesions on CT. The ability of GFAP-BDP level to discriminate patients with traumatic lesions on CT as evaluated by AUC was 0.88 (95% confidence interval [CI], 0.84-0.93). The optimal cutoff of 0.68 ng/mL for plasma GFAP-BDP level was associated with a 21.61 odds ratio for traumatic findings on head CT. Discriminatory ability of unfavorable 6 month outcome was lower, AUC 0.65 (95% CI, 0.55-0.74), with a 2.07 odds ratio. GFAP-BDP levels reliably distinguish the presence and severity of CT scan findings in TBI patients. Although these findings confirm and extend prior studies, a larger prospective trial is still needed to validate the use of GFAP-BDP as a routine diagnostic biomarker for patient care and clinical research. The term "mild" continues to be a misnomer for this patient population, and underscores the need for evolving classification strategies for TBI targeted therapy. (ClinicalTrials.gov number NCT01565551; NIH Grant 1RC2 NS069409).

  11. Role of Sigma Receptor in Cocaine-Mediated Induction of Glial Fibrillary Acidic Protein: Implications for HAND.

    Science.gov (United States)

    Yang, Lu; Yao, Honghong; Chen, Xufeng; Cai, Yu; Callen, Shannon; Buch, Shilpa

    2016-03-01

    Cocaine abuse has been shown to accelerate the progression of human immunodeficiency virus (HIV)-1-associated neurological disorders (HANDs) partially through increasing neuroinflammatory response mediated by activated astrocytes; however, the detailed molecular mechanism of cocaine-mediated astrocyte activation is unclear. In the current study, we demonstrated increased astrogliosis in the cortical regions of brains from HIV(+) cocaine abusers compared with the HIV(+) group without cocaine abuse. We next sought to explore whether cocaine exposure could result in increased expression of glial fibrillary acidic protein (GFAP), a filament protein critical for astrocyte activation. Exposure of cocaine to astrocytes resulted in rapid translocation of sigma receptor to the plasma membrane with subsequent activation of downstream signaling pathways. Using a pharmacological approach, we provide evidence that cocaine-mediated upregulation of GFAP expression involved activation of mitogen-activated protein kinase (MAPK) signaling with subsequent downstream activation of the early growth response gene 1 (Egr-1). Egr-1 activation, in turn, caused transcriptional regulation of GFAP. Corroboration of these findings in vivo demonstrated increased expression of GFAP in the cortical region of mice treated with cocaine compared with the saline injected controls. A thorough understanding of how cocaine mediates astrogliosis could have implications for the development of therapeutic interventions aimed at HIV-infected cocaine abusers.

  12. Prenatal X-irradiation increases GFAP- and calbindin D28k-immunoreactivity in the medial subdivision of the nucleus of solitary tract in the rat.

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    Jacquin, T D; Xie, Q; Miki, T; Satriotomo, I; Itoh, M; Takeuchi, Y

    2000-04-12

    Glial fibrillary acidic protein- (GFAP) and calbindin D28k-immunoreactivity (IR) were investigated in the medial subdivision of the nucleus of the solitary tract (mNST) of prenatally X-irradiated rats. Pregnant rats were exposed to a single whole-body X-irradiation on day 11 or 16 of gestation at a dose of 1. 3 Gy. The offspring were killed at 7-14 days of age for the immunohistochemical observations. Rat pups showed strong GFAP-IR at the level rostral to the obex when receiving X-rays on day 11 of gestation, with hypertrophy of astrocyte cell bodies and cytoplasmic processes, but weak GFAP-IR when receiving X-rays on day 16 of gestation. Calbindin D28k-IR was stronger in the animals receiving X-rays on day 11 or 16 of gestation compared to that in the control animals. In the present study, the increase of GFAP- and calbindin D28k-IR cells in the mNST might indicate that adaptative mechanisms are taking place to preserve integrated nervous system function and possibly, to provide neuroprotection.

  13. Effect of taurine on GFAP and TauT expressions in rat retinal Müller cells in high glucose culture

    Institute of Scientific and Technical Information of China (English)

    ZHANG Ya-jie; XU Hong-xia; ZENG Kai-hong; MI Man-tian

    2007-01-01

    Objective:To detect the expression of glial fibrillary acid protein (GFAP) and taurine transporter (TauT) in the retinal Müller cells in high glucose culture with taurine and to explore the influence of glucose on the taurine transporting, and the possible protective effects of taurine on Müller cells in early diabetic retinopathy. Methods: The Müller cells from the rat retina were cultured in high glucose, and GFAP and TauT expressions were detected in the cells treated with different doses of taurine by immuocytochemical fluorescein staining and Western blotting. Results: High glucose enhanced the expression of GFAP and decreased the expression of TauT in Müller cells. Taurine decreased the up-regulation of GFAP in the cells which was induced by high glucose; 0. 1-10 mmol/L taurine increased the expression of TauT in Müller cells. Conclusion: Taurine can inhibit the changes in Müller cell resulted from high glucose.

  14. Activation of a pro-survival pathway IL-6/JAK2/STAT3 contributes to glial fibrillary acidic protein induction during the cholera toxin-induced differentiation of C6 malignant glioma cells.

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    Shu, Minfeng; Zhou, Yuxi; Zhu, Wenbo; Wu, Sihan; Zheng, Xiaoke; Yan, Guangmei

    2011-06-01

    Differentiation-inducing therapy has been proposed to be a novel potential approach to treat malignant gliomas. Glial fibrillary acidic protein (GFAP) is a well-known specific astrocyte biomarker and acts as a tumor suppressor gene (TSG) in glioma pathogenesis. Previously we reported that a traditional biotoxin cholera toxin could induce malignant glioma cell differentiation characterized by morphologic changes and dramatic GFAP expression. However, the molecular mechanisms underlying GFAP induction are still largely unknown. Here we demonstrate that an oncogenic pathway interleukin-6/janus kinase-2/signal transducer and activator of transcription 3 (IL-6/JAK2/STAT3) cascade mediates the cholera toxin-induced GFAP expression. Cholera toxin dramatically stimulated GFAP expression at the transcriptional level in C6 glioma cells. Meanwhile, phosphorylation of STAT3 and JAK2 was highly induced in a time-dependent manner after cholera toxin incubation, whereas no changes of STAT3 and JAK2 were observed. Furthermore, the IL-6 gene was quickly induced by cholera toxin and subsequent IL-6 protein secretion was stimulated. Importantly, exogenous recombinant rat IL-6 can also induce phosphorylation of STAT3 concomitant with GFAP expression while JAK2 specific inhibitor AG490 could effectively block both cholera toxin- and IL-6-induced GFAP expression. Given that the methylation of the STAT3 binding element can suppress GFAP expression, we detected the methylation status of the critical recognition sequence of STAT3 in the promoter of GFAP gene (-1518 ∼ -1510) and found that it was unmethylated in C6 glioma cells. In addition, neither DNA methyltransferase1 (DNMT1) inhibitor 5-Aza-2'-deoxycytidine (5-AZa-CdR) nor silencing DNMT1 can stimulate GFAP expression, indicating that the loss of GFAP expression in C6 cells is not caused by its promoter hypermethylation. Taken together, our findings suggest that activation of a pro-survival IL-6/JAK2/STAT3 cascade contributes to

  15. Protein and amino acid nutrition

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    Dairy cow protein and amino acid nutrition have a significant role in sustainable dairying. Protein, amino acids, and nitrogen are inextricably linked through effects in the rumen, metabolism of the cow, and environmental nutrient management. Feeding systems have been making progress toward emphasiz...

  16. Immunohistochemical expression of neural tissue markers (neuron-specific enolase, glial fibrillary acidic protein, S100 protein) in ameloblastic fibrodentinoma: a comparative study with ameloblastic fibroma.

    Science.gov (United States)

    Takeda, Y; Sato, H; Satoh, M; Nakamura, S I; Yamamoto, H

    2000-08-01

    Formalin-fixed paraffin-embedded sections of three cases of ameloblastic fibrodentinoma (AFD) were studied by the avidin-biotin-peroxidase complex method using antibodies against neuron-specific enolase (NSE), glial fibrillary acidic protein (GFAP) and S100 protein and the results were compared with those in ameloblastic fibroma (AF). A striking histopathological characteristic of AFD was the formation of abortive dentin with various degrees of maturation at the epithelial-mesenchymal tissue interface. Central cells of enamel organ-like epithelia with various stages of abortive dentin induction in AFD were generally positive for NSE. Dental lamina-like epithelial cells also showed positive staining in some areas. No cells were positive for NSE in AF. Positive staining for GFAP was observed in the juxta-epithelial mesenchymal tissue of the formation stage of immature dentin with various numbers of entrapped cells in AFD, but GFAP staining was negative in other mesenchymal and epithelial tissues at other stages. In AF, no GFAP-positive cells were found. There were a few S100 protein-positive cells found in the foci of epithelial components in both AFD and AF. Mesenchymal cells showing a dendritic or spindle shape were positive for S100 protein in some areas of AFD and AF. Although such cells in the mesenchymal component of pigmented AFD were more numerous than in non-pigmented AFD and AF, their distribution pattern in the former condition was basically similar to that in the latter. Although the present results, obtained from conventional immunohistochemical procedures, do not directly reflect the expression of neural crest-derived cells in the dentinogenesis of AFD, such results do not disprove the possibility of the expression of neural proteins probably related to neural crest-derived cells in dentinogenesis under certain pathologic conditions in odontogenic mixed tumors. Such a phenomenon may also occur during dentinogenesis in other odontogenic mixed tumors and

  17. [The expression of GFAP after brain concussion in rats].

    Science.gov (United States)

    Zhang, Chun-Bing; Li, Yong-Hong

    2006-04-01

    To study the expression of GFAP and pathologic changes after rats brain concussion, so that to provide evidence on brain concussion for forensic identification. Forty-five SD rats were divided into 3, 6, 12, 24 h and 2, 4, 7, 10 d and normal control groups in terms of different wounding time after brain concussion model established, and the expression of GFAP after rats brain concussion were then observed by using SP immunohistochemical method. In normal control brain, low-level GFAP expressions could be observed. After six hours' brain concussion, GFAP positive cells increased obviously. The trend reached to the peak at 7d, partly declined at 10d, then decreased gradually. Brain concussion induced the expression of GFAP. The detection of GFAP could be useful for diagnosis of brain concussion on forensic pathology, and could be a reference index for timing of injury after brain concussion.

  18. The use of serum glial fibrillary acidic protein measurements in the diagnosis of neuromyelitis optica spectrum optic neuritis.

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    Mithu Storoni

    Full Text Available BACKGROUND: Glial fibrillary acidic protein (GFAP is a specific intermediate filament of the cytoskeleton of the astrocyte and may be used as a specific marker for astrocytic damage. It is detectable in the cerebrospinal fluid following a relapse caused by Multiple Sclerosis (MS and Neuromyelitis Optica (NMO spectrum disease. Higher levels are found following an NMO-related relapse. It is not known if GFAP is also detectable in the serum following such relapses. In particular, it is not known if lesions limited to the optic nerve release GFAP in sufficient quantities to be detectable within the serum. The aim of this study was to ascertain the extent to which serum GFAP levels can distinguish between an episode of optic neuritis (ON related to NMO spectrum disease and ON from other causes. METHODOLOGY/PRINCIPAL FINDINGS: Out of 150 patients consecutively presenting to our eye hospital over the period March 2009 until July 2010, we were able to collect a serum sample from 12 patients who had presented with MS-related ON and from 10 patients who had presented with NMO spectrum disease-related ON. We also identified 8 patients with recurrent isolated ON and 8 patients with a corticosteroid-dependent optic neuropathy in the absence of any identified aetiology. GFAP was detectable in the serum of all but three patients (two patients with MS-related ON and one with recurrent optic neuritis. The median serum GFAP level in the patient group with NMO spectrum disease was 4.63 pg/mL whereas in all other cases combined together, this was 2.14 pg/mL. The difference was statistically significant (P = 0.01. A similar statistically significant difference was found when cases with pathology limited to the optic nerve were compared (P = 0.03. CONCLUSIONS: Glial pathology in NMO related optic neuritis is reflected in elevated serum GFAP levels independently of whether or not there is extra-optic nerve disease.

  19. Immunohistochemical analysis of brain lesions using S100B and glial fibrillary acidic protein antibodies in arundic acid- (ONO-2506) treated stroke-prone spontaneously hypertensive rats.

    Science.gov (United States)

    Higashino, Hideaki; Niwa, Atsuko; Satou, Takao; Ohta, Yoshio; Hashimoto, Shigeo; Tabuchi, Masaki; Ooshima, Kana

    2009-10-01

    Stroke-prone spontaneously hypertensive rats (SHRSP) used as a model of essential hypertension cause a high incidence of brain stroke on the course of hypertension. Incidences and sizes of brain lesions are known to relate to the astrocyte activities. Therefore, relation between brain damage and the expression profile of the astrocytes was investigated with morphometric and immunohistochemical analyses using astrocyte marker antibodies of S100B and glial fibrillary acidic protein (GFAP) with or without arundic acid administration, a suppressor on the activation of astrocytes. Arundic acid extended the average life span of SHRSP. An increase in brain tissue weight was inhibited concomitant with a lower rate of gliosis/hemosiderin deposit/scarring in brain lesions. S100B- or GFAP-positive dot and filamentous structures were decreased in arundic acid-treated SHRSP, and this effect was most pronounced in the cerebral cortex, white matter, and pons, and less so in the hippocampus, diencephalon, midbrain, and cerebellum. Blood pressure decreased after administration of arundic acid in the high-dose group (100 mg/kg/day arundic acid), but not in the low-dose group (30 mg/kg/day). These data indicate that arundic acid can prevent hypertension-induced stroke, and may inhibit the enlargement of the stroke lesion by preventing the inflammatory changes caused by overproduction of the S100B protein in the astrocytes.

  20. Specific human astrocyte subtype revealed by affinity purified GFAP antibody; unpurified serum cross-reacts with neurofilament-L in Alzheimer.

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    Jinte Middeldorp

    Full Text Available The human GFAP splice variants GFAPDelta164 and GFAPDeltaexon6 both result in a GFAP protein isoform with a unique out-of-frame carboxy-terminus that can be detected by the GFAP+1 antibody. We previously reported that GFAP+1 was expressed in astrocytes and in degenerating neurons in Alzheimer's disease brains. In this study we aimed at further investigating the neuronal GFAP+1 expression and we started by affinity purifying the GFAP+1 antibody. The purified antibody resulted in a loss of neuronal GFAP+1 signal, although other antibodies directed against the amino- and carboxy-terminus of GFAPalpha still revealed GFAP-immunopositive neurons, as described before. With an in-depth analysis of a western blot, followed by mass spectrometry we discovered that the previously detected neuronal GFAP+1 expression was due to cross-reactivity of the antibody with neurofilament-L (NF-L. This was confirmed by double-label fluorescent immunohistochemistry and western blotting with the unpurified GFAP+1 antibody and an antibody against NF-L. Our data imply that NF-L can accumulate in some tangle-like structures in Alzheimer brains. More importantly, the purified GFAP+1 antibody clearly revealed a specific subtype of astrocytes in the adult human brain. These large astrocytes are present throughout the brain, e.g., along the subventricular zone, in the hippocampus, in the striatum and in the spinal cord of controls, Alzheimer, and Parkinson patients. The presence of a specific GFAP-isoform suggests a specialized function of these astrocytes.

  1. Specific Human Astrocyte Subtype Revealed by Affinity Purified GFAP+1 Antibody; Unpurified Serum Cross-Reacts with Neurofilament-L in Alzheimer

    Science.gov (United States)

    Middeldorp, Jinte; van den Berge, Simone A.; Aronica, Eleonora; Speijer, Dave; Hol, Elly M.

    2009-01-01

    The human GFAP splice variants GFAPΔ164 and GFAPΔexon6 both result in a GFAP protein isoform with a unique out-of-frame carboxy-terminus that can be detected by the GFAP+1 antibody. We previously reported that GFAP+1 was expressed in astrocytes and in degenerating neurons in Alzheimer's disease brains. In this study we aimed at further investigating the neuronal GFAP+1 expression and we started by affinity purifying the GFAP+1 antibody. The purified antibody resulted in a loss of neuronal GFAP+1 signal, although other antibodies directed against the amino- and carboxy-terminus of GFAPα still revealed GFAP-immunopositive neurons, as described before. With an in-depth analysis of a western blot, followed by mass spectrometry we discovered that the previously detected neuronal GFAP+1 expression was due to cross-reactivity of the antibody with neurofilament-L (NF-L). This was confirmed by double-label fluorescent immunohistochemistry and western blotting with the unpurified GFAP+1 antibody and an antibody against NF-L. Our data imply that NF-L can accumulate in some tangle-like structures in Alzheimer brains. More importantly, the purified GFAP+1 antibody clearly revealed a specific subtype of astrocytes in the adult human brain. These large astrocytes are present throughout the brain, e.g., along the subventricular zone, in the hippocampus, in the striatum and in the spinal cord of controls, Alzheimer, and Parkinson patients. The presence of a specific GFAP-isoform suggests a specialized function of these astrocytes. PMID:19888461

  2. Estudo da imunorreatividade astrocitária para GFAP e vimentina no tronco encefálico de ratos Wistar submetidos ao modelo gliotóxico do brometo de etídio Investigation into the astrocytic immunoreactivity to GFAP and vimentin in the brainstem of Wistar rats submitted to the ethidium bromide gliotoxic model

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    Eduardo Fernandes Bondan

    2003-09-01

    Full Text Available INTRODUÇÃO: O brometo de etídio (BE é reconhecido como um agente gliotóxico que causa desaparecimento focal astrocitário e oligodendroglial. OBJETIVO: Investigou-se a imunorreatividade astrocitária à proteína glial fibrilar ácida (GFAP e à vimentina (VIM após injeção do BE. MÉTODO: Ratos Wistar adultos foram tomados como controles histológicos (grupo H ou injetados na cisterna basal com BE a 0,1% (grupo E ou salina a 0,9% (grupo C. Fragmentos do tronco encefálico foram colhidos das 24 horas aos 31 dias pós-injeção para estudo imuno-histoquímico da GFAP e VIM pelo método da avidina-biotina. RESULTADOS: No grupo E, foram observadas extensas lesões na ponte e no mesencéfalo, com desaparecimento astrocitário da área central 24 horas pós-BE, bem como infiltração macrofágica e astrogliose periférica a partir do 3º dia. Os astrócitos marginais apresentaram imunorreatividade aumentada à GFAP e reexpressão de VIM, esta confinada às bordas imediatas do sítio lesional. No grupo C, foram visualizadas lesões pontinas discretas, com preservação astrocitária central e marcação menos intensa para GFAP nos bordos em relação ao grupo E. Nenhuma imunorreatividade para VIM foi notada em tais astrócitos. CONCLUSÃO: Os astrócitos das margens das lesões induzidas pelo BE apresentaram imunorreatividade aumentada para GFAP e reexpressão de VIM.BACKGROUND: Ethidium bromide (EB is known as a gliotoxic agent that causes focal astrocytic and oligodendroglial disappearance. OBJECTIVE: Astrocyte immunoreactivity to glial fibrillary acidic protein (GFAP and vimentin (VIM was investigated after EB injection. METHOD: Adult male Wistar rats were taken as histologic controls (group H or injected into cisterna pontis with 0.1% EB (group E or 0.9% saline solution (group C. Brainstem samples were collected from 24 hours to 31 days post-injection for GFAP and VIM immunohistochemical staining using avidin-biotin method. RESULTS: In

  3. Effects of Chloroquine on GFAP, PCNA and Cyclin D1 in Hippocampus and Cerebral Cortex of Rats with Seizures Induced by Pentylenetetrazole

    Institute of Scientific and Technical Information of China (English)

    ZHANG Shuhua; ZHU Changgeng; LIU Qingying; WANG Wei

    2005-01-01

    The effects of chloroquine on glial fibrillary acidic protein (GFAP), proliferation cell nuclear antigen (PCNA) and Cyclin D1 in hippocampus and cerebral cortex of rats with seizures induced by pentylenetetrazole (PTZ) were observed in the present study. Forty-eight male adult Sprague-Dawley (SD) rats were randomly divided into control group, chloroquine intervening group, and PTZ group. The behavior and electroencephalogram (EEG) were observed and recor ded. GFAP and PCNA were examined with immunohistochemistry. The content of Cyclin D1 in hippocampus and cerebral cortex was inspected with Western blot. The results showed no seizure activity in the control group, severe seizure activity in the PTZ group (Ⅳ-Ⅴ degree), and slight seizure activity ( Ⅰ - Ⅲ degree) in the chloroquine intervening group (P<0. 05). EEG recordings showed no epileptic spikes in the control group, high amplitude with fast frequency in the PTZ group, low-amplitude and slow frequency in the chloroquine intervening group. The expression of GFAP and the positive index of PCNA in the PTZ group were higher than those of control group (P <0.05 and P<0.01, respectively). No differences in GFAP expression and PCNA index were observed between chloroquine intervening and control groups (P>0.05). The content of Cyclin D1 in hippocampus and cerebral cortex was significantly higher in the PTZ group than in control and chloroquine intervening groups (P< 0.05). Therefore, it is considered that chloroquine, by inhibiting the functions and proliferation of glial cells in the hippocampus and cerebral cortex, can alleviate the seizure activities. These results suggest that chloroquine may be an ideal anticonvulsant in preventing and treating epilepsy.

  4. CUTANEOUS LUPUS ERYTHEMATOSUS WITH AUTOANTIBODIES COLOCALIZING WITH GLIAL FIBRILLARY ACIDIC PROTEIN

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    Abreu-Velez Ana Maria

    2011-01-01

    Full Text Available Background: Discoid lupus erythematosus (DLE is a chronic, inflammatory skin disorder, presenting with scarring lesions predominating on sun exposed areas of the face and scalp. Case Report: A 43-year-old African American female was evaluated for possible DLE. Methods: Skin biopsies for hematoxylin and eosin (H&E examination, as well as for direct immunofluorescence (DIF analysis were performed. Results: H&E staining demonstrated classic features of cutaneous lupus erythematosus, with the pertinent presence of perineural lymphohistiocitic infiltrates, especially those associated with skin appendices. DIF revealed strong deposits of immunoglobulins IgG, IgM, fibrinogen and Complement/C3, present in a shaggy, linear pattern at or near the basement membrane zone (BMZ of selected eccrine and sebaceous glands, and around some blood vessels. The BMZ positivity in these structures consistently colocalized with positive staining in multiple, punctate areas for glial fibrillary acidic protein (GFAP, including within cytoid bodies. Conclusions:. The observed colocalization of the patient’s autoantibodies in cutaneous lupus with GFAP may have pathophysiologic relevance. Specifically, our data could be consistent with previously described DLE patients with or without overt central nervous system manifestations, or could represent an epiphenomenon. Additional, larger studies are needed to satisfactorily address this possibility.

  5. Generation of GFAP::GFP astrocyte reporter lines from human adult fibroblast-derived iPS cells using zinc-finger nuclease technology.

    Science.gov (United States)

    Zhang, Ping-Wu; Haidet-Phillips, Amanda M; Pham, Jacqueline T; Lee, Youngjin; Huo, Yuqing; Tienari, Pentti J; Maragakis, Nicholas J; Sattler, Rita; Rothstein, Jeffrey D

    2016-01-01

    Astrocytes are instrumental to major brain functions, including metabolic support, extracellular ion regulation, the shaping of excitatory signaling events and maintenance of synaptic glutamate homeostasis. Astrocyte dysfunction contributes to numerous developmental, psychiatric and neurodegenerative disorders. The generation of adult human fibroblast-derived induced pluripotent stem cells (iPSCs) has provided novel opportunities to study mechanisms of astrocyte dysfunction in human-derived cells. To overcome the difficulties of cell type heterogeneity during the differentiation process from iPSCs to astroglial cells (iPS astrocytes), we generated homogenous populations of iPS astrocytes using zinc-finger nuclease (ZFN) technology. Enhanced green fluorescent protein (eGFP) driven by the astrocyte-specific glial fibrillary acidic protein (GFAP) promoter was inserted into the safe harbor adeno-associated virus integration site 1 (AAVS1) locus in disease and control-derived iPSCs. Astrocyte populations were enriched using Fluorescence Activated Cell Sorting (FACS) and after enrichment more than 99% of iPS astrocytes expressed mature astrocyte markers including GFAP, S100β, NFIA and ALDH1L1. In addition, mature pure GFP-iPS astrocytes exhibited a well-described functional astrocytic activity in vitro characterized by neuron-dependent regulation of glutamate transporters to regulate extracellular glutamate concentrations. Engraftment of GFP-iPS astrocytes into rat spinal cord grey matter confirmed in vivo cell survival and continued astrocytic maturation. In conclusion, the generation of GFAP::GFP-iPS astrocytes provides a powerful in vitro and in vivo tool for studying astrocyte biology and astrocyte-driven disease pathogenesis and therapy.

  6. Altered astrocytic swelling in the cortex of α-syntrophin-negative GFAP/EGFP mice.

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    Miroslava Anderova

    Full Text Available Brain edema accompanying ischemic or traumatic brain injuries, originates from a disruption of ionic/neurotransmitter homeostasis that leads to accumulation of K(+ and glutamate in the extracellular space. Their increased uptake, predominantly provided by astrocytes, is associated with water influx via aquaporin-4 (AQP4. As the removal of perivascular AQP4 via the deletion of α-syntrophin was shown to delay edema formation and K(+ clearance, we aimed to elucidate the impact of α-syntrophin knockout on volume changes in individual astrocytes in situ evoked by pathological stimuli using three dimensional confocal morphometry and changes in the extracellular space volume fraction (α in situ and in vivo in the mouse cortex employing the real-time iontophoretic method. RT-qPCR profiling was used to reveal possible differences in the expression of ion channels/transporters that participate in maintaining ionic/neurotransmitter homeostasis. To visualize individual astrocytes in mice lacking α-syntrophin we crossbred GFAP/EGFP mice, in which the astrocytes are labeled by the enhanced green fluorescent protein under the human glial fibrillary acidic protein promoter, with α-syntrophin knockout mice. Three-dimensional confocal morphometry revealed that α-syntrophin deletion results in significantly smaller astrocyte swelling when induced by severe hypoosmotic stress, oxygen glucose deprivation (OGD or 50 mM K(+. As for the mild stimuli, such as mild hypoosmotic or hyperosmotic stress or 10 mM K(+, α-syntrophin deletion had no effect on astrocyte swelling. Similarly, evaluation of relative α changes showed a significantly smaller decrease in α-syntrophin knockout mice only during severe pathological conditions, but not during mild stimuli. In summary, the deletion of α-syntrophin markedly alters astrocyte swelling during severe hypoosmotic stress, OGD or high K(+.

  7. GABAρ subunits confer a bicuculline-insensitive component to GFAP+ cells of cerebellum

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    Pétriz, Adriana; Reyes-Haro, Daniel; González-González, María Alejandra; Miledi, Ricardo; Martínez-Torres, Ataúlfo

    2014-01-01

    GABA-A receptors mediating synaptic or extrasynaptic transmission are molecularly and functionally distinct, and glial cells are known to express a plethora of GABA-A subunits. Here we demonstrate that GFAP+ cells of the granular layer of cerebellum express GABAρ subunits during early postnatal development, thereby conferring peculiar pharmacologic characteristics to GABA responses. Electron microscopy revealed the presence of GABAρ in the plasma membrane of GFAP+ cells. In contrast, expression in the adult was restricted to Purkinje neurons and a subset of ependymal cells. Electrophysiological studies in vitro revealed that astrocytes express functional receptors with an EC50 of 52.2 ± 11.8 μM for GABA. The evoked currents were inhibited by bicuculline (100 μM) and TPMPA (IC50, 5.9 ± 0.6 μM), indicating the presence of a GABAρ component. Coimmunoprecipitation demonstrated protein–protein interactions between GABAρ1 and GABAα1, and double immunofluorescence showed that these subunits colocalize in the plasma membrane. Three populations of GABA-A receptors in astrocytes were identified: classic GABA-A, bicuculline-insensitive GABAρ, and GABA-A–GABAρ hybrids. Clusters of GABA-A receptors were distributed in the perinuclear space and along the processes of GFAP+ cells. Time-lapse microscopy showed GABAρ2-GFP accumulation in clusters located in the soma and along the processes. The clusters were relatively immobile, with mean displacement of 9.4 ± 0.9 μm and a net distance traveled of 1–2 μm, owing mainly to directional movement or simple diffusion. Modulation of GABAρ dynamics may be a novel mechanism of extrasynaptic transmission regulating GABAergic control of GFAP+ cells during early postnatal development. PMID:25422464

  8. Dopamine D1 Receptor Immunoreactivity on Fine Processes of GFAP-Positive Astrocytes in the Substantia Nigra Pars Reticulata of Adult Mouse

    Science.gov (United States)

    Nagatomo, Katsuhiro; Suga, Sechiko; Saitoh, Masato; Kogawa, Masahito; Kobayashi, Kazuto; Yamamoto, Yoshio; Yamada, Katsuya

    2017-01-01

    Substantia nigra pars reticulata (SNr), the major output nucleus of the basal ganglia, receives dopamine from dendrites extending from dopaminergic neurons of the adjacent nucleus pars compacta (SNc), which is known for its selective degeneration in Parkinson's disease. As a recipient for dendritically released dopamine, the dopamine D1 receptor (D1R) is a primary candidate due to its very dense immunoreactivity in the SNr. However, the precise location of D1R remains unclear at the cellular level in the SNr except for that reported on axons/axon terminals of presumably striatal GABAergic neurons. To address this, we used D1R promotor-controlled, mVenus-expressing transgenic mice. When cells were acutely dissociated from SNr of mouse brain, prominent mVenus fluorescence was detected in fine processes of glia-like cells, but no such fluorescence was detected from neurons in the same preparation, except for the synaptic bouton-like structure on the neurons. Double immunolabeling of SNr cells dissociated from adult wild-type mice brain further revealed marked D1R immunoreactivity in the processes of glial fibrillary acidic protein (GFAP)-positive astrocytes. Such D1R imunoreactivity was significantly stronger in the SNr astrocytes than that in those of the visual cortex in the same preparation. Interestingly, GFAP-positive astrocytes dissociated from the striatum demonstrated D1R immunoreactivity, either remarkable or minimal, similarly to that shown in neurons in this nucleus. In contrast, in the SNr and visual cortex, only weak D1R immunoreactivity was detected in the neurons tested. These results suggest that the SNr astrocyte may be a candidate recipient for dendritically released dopamine. Further study is required to fully elucidate the physiological roles of divergent dopamine receptor immunoreactivity profiles in GFAP-positive astrocytes. PMID:28203148

  9. Astrocytes in development, aging and disease: starring GFAP

    NARCIS (Netherlands)

    Middeldorp, J.

    2010-01-01

    We show in this thesis that different subtypes of astrocytes comprise specialized GFAP-IF networks, that change during development, aging and Alzheimer’s disease. The novel functions that have emerged for the IF network suggest these changes can play an important part in the specialized function of

  10. Relationship between serum level of GFAP and cognitive function in patients with first episode schizophrenia.%首发精神分裂症患者血清GFAP与认知功能的研究

    Institute of Scientific and Technical Information of China (English)

    李国华; 李继荣; 吴秋霞; 熊鹏; 曾勇; 徐飞; 卢瑾; 姜林伶

    2013-01-01

    目的 研究首发精神分裂症患者血清中胶质纤维酸性蛋白(Glial Fibrillary Acidic Protein,GFAP)水平的变化与认知功能障碍之间的关系.方法 用酶联免疫吸附技术(Enzyme-linked Immunoadsordent Assay,ELISA)测定48例精神分裂症患者(患者组)和42例正常对照者(对照组)血清GFAP的浓度,并用威斯康星卡片分类测验(Wisconsin Cord Sorting Test,WCST)检测首发精神分裂症患者和正常人的认知功能.比较两组血清GFAP浓度,同时探讨GFAP浓度的变化与认知功能的关系.结果 (1)患者组血清浓度高于对照组(P<0.01);(2)患者组WCST成绩低于对照组(P<0.01),表明首发精神分裂症患者认知功能及执行能力比正常人差;(3)相关分析表明WCST中错误应答数、持续性错误数与患者血清GFAP浓度呈正相关,完成分类个数与患者血清GFAP浓度呈负相关(P<0.05).结论 首发精神分裂症患者存在认知功能障碍和神经胶质细胞损害,且代表神经胶质细胞损害的血清GFAP浓度与认知功能障碍程度呈正相关.%Objective To study the relevance between serum level of glial fibrillary acidic protein (GFAP) and cognitive dysfunction in patients with first episode schizophrenia. Methods Serum GFAP level was measured with Enzyme-linked Immunosorbent method and the cognitive function was assessed with Wisconsin cord sorting test in 48 patients with first episode schizophrenia ( study group) and 42 normal controls ( control group) . Serum GFAP level was compared between the two groups and the relationship between GFAP level and cognitive dysfunction was analyzed. Results (1) Serum GFAP level in study group was significantly higher than that in control group ( P <0. 01). (2) Score of WCST in study group was significantly lower than that in control group (P<0.01). (3)Serum GFAP level in study group was positively correlated with number of wrong answers and number of continued errors, and was negatively correlated

  11. Yohimbine prevents morphine-induced changes of glial fibrillary acidic protein in brainstem and alpha2-adrenoceptor gene expression in hippocampus.

    Science.gov (United States)

    Alonso, Elba; Garrido, Elisa; Díez-Fernández, Carmen; Pérez-García, Carmen; Herradón, Gonzalo; Ezquerra, Laura; Deuel, Thomas F; Alguacil, Luis F

    2007-01-29

    The alpha(2)-adrenoceptor antagonist yohimbine is known to oppose to several pharmacological effects of opioid drugs, but the consequences and the mechanisms involved remain to be clearly established. In the present study we have checked the effects of yohimbine on morphine-induced alterations of the expression of key proteins (glial fibrillary acidic protein, GFAP) and genes (alpha(2)-adrenoceptors) in rat brain areas known to be relevant in opioid dependence, addiction and individual vulnerability to drug abuse. Rats were treated with morphine in the presence or absence of yohimbine. The effects of the treatments on GFAP expression were studied by immunohistochemical staining in Locus Coeruleus (LC) and Nucleus of the Solitary Tract (NST), two important noradrenergic nuclei. In addition, drug effects on alpha(2)-adrenoceptor gene expression were determined by real time RT-PCR in the hippocampus, a brain area that receives noradrenergic input from the brainstem. Morphine administration increased GFAP expression both in LC and NST as it was previously reported in other brain areas. Yohimbine was found to efficiently prevent morphine-induced GFAP upregulation. Chronic (but not acute) morphine downregulated mRNA levels of alpha(2A)- and alpha(2C)-adrenoceptors in the hippocampus, while simultaneously increased the expression of the alpha(2B)-adrenoceptor gene. Again, yohimbine was able to prevent morphine-induced changes in the levels of expression of the three alpha(2)-adrenoceptor genes. These results correlate the well-established reduction of opioid dependence and addiction by yohimbine and suggest that this drug could interfere with the neural plasticity induced by chronic morphine in central noradrenergic pathways.

  12. Fetal development of the human epiglottis revisited: appearance of GFAP-positive mesenchymal cells and fibrous connections with other laryngeal and lingual structures.

    Science.gov (United States)

    Katori, Yukio; Takeuchi, Hiromi; Rodríguez-Vázquez, Jose Francisco; Kitano, Hiroya; Murakami, Gen; Kawase, Tetsuaki

    2011-03-01

    The fetal anatomy of the human epiglottis has not yet been fully described. We investigated the histology (paraffin-embedding) of 18 mid-term fetuses at 7-25 weeks of gestation (three fetuses each at 7, 9, 12, 15, 20 and 25 weeks). A mesenchymal condensation of the epiglottic cartilage appears posterior and somewhat superior to the hyoid body at 9 weeks, but at 12 and 15 weeks, the root or inferior part descends to the level of the thyroid cartilage. The covering epithelium stains much darker with hematoxylin than other pharyngeal epithelia. After 20 weeks, the epiglottis again protrudes superiorly beyond the hyoid body. In contrast to other laryngeal cartilage anlagen, the mesenchymal condensation of the epiglottis begins to express glial fibrillary acidic protein (GFAP) at 15 weeks. At the same stage, mucosal glands begin invading into the epiglottic mesenchyme. The developing cartilage becomes penetrated and fragmented by abundant mucosal glands up until 25 weeks. The thyro-epiglottic ligament seems to develop from the GFAP-positive mesenchymal condensation, whereas the hyo-epiglottic ligament is likely to originate from the fasciae of lingual muscles. Epithelial-mesenchymal interaction is strongly suggested in the development of the epiglottic cartilage and concomitant glands.

  13. Effect of a antisense oligonucleotide to noggin on the expression of nestin and GFAP in the hippocampus of adult rats%反义Noggin基因对成年大鼠海马内Nestin及GFAP表达的影响

    Institute of Scientific and Technical Information of China (English)

    徐海伟; 范晓棠

    2005-01-01

    目的探讨Noggin基因对成年大鼠海马内Nestin及GFAP表达的影响.方法反义寡核苷酸技术封闭内源性Noggin基因的表达,免疫组化法检测成年大鼠海马内Nestin与GFAP的表达.结果侧脑室连续4 d注射Noggin基因的反义寡核苷酸后,可见海马齿状回(dentate gyrus,DG)内Nestin阳性细胞数与GFAP阳性细胞数较对照组显著增加;室下区GFAP阳性细胞数亦明显增加.结论Noggin对成年海马干细胞的分化有重要作用,内源性Noggin基因的表达可使神经干细胞向神经元方向分化.%Objective To examine the role of noggin on the expression of nestin and glial fibrillary acidic protein (GFAP) in the hippocampus of adult rats. Methods Antisense oligodeoxynucleotide (ASODN) technique was employed to inhibit endogenous noggin expression and immunohistochemistry was used to detect the expressions of Nestin and GFAP in the hippocampus of adult rats. Results It was observed that the number of nestin and GFAP immunoreactive cells in the dentate gyrus (DG) of hippocampus was increased in adult rats treated with antisense oligodeoxynucleotide to noggin. Moreover, the number of GFAP immunoreactive cells was increased in the subventricular zone of the rats treated with antisense oligodeoxynucleotide to noggin. Conclusion The results in the present study indicates that noggin may play a role in the differentiation of neural stem cells in the adult hippocampus, and it promotes the differentiation of neural stem cells in the DG to neuronal fate.

  14. Application of 4-wavelength optical intrinsic signal imaging in monitoring peri-infarct depo-larizations in GFAP-/-Vim-/-mice%四波长内源光信号成像在GFAP-/-Vim-/-小鼠脑梗塞周围扩散性去极化中的应用

    Institute of Scientific and Technical Information of China (English)

    吕建平; 曹志恺; Lee Jin-Moo

    2015-01-01

    Objective To study optical intrinsic signal (OIS) imaging of peri-infarct depolarizations (PIDs) in mice and investigate the influence of knockout of glial fibrillary acidic protein and vimentin on PIDs. Methods GFAP-/-Vim-/-mice and GFAP+/+Vim+/+mice were subjected to MCAO by standard intraluminal filament method. The main characteristics of PIDs in 4 h were studied by 4-wavelength OIS imaging technique. Results PIDs were identified as consistent, red and blue interaction waves in the cortical reflectance that slowly propagated peripherally from the origin site. There were 5 patterns of PID propagation, namely rostro-caudal, latero-medial, caudo-rostral, contralateral and medial-lateral. No significant differences were found in PID frequency, propagation patterns, velocity or duration time between the two groups (P>0.05). Conclusion The 4-wavelength OIS system allows acquisition of high temporal-spatial resolution color images for analyzing temporal-spatial characteristics of PIDs in detail. Knockout of GFAP and vimentin do not affect PIDs in 4 h following middle cerebral artery occlusion.%目的:观测小鼠脑梗塞周围扩散性去极化(PIDs)的内源光信号变化特征,探索胶质纤维酸性蛋白(GFAP)和波形蛋白(Vim)基因敲除对PIDs的影响。方法以GFAP和Vim基因敲除(GFAP-/-Vim-/-)小鼠及其野生型(GFAP+/+Vim+/+)小鼠为研究对象,线栓法制备大脑中动脉梗塞(MCAO)模型,应用四波长内源光信号(OIS)成像技术监测两组动物4 h内PIDs的发作情况。结果 OIS成像显示PIDs在空间分布上表现为连续的、红蓝相间的弧形波,由发源处向四周缓慢播散;PIDs存在5种不同空间播散类型:喙-尾播散类型、侧方-内侧播散类型、尾-喙播散类型、对侧播散类型和内侧-侧方播散类型。GFAP-/-Vim-/-小鼠和GFAP+/+Vim+/+小鼠PIDs的发作次数、播散类型、速度及时程组间比较无统计学差异(P>0.05)。结

  15. Developmental analysis of GFAP immunoreactivity in the cerebellum of the meander tail mutant mouse.

    Science.gov (United States)

    Grishkat, H L; Schwartz, E; Jain, G; Eisenman, L M

    1996-08-01

    It is thought that Bergmann glial fibers assist in the inward migration of granule cells. Model systems in which there is a perturbation of either the migrating cells or the glial cell population have been useful in understanding the migratory process. In the meander tail mutant mouse, the anterior cerebellar region is agranular, whereas the posterior cerebellum is relatively unaffected by the mutation. This study presents a qualitative analysis of the development of cerebellar radial glia in mea/mea and +/mea mice aged from postnatal day 0 to adult, using an antibody against the glia specific antigen, glial fibrillary acidic protein. The results indicate a slight delay in the onset of immunoreactivity in the mea/mea cerebellum and abnormal glial formation in the anterior and posterior regions by postnatal day 5. At postnatal day 11, the full complement of labeled fibers appears to be present and although they appear abnormal in formation, they eventually reach the surface and terminate in oddly shaped and irregularly spaced endfeet. In adult mea/mea and +/mea mice, as compared to the early postnatal stages, there is a significant reduction in GFAP immunoreactive fibers. Cresyl violet stained adult mea/mea sections revealed the presence of ectopic granule cells in radial columns and small clumps at the surface of and within the molecular layer of the caudal cerebellum. Quantitative analyses revealed a 4- to 5-fold increase in the number of ectopic granule cells in lobule VIII of the mea/mea when compared with the +/mea cerebellum. These results suggest that the radial glia in the mea/mea cerebellum exhibit some uncharacteristic morphologies, but that these abnormalities are most likely the consequence of environmental alterations produced by the mutant gene.

  16. Adalimumab (tumor necrosis factor-blocker) reduces the expression of glial fibrillary acidic protein immunoreactivity increased by exogenous tumor necrosis factor alpha in an organotypic culture of porcine neuroretina.

    Science.gov (United States)

    Fernandez-Bueno, I; Garcia-Gutierrez, M T; Srivastava, G K; Gayoso, M J; Gonzalo-Orden, J M; Pastor, J C

    2013-01-01

    To determine if exogenous addition of tumor necrosis factor alpha (TNFα) exacerbates retinal reactive gliosis in an organotypic culture of porcine neuroretina and to evaluate if concomitant adalimumab, a TNF-blocker, diminishes it. Porcine retinal explants from 20 eyeballs were cultured. Cultures with 100 pg/ml TNFα, 10 µg/ml adalimumab, 100 pg/ml TNFα plus 10 µg/ml adalimumab, or controls without additives were maintained for 9 days. Freshly detached retinas were processed in parallel. TNFα levels in control culture supernatants were quantified with enzyme-linked immunosorbent assay. Cryostat sections were doubly immunostained for glial fibrillary acidic protein (GFAP), a marker for reactive gliosis, and cellular retinaldehyde-binding protein (CRALBP), a marker for Müller cells. Sections were also labeled with the isolectin IB4, a label for microglia/macrophages. TNFα in control culture supernatants was detected only at day 1. Compared to the fresh neuroretinal samples, upregulation of GFAP and downregulation of CRALBP occurred during the 9 days of culture. Exogenous TNFα stimulated glial cells to upregulate GFAP and downregulate CRALBP immunoreactivity. TNFα-treated cultures also initiated the growth of gliotic membranes and underwent retinal disorganization. Adalimumab inhibited the spontaneous increases in GFAP and maintained CRALBP. In combination with TNFα, adalimumab reduced GFAP expression and conserved CRALBP, with only slight retinal disorganization. No appreciable changes in IB4 labeling were observed under the different culture conditions. In cultured porcine neuroretina, spontaneous reactive gliosis and retinal disorganization were exacerbated by exogenous TNFα. Adalimumab reduced spontaneous changes and those induced by TNFα. Therefore, inhibiting TNFα may represent a novel approach to controlling retinal fibrosis observed in some human diseases.

  17. Effect of basic fibroblast growth factor on the expression of glial fibrillary acidic protein after tractive spinal cord injury in rats

    Institute of Scientific and Technical Information of China (English)

    LIU Lei; L(U) Bo; TU Chong-qi; CHI Lei-ting; WANG Guang-lin; PEI Fu-xing

    2005-01-01

    Objective: To investigate the effects of basic fibroblast growth factor (bFGF) on the expression of glial fibrillary acidic protein (GFAP) after tractive spinal cord injury in rats and to explore the recovery of spinal cord function.Methods: The rats were subjected to tractive spinal cord injury at T13-L2. Cortical somatosensory-evoked potential (CSEP) was closely monitored and when P1-N1 wave amplitude decreased to 70% of that before operation, a small-bore catheter was inserted below the injured plane through subarachnoid cavity. In the treatment groups, 20 μl of bFGF solution (containing 20 μg of bFGF) was injected through the catheter right after the operation and 1,2, 3, 4, 8, 12 and 24 h postoperatively. In the control group, same volume of normal saline was injected and every four rats were killed at 1, 4, 7, 14 and 21 d after the operation. Combined behavior score (CBS) and electro-physiological examination were adopted to evaluate function recovery. Expression of GFAP was observed by immuno-histochemical staining and was analyzed quantitatively by computer image analysis.Results: There was statistically significant difference in GFAP-positive cells between bFGF treatment group and the control group (P<0.01). Similar tendency was indicated by the results of CBS and CSEP.Conclusions: bFGF can induce large expression of GFAP after tractive spinal cord injury in rats and promote spinal function recovery, which is highly important for spinal cord regeneration.

  18. Time Course and Diagnostic Accuracy of Glial and Neuronal Blood Biomarkers GFAP and UCH-L1 in a Large Cohort of Trauma Patients With and Without Mild Traumatic Brain Injury.

    Science.gov (United States)

    Papa, Linda; Brophy, Gretchen M; Welch, Robert D; Lewis, Lawrence M; Braga, Carolina F; Tan, Ciara N; Ameli, Neema J; Lopez, Marco A; Haeussler, Crystal A; Mendez Giordano, Diego I; Silvestri, Salvatore; Giordano, Philip; Weber, Kurt D; Hill-Pryor, Crystal; Hack, Dallas C

    2016-05-01

    Glial fibrillary acidic protein (GFAP) and ubiquitin C-terminal hydrolase L1 (UCH-L1) have been widely studied and show promise for clinical usefulness in suspected traumatic brain injury (TBI) and concussion. Understanding their diagnostic accuracy over time will help translate them into clinical practice. To evaluate the temporal profiles of GFAP and UCH-L1 in a large cohort of trauma patients seen at the emergency department and to assess their diagnostic accuracy over time, both individually and in combination, for detecting mild to moderate TBI (MMTBI), traumatic intracranial lesions on head computed tomography (CT), and neurosurgical intervention. This prospective cohort study enrolled adult trauma patients seen at a level I trauma center from March 1, 2010, to March 5, 2014. All patients underwent rigorous screening to determine whether they had experienced an MMTBI (blunt head trauma with loss of consciousness, amnesia, or disorientation and a Glasgow Coma Scale score of 9-15). Of 3025 trauma patients assessed, 1030 met eligibility criteria for enrollment, and 446 declined participation. Initial blood samples were obtained in 584 patients enrolled within 4 hours of injury. Repeated blood sampling was conducted at 4, 8, 12, 16, 20, 24, 36, 48, 60, 72, 84, 96, 108, 120, 132, 144, 156, 168, and 180 hours after injury. Diagnosis of MMTBI, presence of traumatic intracranial lesions on head CT scan, and neurosurgical intervention. A total of 1831 blood samples were drawn from 584 patients (mean [SD] age, 40 [16] years; 62.0% [362 of 584] male) over 7 days. Both GFAP and UCH-L1 were detectible within 1 hour of injury. GFAP peaked at 20 hours after injury and slowly declined over 72 hours. UCH-L1 rose rapidly and peaked at 8 hours after injury and declined rapidly over 48 hours. Over the course of 1 week, GFAP demonstrated a diagnostic range of areas under the curve for detecting MMTBI of 0.73 (95% CI, 0.69-0.77) to 0.94 (95% CI, 0.78-1.00), and UCH-L1 demonstrated

  19. Tuberal hypothalamic expression of the glial intermediate filaments, glial fibrillary acidic protein and vimentin across the turkey hen (Meleagris gallopavo) reproductive cycle: Further evidence for a role of glial structural plasticity in seasonal reproduction.

    Science.gov (United States)

    Steinman, Michael Q; Valenzuela, Anthony E; Siopes, Thomas D; Millam, James R

    2013-11-01

    Glia regulate the hypothalamic-pituitary-gonadal (HPG) axis in birds and mammals. This is accomplished mechanically by ensheathing gonadotrophin-releasing hormone I (GnRH) nerve terminals thereby blocking access to the pituitary blood supply, or chemically in a paracrine manner. Such regulation requires appropriate spatial associations between glia and nerve terminals. Female turkeys (Meleagris gallopavo) use day length as a primary breeding cue. Long days activate the HPG-axis until the hen enters a photorefractory state when previously stimulatory day lengths no longer support HPG-axis activity. Hens must then be exposed to short days before reactivation of the reproductive axis occurs. As adult hens have discrete inactive reproductive states in addition to a fertile state, they are useful for examining the glial contribution to reproductive function. We immunostained tuberal hypothalami from short and long-day photosensitive hens, plus long-day photorefractory hens to examine expression of two intermediate filaments that affect glial morphology: glial fibrillary acidic protein (GFAP) and vimentin. GFAP expression was drastically reduced in the central median eminence of long day photosensitive hens, especially within the internal zone. Vimentin expression was similar among groups. However, vimentin-immunoreactive fibers abutting the portal vasculature were significantly negatively correlated with GFAP expression in the median eminence, which is consistent with our hypothesis for a reciprocal relationship between GFAP and vimentin expression. It appears that up-regulation of GFAP expression in the central median eminence of turkey hens is associated with periods of reproductive quiescence and that photofractoriness is associated with the lack of a glial cytoskeletal response to long days.

  20. SLC27 fatty acid transport proteins.

    Science.gov (United States)

    Anderson, Courtney M; Stahl, Andreas

    2013-01-01

    The uptake and metabolism of long chain fatty acids (LCFA) are critical to many physiological and cellular processes. Aberrant accumulation or depletion of LCFA underlie the pathology of numerous metabolic diseases. Protein-mediated transport of LCFA has been proposed as the major mode of LCFA uptake and activation. Several proteins have been identified to be involved in LCFA uptake. This review focuses on the SLC27 family of fatty acid transport proteins, also known as FATPs, with an emphasis on the gain- and loss-of-function animal models that elucidate the functions of FATPs in vivo and how these transport proteins play a role in physiological and pathological situations.

  1. Macromolecular mimicry of nucleic acid and protein

    DEFF Research Database (Denmark)

    Nautrup Pedersen, Gitte; Nyborg, Jens; Clark, Brian F

    1999-01-01

    of the concept of macromolecular mimicry. Macromolecular mimicry has further been proposed among initiation and release factors, thereby adding a new element to the description of protein synthesis in bacteria. Such mimicry has also been observed in other biological processes such as autoimmunity, DNA repair......Although proteins and nucleic acids consist of different chemical components, proteins can mimic structures and possibly also functions of nucleic acids. Recently, structural mimicry was observed between two elongation factors in bacterial protein biosynthesis leading to the introduction...

  2. Effect of erhuangfang on cerebral and spinal demyelination and regeneration as well as expression of glial fibrillary acidic protein in rats with experimental allergic encephalomyelitis

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    ventricle) and spinal cord (cervical enlargement and lumbar enlargement) collections,and then haematine-eosin (HE) staining and SLG myelin staining were used to observe demyelination and regeneration; meanwhile, immunohistochemical staining was used to observe the expression of glial fibriliary acidic protein (GFAP).MAIN OUTCOME MEASURES: Cerebral and spinal demyelination and regeneration as well as expression of GFAP in EAE rats.RESULTS: All 70 Lewis rats were involved in the final analysis. ① Demyelination and regeneration:Infiltration of inflammatory cells surrounding cerebrum and small venous vessels of spinal cord white matter,demyelination surrounding vessels and plentiful foam cells at myelinolysis sites were observed in the model group. Symptoms were relieved in the western medicine group and the Chinese herb group as compared with those in the model group. While, numbers of inflammatory infiltrated cells and vascular cuffs were decreased in focal region as compared with those in the model group; in addition, areas of softening focus and demyelination were decreased. ② Expression of GFAP: Volumes and numbers of positive cells of GFAP in white matter region were respectively bigger and higher than those of normal cells in the model group.Plentiful positive cells of GFAP were disorderly aggregated in hippocampus and surrounding small vessel cuffs. While, expression of GFAP was mildly increased surrounding focus in the Chinese herb group;however, GFAP did not express surrounding focus in the western medicine group. In addition, expressions of GFAP were not increased in non-focal region in both Chinese herb group and western medicine group.CONCLUSION: Both erhuangfang and hormone can relieve inflammatory reaction of central nervous system and demyelination of EAE rats. On one hand, erhuangfang can regulate reaction of astrocyte in two ways, relieve reaction and proliferation of astrocyte in non-focal region and maintain the protective effect of astrocyte on brain

  3. Cytokines: muscle protein and amino acid metabolism

    DEFF Research Database (Denmark)

    van Hall, Gerrit

    2012-01-01

    raises TNF-α and IL-6 to moderate levels, has only identified IL-6 as a potent cytokine, decreasing systemic amino acid levels and muscle protein metabolism. The marked decrease in circulatory and muscle amino acid concentrations was observed with a concomitant reduction in both the rates of muscle...... of IL-6 on the regulation of muscle protein metabolism but indirectly via IL-6 reducing amino acid availability. SUMMARY: Recent studies suggest that the best described cytokines TNF-α and IL-6 are unlikely to be the major direct mediators of muscle protein loss in inflammatory diseases. However...... protein synthesis and breakdown, that is, reduced turnover with a minor increase in net muscle degradation. Very similar observations have been made in models of acute inflammation, induced by high-dose endotoxin injection. However, these changes were suggested not to be attributed to a direct effect...

  4. Cytokines: muscle protein and amino acid metabolism

    DEFF Research Database (Denmark)

    van Hall, Gerrit

    2012-01-01

    raises TNF-α and IL-6 to moderate levels, has only identified IL-6 as a potent cytokine, decreasing systemic amino acid levels and muscle protein metabolism. The marked decrease in circulatory and muscle amino acid concentrations was observed with a concomitant reduction in both the rates of muscle...... protein synthesis and breakdown, that is, reduced turnover with a minor increase in net muscle degradation. Very similar observations have been made in models of acute inflammation, induced by high-dose endotoxin injection. However, these changes were suggested not to be attributed to a direct effect...... of IL-6 on the regulation of muscle protein metabolism but indirectly via IL-6 reducing amino acid availability. SUMMARY: Recent studies suggest that the best described cytokines TNF-α and IL-6 are unlikely to be the major direct mediators of muscle protein loss in inflammatory diseases. However...

  5. 氯化甲基汞对成年大鼠脑组织中GFAP和MBP的影响%Effect of different doses of methyl-mercuric chloride on GFAP,MBP of adult rats in different courses

    Institute of Scientific and Technical Information of China (English)

    范玉芹; 吕伟; 王树才; 曹秉振; 胡怀强

    2016-01-01

    目的:探讨不同剂量氯化甲基汞蓄积对成年大鼠脑组织胶质纤维酸性蛋白(glial fibrillary acidic protein,GFAP)、髓鞘碱性蛋白(myelin basic protein,MBP)的影响。方法120只 SD大鼠随机分为正常对照组、氯化甲基汞小剂量组(0.5 mg/kg )、氯化甲基汞中剂量组(1.0 mg/kg )、氯化甲基汞大剂量组(2.0 mg/kg)。每天灌胃1次,连续染毒,各组染毒时间分别为1、2、4周。采用免疫组织化学方法观察不同时间点各组大鼠脑组织中 GFAP、MBP的变化。结果各染毒时间组中氯化甲基汞剂量增加,大鼠脑组织 GFAP表达升高和 MBP表达降低,差异均有统计学意义(P<0.01)。同一剂量组不同染毒时间比较,随染毒时间延长,GFAP表达升高;MBP表达降低,差异均有统计学意义(P<0.01)。结论氯化甲基汞促进星形胶质细胞增殖,破坏髓鞘,并与汞的蓄积量与染毒时间相关。%Objective To investigate the effect of different doses of methyl-mercuric chloride(MMC)on glial fibrillary acidic protein (GFAP)and myelin basic protein (MBP)of adult rats in different courses. Methods 120 Sprague Dawley rats were randomly divided into the normal control group and MMC low dose group (0.5 mg/kg),middle dose(1 mg/Kg),high dose (2 mg/kg).MMC group rats were given by different dose MMC gavage once a day for 1w, 2w and 4w respectively.GFAP and MBP were measured by immunohistochemistry at different times after administrating various dosages.Results The GFAP expression increased and MBP expression decreased with MMC dosage in various times,the differences of groups were significant (P<0.01).As the time went on,the GFAP expression increased and MBP expression decreased in the same dose group,the differences of groups were significant (P <0.01).Conclusions MMC can destroy myelin sheath and promote GFAP,which shows a significant dose-effect and time-effect correlation.

  6. Glial and neuronal proteins in serum predict outcome after severe traumatic brain injury.

    NARCIS (Netherlands)

    Vos, P.E.; Lamers, K.J.B.; Hendriks, J.C.M.; Haaren, M. van; Beems, T.; Zimmerman, C.; Geel, W.J.A. van; Reus, H.P.M. de; Biert, J.; Verbeek, M.M.

    2004-01-01

    OBJECTIVE: To study the ability of glial (glial fibrillary acidic protein [GFAP] and S100b) and neuronal (neuron specific enolase [NSE]) protein levels in peripheral blood to predict outcome after severe traumatic brain injury. METHODS: Eighty-five patients with severe traumatic brain injury (admiss

  7. Methylprednisolone Inhibits the Expression of Glial Fibrillary Acidic Protein and Chondroitin Sulfate Proteoglycans in Reactivated Astrocytes

    Institute of Scientific and Technical Information of China (English)

    WEI-LIN LIU; YI-HSUAN LEE; SHIH-YING TSAI; CHUNG YI HSU; YU-YO SUN; LIANG-YO YANG; SHING-HAN TSAI; WEI-CHUNG VIVIAN YANG

    2008-01-01

    创伤后的神经胶质增生导致硫酸软骨素蛋白聚糖(CSPG)的显著表达,从而抑制轴突生长和再生.甲基强地松龙(MP),一种合成的糖皮质激素,在急性脊髓损伤(SCI)的治疗中有神经保护作用和抗炎效应.但是,MP对于CSPG在活性胶质细胞中的表达的作用尚不清楚.本文用a-氨基-3-羟基-5-甲基-4-异恶唑丙酸酯(AM-PA)诱导星形胶质细胞再活化,用环噻嗪模拟SCI的兴奋性中毒刺激.AMPA治疗后,星形胶质细胞再活化的标志物-胶质纤维酸性蛋白(GFAP)、CSPG神经聚糖和磷酸盐的表达都显著上调.AMPA治疗星形胶质细胞的条件培养液强烈抑制大鼠背根神经节中神经元的轴突生长,但这种作用能被MP的预处理所逆转.此外,MP下调成年SCI大鼠中GFAP和CSPG的表达,对抗RU486的糖皮质激素受体(GR)和GR siRNA能逆转MP对GFAP和神经聚糖表达的抑制作用.这些结果提示,MP能在兴奋性中毒损伤后通过GR介导的星形胶质细胞再活化下调和GSPG表达抑制来改善神经修复,促进轴突生长.%Reactive gliosis caused by post-traumatic injury often results in marked expression of chondroitin sul-fate proteoglycan(CSPG), which inhibits neurite outgrowth and regeneration. Methylprednisolone (MP), a synthet-ic glucocorticoid, has been shown to have neuroprotective and anti-inflammatory effects for the treatment of acute spinal cord injury (SCI). However, the effect of MP on CSPG expression in reactive glial cells remains unclear. In our study, we induced astrocyte reactivation using a-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) and cyclothiazide to mimic the exciotoxic stimuli of SCI. The expression of glial fibrillary acidic protein (GFAP), a marker of astrocyte reactivation, and CSPG neurocan and phosphacan were significantly elevated by AMPA treat-ment. The conditioned media from AMPA-treated astrocytes strongly inhibited neurite outgrowth of rat dorsal root ganglion neurons, and this

  8. 不同剂量羊瘙痒因子263K颅内感染仓鼠临床终末期脑中GFAP表达的研究%Analyses of the expressions of GFAP in the brain tissues of hamsters infected with various amounts of scrapie strain 263K at terminal stage

    Institute of Scientific and Technical Information of China (English)

    田婵; 张宝云; 石琦; 韩俊; 高晨; 韩露; 董小平

    2008-01-01

    Objective To investigate whether gliosis in the brain tissues of the hamsters infected with various amounts of scrapie strain 263K is correlated with the inoculation doses or the incubation times. Methods The total values of glial fibrillary acidic protein (GFAP) in brains were evaluated by Western Blots and the GFAP-stained cells were detected by immunohistochemistry (IHC). The characteristics of GFAP distributions among various groups were defined by quantitive and statistic analyses. Results Compared with the brain tissues of normal hamsters, remarkably higher total GFAP levels and more GFAP-stained cells were observed in the brain tissues of infected ones, howbeit, no significant difference was addressed among the infected groups. Conclusion Inoculations of various amounts of scrapie strain 263K into experimental hamsters intracerebrally induced the similar patterns of gliosis in the brains at the clinically terminal stage, regardless of infectious doses and incubation times.%目的 研究不同剂量羊瘙痒因子263 K经颅内注射感染仓鼠后,在发病的终末期星形胶质细胞增生程度是否与注射剂量及潜伏期长短有关.方法 以胶质纤维酸性蛋白(glial fibrill aryacidic protein,GFAP)作为星形胶质细胞增生的分子标志物,采用Western Blot和免疫组化方法检测感染仓鼠终末期脑匀浆和脑组织病理切片中的GFAP表达,经定量分析比较各感染剂量组间是否存在差异.结果与正常对照相比,不同剂量感染仓鼠发病终末期脑组织GFAP阳性细胞数量和总GFAP含量均明显升高,但各感染剂量组间无显著差异.结论 不同感染剂量羊瘙痒因子263K经颅内注射感染仓鼠在发病终末期脑中星形胶质细胞的增生程度相似,与感染剂量及潜伏期无关联性.

  9. Nucleic acids, proteins, and chirality

    Science.gov (United States)

    Usher, D. A.; Profy, A. T.; Walstrum, S. A.; Needels, M. C.; Bulack, S. C.; Lo, K. M.

    1984-01-01

    The present investigation is concerned with experimental results related, in one case, to the chirality of nucleotides, and, in another case, to the possibility of a link between the chirality of nucleic acids, and that of peptides. It has been found that aminoacylation of the 'internal' hydroxyl group of a dinucleoside monophosphate can occur stereoselectively. However, this reaction has not yet been made a part of a working peptide synthesis scheme. The formation and cleavage of oligonucleotides is considered. In the event of the formation of a helical complex between the oligonucleotide and the polymer, 1-prime,5-prime-bonds in the oligomer are found to become more resistant towards cleavage. The conditions required for peptide bond formation are examined, taking into account the known structures of RNA and possible mechanisms for prebiotic peptide bond formation. The possibility is considered that the 2-prime,5-prime-internucleotide linkage could have played an important part in the early days of biological peptide synthesis.

  10. Functional characterization of a GFAP variant of uncertain significance in an Alexander disease case within the setting of an individualized medicine clinic.

    Science.gov (United States)

    Boczek, Nicole J; Sigafoos, Ashley N; Zimmermann, Michael T; Maus, Rachel L; Cousin, Margot A; Blackburn, Patrick R; Urrutia, Raul; Clark, Karl J; Patterson, Marc C; Wick, Myra J; Klee, Eric W

    2016-09-01

    A de novo GFAP variant, p.R376W, was identified in a child presenting with hypotonia, developmental delay, and abnormal brain MRI. Following the 2015 ACMG variant classification guidelines and the functional studies showing protein aggregate formation in vitro, p.R376W should be classified as a pathogenic variant, causative for Alexander disease.

  11. Anchoring of proteins to lactic acid bacteria

    NARCIS (Netherlands)

    Leenhouts, K; Buist, Girbe; Kok, Jan

    1999-01-01

    The anchoring of proteins to the cell surface of lactic acid bacteria (LAB) using genetic techniques is an exciting and emerging research area that holds great promise for a wide variety of biotechnological applications. This paper reviews five different types of anchoring domains that have been exp

  12. Anchoring of proteins to lactic acid bacteria

    NARCIS (Netherlands)

    Leenhouts, K; Buist, Girbe; Kok, Jan

    1999-01-01

    The anchoring of proteins to the cell surface of lactic acid bacteria (LAB) using genetic techniques is an exciting and emerging research area that holds great promise for a wide variety of biotechnological applications. This paper reviews five different types of anchoring domains that have been exp

  13. 慢性砷中毒对小鼠齿状回GFAP表达的影响%Effects of chronic arsenic poisoning on GFAP expression in the dentate gyms of adult mice

    Institute of Scientific and Technical Information of China (English)

    康朝胜; 孙宝飞; 余资江; 李玉飞

    2011-01-01

    Objective To investigate activation of glial fibrillary acidic protein (GFAP) in the dentate gyrus of adult mice after chronic arsenic poisoning. Methods 80 healthy adult Kunming mice, weighing 20-22g, were divided into four groups: the normal control group, the low-dose group, the medium-dose group and the high-dose group ( 10 males and 10 females in each group). Mice in the four groups were respectively fed with distilled water, 1/5 LD50, 1/10 LD50 and 1/40 LD50 AS2O3 for 3 months, and the dosage was adjusted according to changes of weight. Then ability of learning and memory was tested by a Y-maze, and expression of the GFAP protein in the dentate gyrus by Western blot and immunohistochemistry. Results Ability of learning and memory in the high-dose group was significantly lower than that in the normal control group ( P < 0.05 ). Immunohistochemical results showed that the number of GFAP-positive cells in the dentate gyrus was significantly more increased in the dose groups compared with the normal control group(P<0.01 ), and the average optical density was also increased (P <0.01 ). Western blot results showed the GFAP protein content increased with the dosage increasing (P <0.01 ). Conclusion Chronic arsenic poisoning might damage ability of learning and memory in mice, which may be related to proliferation of astroeytes and expression of GFAP in the dentate gyrus.%目的 研究砷中毒后小鼠齿状回胶质原纤维酸性蛋白(GFAP)的变化.方法 选取健康成年昆明小鼠80只,雌雄各半,分为对照组及慢性砷中毒高、中、低剂量组,每组20只,分别以蒸馏水、1/5 LD50、1/10 LD50、1/40LD50 As2O3连续灌胃3个月,根据其体质量变化随时调整用药剂量,采用Y-电迷宫检测各组小鼠学习记忆行为,采用免疫组织化学和蛋白印迹技术检测不同浓度砷中毒对小鼠齿状回部位GFAP表达的影响.结果 与正常对照组比较,高剂量砷中毒组学习、记忆Y-迷

  14. Alimentary proteins, amino acids and cholesterolemia.

    Science.gov (United States)

    Blachier, François; Lancha, Antonio H; Boutry, Claire; Tomé, Daniel

    2010-01-01

    Numerous data from both epidemiological and experimental origins indicate that some alimentary proteins and amino acids in supplements can modify the blood LDL cholesterol, HDL cholesterol and total cholesterol. After an initial approval of the health claim for soy protein consumption for the prevention of coronary heart disease, more recently it has been concluded from an overall analysis of literature that isolated soy protein with isoflavones only slightly decrease LDL and total cholesterol. Other plant extracts and also some proteins from animal origin have been reported to exert a lowering effect on blood cholesterol when compared with a reference protein (often casein). The underlying mechanisms are still little understood. Individual amino acids and mixture of amino acids have also been tested (mostly in animal studies) for their effects on cholesterol parameters and on cholesterol metabolism. Methionine, lysine, cystine, leucine, aspartate and glutamate have been tested individually and in combination in different models of either normo or hypercholesterolemic animals and found to be able to modify blood cholesterol and/or LDL cholesterol and/or HDL cholesterol. It is however not known if these results are relevant to human nutrition.

  15. Protein and Amino Acid Profiles of Different Whey Protein Supplements.

    Science.gov (United States)

    Almeida, Cristine C; Alvares, Thiago S; Costa, Marion P; Conte-Junior, Carlos A

    2016-01-01

    Whey protein (WP) supplements have received increasing attention by consumers due to the high nutritional value of the proteins and amino acids they provide. However, some WP supplements may not contain the disclosed amounts of the ingredients listed on the label, compromising the nutritional quality and the effectiveness of these supplements. The aim of this study was to evaluate and compare the contents of total protein (TP), α-lactalbumin (α-LA), β-lactoglobulin (β-LG), free essential amino acids (free EAA), and free branched-chain amino acids (free BCAA), amongst different WP supplements produced by U.S. and Brazilian companies. Twenty commercial brands of WP supplements were selected, ten manufactured in U.S. (WP-USA) and ten in Brazil (WP-BRA). The TP was analyzed using the Kjeldahl method, while α-LA, β-LG, free EAA, and free BCAA were analyzed using HPLC system. There were higher (p 0.05) in the content of free EAA between WP-USA and WP-BRA. Amongst the 20 brands evaluated, four WP-USA and seven WP-BRA had lower (p < 0.05) values of TP than those specified on the label. In conclusion, the WP-USA supplements exhibited better nutritional quality, evaluated by TP, α-LA, β-LG, and free BCAA when compared to WP-BRA.

  16. Inadequacy of prebiotic synthesis as origin of proteinous amino acids.

    Science.gov (United States)

    Wong, J T; Bronskill, P M

    1979-07-18

    The production of some nonproteinous, and lack of production of other proteinous, amino acids in model prebiotic synthesis, along with the instability of glutamine and asparagine, suggest that not all of the 20 present day proteinous amino acids gained entry into proteins directly from the primordial soup. Instead, a process of active co-evolution of the genetic code and its constituent amino acids would have to precede the final selection of these proteinous amono acids.

  17. CSF Neurofilament Proteins Levels are Elevated in Sporadic Creutzfeldt-Jakob Disease

    NARCIS (Netherlands)

    van Eijk, Jeroen J. J.; van Everbroeck, Bart; Abdo, W. Farid; Kremer, Berry P. H.; Verbeek, Marcel M.

    2010-01-01

    In this study we investigated the cerebrospinal fluid (CSF) levels of neurofilament light (NFL) and heavy chain (NFHp35), total tau (t-tau), and glial fibrillary acidic protein (GFAP) to detect disease specific profiles in sporadic Creutzfeldt Jakob disease (sCJD) patients and Alzheimer's disease

  18. CSF neurofilament proteins levels are elevated in sporadic Creutzfeldt-Jakob disease.

    NARCIS (Netherlands)

    Eijk, J.J.J. van; Everbroeck, B. van; Abdo, W.; Kremer, H.P.H.; Verbeek, M.M.

    2010-01-01

    In this study we investigated the cerebrospinal fluid (CSF) levels of neurofilament light (NFL) and heavy chain (NFHp35), total tau (t-tau), and glial fibrillary acidic protein (GFAP) to detect disease specific profiles in sporadic Creutzfeldt Jakob disease (sCJD) patients and Alzheimer's disease

  19. CSF Neurofilament Proteins Levels are Elevated in Sporadic Creutzfeldt-Jakob Disease

    NARCIS (Netherlands)

    van Eijk, Jeroen J. J.; van Everbroeck, Bart; Abdo, W. Farid; Kremer, Berry P. H.; Verbeek, Marcel M.

    2010-01-01

    In this study we investigated the cerebrospinal fluid (CSF) levels of neurofilament light (NFL) and heavy chain (NFHp35), total tau (t-tau), and glial fibrillary acidic protein (GFAP) to detect disease specific profiles in sporadic Creutzfeldt Jakob disease (sCJD) patients and Alzheimer's disease (A

  20. Increased in vitro glial fibrillary acidic protein expression, telomerase activity, and telomere length after productive human immunodeficiency virus-1 infection in murine astrocytes.

    Science.gov (United States)

    Ojeda, Diego; López-Costa, Juan José; Sede, Mariano; López, Ester María; Berria, María Isabel; Quarleri, Jorge

    2014-02-01

    Although HIV-associated neurocognitive disorders (HAND) result from injury and loss of neurons, productive infection routinely takes place in cells of macrophage lineage. In such a complex context, astrocytosis induced by local chemokines/cytokines is one of the hallmarks of HIV neuropathology. Whether this sustained astrocyte activation is able to alter telomere-aging process is unknown. We hypothesized that interaction of HIV with astrocytes may impact astrocyte telomerase activity (TA) and telomere length in a scenario of astrocytic activation measured by expression of glial fibrillary acidic protein (GFAP). To test this hypothesis, cultured murine astrocytes were challenged with pseudotyped HIV/vesicular stomatitis virus (HIV/VSV) to circumvent the absence of viral receptors; and GFAP, telomerase activity, and telomere length were quantified. As an early and transient event after HIV infection, both TA activity and telomere length were significantly augmented (P telomere length, that may attenuate cell proliferation and enhance the astrocyte dysregulation, contributing to HIV neuropathogenesis. Understanding the mechanisms involved in HIV-mediated persistence by altering the telomere-related aging processes could aid in the development of therapeutic modalities for neurological complications of HIV infection.

  1. In Silico Prediction and Validation of Gfap as an miR-3099 Target in Mouse Brain.

    Science.gov (United States)

    Abidin, Shahidee Zainal; Leong, Jia-Wen; Mahmoudi, Marzieh; Nordin, Norshariza; Abdullah, Syahril; Cheah, Pike-See; Ling, King-Hwa

    2017-08-01

    MicroRNAs are small non-coding RNAs that play crucial roles in the regulation of gene expression and protein synthesis during brain development. MiR-3099 is highly expressed throughout embryogenesis, especially in the developing central nervous system. Moreover, miR-3099 is also expressed at a higher level in differentiating neurons in vitro, suggesting that it is a potential regulator during neuronal cell development. This study aimed to predict the target genes of miR-3099 via in-silico analysis using four independent prediction algorithms (miRDB, miRanda, TargetScan, and DIANA-micro-T-CDS) with emphasis on target genes related to brain development and function. Based on the analysis, a total of 3,174 miR-3099 target genes were predicted. Those predicted by at least three algorithms (324 genes) were subjected to DAVID bioinformatics analysis to understand their overall functional themes and representation. The analysis revealed that nearly 70% of the target genes were expressed in the nervous system and a significant proportion were associated with transcriptional regulation and protein ubiquitination mechanisms. Comparison of in situ hybridization (ISH) expression patterns of miR-3099 in both published and in-house-generated ISH sections with the ISH sections of target genes from the Allen Brain Atlas identified 7 target genes (Dnmt3a, Gabpa, Gfap, Itga4, Lxn, Smad7, and Tbx18) having expression patterns complementary to miR-3099 in the developing and adult mouse brain samples. Of these, we validated Gfap as a direct downstream target of miR-3099 using the luciferase reporter gene system. In conclusion, we report the successful prediction and validation of Gfap as an miR-3099 target gene using a combination of bioinformatics resources with enrichment of annotations based on functional ontologies and a spatio-temporal expression dataset.

  2. 首发精神分裂症治疗前后血清 NGF、BDNF、GFAP与临床症状的相关性研究%Correlation between pre-and post-treatment serum levels of NGF,BDNF,GFAP and severity of clinical symptoms in ;patients with first-episode schizophrenia

    Institute of Scientific and Technical Information of China (English)

    牛莉莉; 温科奇; 熊鹏; 曾勇; 徐飞; 李明; 黄晓江

    2015-01-01

    Objective To explore the changes of serum levels of Nerve Growth Factor (NGF),Brain-Derived Neurotrophic Factor (BDNF),Glial Fibrillary Acidic Protein (GFAP)in patients with first-episode schizophrenia,and to explore the correlation with severity of clinical symptoms pre-and post-treatment.Methods Serum levels of NGF,BDNF and GFAP were measured by using ELISA in 50 patients with first-episode schizophrenia (study group)and 78 healthy controls (control group), and were re-measured in subjects in study group after 3-month risperidone treatment.All patients in study group were assessed with Positive and Negative Syndrome Scale(PANSS)at baseline and at the end of the 3-month treatment to evaluate the severity of clinical symptoms.Results The pre-treatment serum level of GFAP in study group was significantly higher than that in control group (P <0.01),pre-treatment serum levels of NGF and BDNF in study group were significantly lower than those in control group (P <0.01).Serum levels of the 3 protein factors at the end of the treatment in study group were significantly lower than those at the baseline (P <0.05).Total score of PANSS and factor scores of positive symptoms,negative symptoms and general psychopathology at endpoint in study group were significantly lower than those at baseline (P <0.01).In study group,the pre-treatment serum level of NGF and GFAP were significantly positively correlated with factor score of negative symptoms of PNASS (P <0.05 ).The post-treatment NGF level was significantly negatively correlated with factor score of negative symptoms of PNASS (P =0.000 ).Post -treatment BDNF level was significantly positively correlated with factor score of general psychopathology (P =0.002),while post-treatment GFAP level was significantly negatively correlated with it (P =0.024). Conclusion Serum levels of NGF,BDNF and GFAP change with the improvement of clinical symptoms in patients with first-episode schizophrenia.NGF,BDNF and GFAP may be involved into

  3. Hypochlorous and peracetic acid induced oxidation of dairy proteins.

    Science.gov (United States)

    Kerkaert, Barbara; Mestdagh, Frédéric; Cucu, Tatiana; Aedo, Philip Roger; Ling, Shen Yan; De Meulenaer, Bruno

    2011-02-09

    Hypochlorous and peracetic acids, both known disinfectants in the food industry, were compared for their oxidative capacity toward dairy proteins. Whey proteins and caseins were oxidized under well controlled conditions at pH 8 as a function of the sanitizing concentration. Different markers for protein oxidation were monitored. The results established that the protein carbonyl content was a rather unspecific marker for protein oxidation, which did not allow one to differentiate the oxidant used especially at the lower concentrations. Cysteine, tryptophan, and methionine were proven to be the most vulnerable amino acids for degradation upon hypochlorous and peracetic acid treatment, while tyrosine was only prone to degradation in the presence of hypochlorous acid. Hypochlorous acid induced oxidation gave rise to protein aggregation, while during peracetic acid induced oxidation, no high molecular weight aggregates were observed. Protein aggregation upon hypochlorous acid oxidation could primarily be linked to tryptophan and tyrosine degradation.

  4. The Effect of Silybum marianum on GFAP and Spatial Memory in a Mouse Model of Alzheimer\\'s Disease

    Directory of Open Access Journals (Sweden)

    A Hadinia

    2010-01-01

    Full Text Available Introduction & Objective: Studies have shown that Silybum marianum have high levels of antioxidant polyphenolic substances and have neuro-protective effects on neurodegenerative diseases. Accordingly, this study was conducted to determine the possible effect of Silybum marianum on expression of and spatial memory in a mouse model of Alzheimer's disease. Materials & Methods: This experimental study was conducted at Yasuj University of Medical Sciences in 2009. Thirty adult male Wistar rats were allocated in three groups: sham group, experimental group, and lesion group, each consisting of ten rats. The experimental and lesion groups received Ibotonic acid of the NBM nucleus in stereotaxic apparatus whereas the sham group underwent surgical procedure without any injection. The experimental group received 200mg/kg of Silybum mirianum extract orally, diluted in 1% Arabic gum. Also the sham group received 1% Arabic gum every day for four weeks. The lesion group did not receive anything. The behavioral assessment was measured, after treatment , by using of Y maze test on day 7 and 28 in all groups. The ELISA method was used to measure the GFAP level in Hippocamp at the end of behavioral assessment. The collected data was analyzed by the SPSS software using ANOVA and Repeated Measures of Analysis Variance tests. Results:Improvement of behavioral performance of the experimental animals compared to the lesion and sham groups were increased significantly on day 7 and 28 (P <0.01 & P <0.001 respectively. The ELISA method showed that the level of the GFAP synthesis decreased in the experimental group compared to the lesion and sham groups (P <0.001. Conclusion: The Silybum marianum plant has a protective effect on the nerve tissue in a mouse model of Alzheimer's disease by decreasing of the GFAP synthesis and lead to the improvement of behavioral performance. :

  5. Covalent interactions between proteins and oxidation products of caffeoylquinic acid (chlorogenic acid)

    NARCIS (Netherlands)

    Prigent, S.V.E.; Voragen, A.G.J.; Visser, A.J.W.G.; Koningsveld, van G.A.; Gruppen, H.

    2007-01-01

    BACKGROUND: The interactions between phenolic compounds and proteins can modify protein properties important in the food industry. To understand the effects of these interactions, the covalent interactions between caffeoylquinic acid (chlorogenic acid, CQA) oxidised by polyphenol oxidase (PPO) at ac

  6. Supra- and infratentorial pediatric ependymomas differ significantly in NeuN, p75 and GFAP expression.

    Science.gov (United States)

    Hagel, Christian; Treszl, András; Fehlert, Julia; Harder, Jonas; von Haxthausen, Franziska; Kern, Meike; von Bueren, André O; Kordes, Uwe

    2013-04-01

    Ependymomas comprise 8 % of all intracranial tumors in children infratentorial tumors and GFAP to be expressed at significantly higher levels in infratentorial lesions. In conclusion, immunohistochemical expression of p75, NeuN and GFAP differed in ependymomas depending on tumor topography supporting the view of divergent cells of origin. However, because of the small sample size the results are of preliminary nature and replication in a larger cohort would be desirable.

  7. Enzymic Synthesis of Caffeoylglucaric Acid from Chlorogenic Acid and Glucaric Acid by a Protein Preparation from Tomato Cotyledons 1

    Science.gov (United States)

    Strack, Dieter; Gross, Wiltrud; Wray, Victor; Grotjahn, Lutz

    1987-01-01

    The phenylpropane metabolism of tomato (Lycopersicon esculentum Mill) cotyledons was investigated. The HPLC analysis revealed two hydroxycinnamic-acid conjugates as major components, identified as chlorogenic acid (5-O-caffeoylquinic acid) and caffeoylglucaric acid (2-O- or 5-O-caffeoyl-glucaric acid). Quantitative analyses indicated a precursor-product relationship between the chlorogenic and caffeoylglucaric acids. Protein preparations from tomato cotyledons were found to catalyze the formation of caffeoylglucaric acid with chlorogenic acid as acyl donor and free glucaric acid as acceptor molecule. This enzyme activity, possibly to be classified as hydroxycinnamoylquinic acid:glucaric acid hydroxycinnamoyltransferase, acts together with hydroxycinnamoyl-CoA: quinic acid hydroxycinnamoyltransferase. PMID:16665274

  8. Site specific incorporation of keto amino acids into proteins

    Science.gov (United States)

    Schultz, Peter G [La Jolla, CA; Wang, Lei [San Diego, CA

    2012-02-14

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate keto amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with keto amino acids using these orthogonal pairs.

  9. Site specific incorporation of keto amino acids into proteins

    Energy Technology Data Exchange (ETDEWEB)

    Schultz, Peter G [La Jolla, CA; Wang, Lei [San Diego, CA

    2012-02-14

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate keto amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with keto amino acids using these orthogonal pairs.

  10. Site specific incorporation of keto amino acids into proteins

    Energy Technology Data Exchange (ETDEWEB)

    Schultz, Peter G. (La Jolla, CA); Wang, Lei (San Diego, CA)

    2011-03-22

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate keto amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with keto amino acids using these orthogonal pairs.

  11. Site specific incorporation of keto amino acids into proteins

    Energy Technology Data Exchange (ETDEWEB)

    Schultz, Peter G [La Jolla, CA; Wang, Lei [San Diego, CA

    2011-12-06

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate keto amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with keto amino acids using these orthogonal pairs.

  12. Regulation of intestinal protein metabolism by amino acids.

    Science.gov (United States)

    Bertrand, Julien; Goichon, Alexis; Déchelotte, Pierre; Coëffier, Moïse

    2013-09-01

    Gut homeostasis plays a major role in health and may be regulated by quantitative and qualitative food intake. In the intestinal mucosa, an intense renewal of proteins occurs, at approximately 50% per day in humans. In some pathophysiological conditions, protein turnover is altered and may contribute to intestinal or systemic diseases. Amino acids are key effectors of gut protein turnover, both as constituents of proteins and as regulatory molecules limiting intestinal injury and maintaining intestinal functions. Many studies have focused on two amino acids: glutamine, known as the preferential substrate of rapidly dividing cells, and arginine, another conditionally essential amino acid. The effects of glutamine and arginine on protein synthesis appear to be model and condition dependent, as are the involved signaling pathways. The regulation of gut protein degradation by amino acids has been minimally documented until now. This review will examine recent data, helping to better understand how amino acids regulate intestinal protein metabolism, and will explore perspectives for future studies.

  13. SIFT: predicting amino acid changes that affect protein function

    OpenAIRE

    Ng, Pauline C.; Henikoff, Steven

    2003-01-01

    Single nucleotide polymorphism (SNP) studies and random mutagenesis projects identify amino acid substitutions in protein-coding regions. Each substitution has the potential to affect protein function. SIFT (Sorting Intolerant From Tolerant) is a program that predicts whether an amino acid substitution affects protein function so that users can prioritize substitutions for further study. We have shown that SIFT can distinguish between functionally neutral and deleterious amino acid changes in...

  14. Fatty acid acylation of proteins: specific roles for palmitic, myristic and caprylic acids

    Directory of Open Access Journals (Sweden)

    Rioux Vincent

    2016-05-01

    Full Text Available Fatty acid acylation of proteins corresponds to the co- or post-translational covalent linkage of an acyl-CoA, derived from a fatty acid, to an amino-acid residue of the substrate protein. The cellular fatty acids which are involved in protein acylation are mainly saturated fatty acids. Palmitoylation (S-acylation corresponds to the reversible attachment of palmitic acid (C16:0 via a thioester bond to the side chain of a cysteine residue. N-terminal myristoylation refers to the covalent attachment of myristic acid (C14:0 by an amide bond to the N-terminal glycine of many eukaryotic and viral proteins. Octanoylation (O-acylation typically concerns the formation of an ester bond between octanoic acid (caprylic acid, C8:0 and the side chain of a serine residue of the stomach peptide ghrelin. An increasing number of proteins (enzymes, hormones, receptors, oncogenes, tumor suppressors, proteins involved in signal transduction, eukaryotic and viral structural proteins have been shown to undergo fatty acid acylation. The addition of the acyl moiety is required for the protein function and usually mediates protein subcellular localization, protein-protein interaction or protein-membrane interaction. Therefore, through the covalent modification of proteins, these saturated fatty acids exhibit emerging specific and important roles in modulating protein functions. This review provides an overview of the recent findings on the various classes of protein acylation leading to the biological ability of saturated fatty acids to regulate many pathways. Finally, the nutritional links between these elucidated biochemical mechanisms and the physiological roles of dietary saturated fatty acids are discussed.

  15. Nanopore biosensors for detection of proteins and nucleic acids

    NARCIS (Netherlands)

    Maglia, Giovanni; Soskine, Mikhael

    2014-01-01

    Described herein are nanopore biosensors based on a modified cytolysin protein. The nanopore biosensors accommodate macromoiecules including proteins and nucleic acids, and may additionally comprise ligands with selective binding properties.

  16. Nanopore biosensors for detection of proteins and nucleic acids

    NARCIS (Netherlands)

    Maglia, Giovanni; Soskine, Mikhael

    2014-01-01

    Described herein are nanopore biosensors based on a modified cytolysin protein. The nanopore biosensors accommodate macromoiecules including proteins and nucleic acids, and may additionally comprise ligands with selective binding properties.

  17. Intumescent features of nucleic acids and proteins

    Energy Technology Data Exchange (ETDEWEB)

    Alongi, Jenny, E-mail: jenny.alongi@polito.it; Cuttica, Fabio; Blasio, Alessandro Di; Carosio, Federico; Malucelli, Giulio

    2014-09-10

    Highlights: • The combustion resistance of DNA and caseins to different heat fluxes was studied. • Upon heating, DNA and caseins exhibited an intumescent behaviour. • The char derived from DNA was more stable and coherent than that from caseins. - Abstract: Are nucleic acids and proteins intumescent molecules? In order to get an answer, in the present manuscript, powders of deoxyribose nucleic acids (DNA) and caseins have been exposed to different heat fluxes under a cone calorimeter source and to the direct application of a propane flame. Under these conditions, DNA and caseins exhibited a typical intumescent behaviour, generating a coherent expanded cellular carbonaceous residue (char), extremely resistant to heat exposure. The resulting volumetric expansion as well as the resistance of the formed char turned out to be dependent on (i) the chemical structure of the chosen biomacromolecule, (ii) the evolution of ammonia and (iii) the adopted heat flux in cone calorimetry tests (namely, 25, 35, 50 and 75 kW/m{sup 2}). The presence of ribose units within the DNA backbone determined the formation of highly expanded and coherent residues as compared to those obtained from caseins. Indeed, under a heat flux of 35 kW/m{sup 2}, when a carbon source (i.e. common cane sugar) was added to caseins, the resulting char was similar to that formed by DNA. Furthermore, the char expansion was ascribed to the evolution of ammonia released by these biomacromolecules upon heating, as detected by thermogravimetry coupled to infrared spectroscopy, and confirmed by scanning electron microscopy experiments performed on the bubbles present in the residues of flammability tests.

  18. Self-assembly of proteins and their nucleic acids

    Directory of Open Access Journals (Sweden)

    Terrett Jon

    2003-01-01

    Full Text Available Abstract We have developed an artificial protein scaffold, herewith called a protein vector, which allows linking of an in-vitro synthesised protein to the nucleic acid which encodes it through the process of self-assembly. This protein vector enables the direct physical linkage between a functional protein and its genetic code. The principle is demonstrated using a streptavidin-based protein vector (SAPV as both a nucleic acid binding pocket and a protein display system. We have shown that functional proteins or protein domains can be produced in vitro and physically linked to their DNA in a single enzymatic reaction. Such self-assembled protein-DNA complexes can be used for protein cloning, the cloning of protein affinity reagents or for the production of proteins which self-assemble on a variety of solid supports. Self-assembly can be utilised for making libraries of protein-DNA complexes or for labelling the protein part of such a complex to a high specific activity by labelling the nucleic acid associated with the protein. In summary, self-assembly offers an opportunity to quickly generate cheap protein affinity reagents, which can also be efficiently labelled, for use in traditional affinity assays or for protein arrays instead of conventional antibodies.

  19. Absolute quantitation of proteins by Acid hydrolysis combined with amino Acid detection by mass spectrometry

    DEFF Research Database (Denmark)

    Mirgorodskaya, Olga A; Körner, Roman; Kozmin, Yuri P;

    2012-01-01

    Amino acid analysis is among the most accurate methods for absolute quantification of proteins and peptides. Here, we combine acid hydrolysis with the addition of isotopically labeled standard amino acids and analysis by mass spectrometry for accurate and sensitive protein quantitation...

  20. 脑出血患者血清NSE、GFAP、BDNF水平变化与认知障碍相关性研究%Correlation of serum levels of NSE, GFAP, BDNF and cognitive impairment in patients with intracerebral hemorrhage

    Institute of Scientific and Technical Information of China (English)

    刘宇明; 邓燕华; 许治强; 梁燕玲; 胡佳佳; 林永强

    2016-01-01

    Objective To investigate the serum neuron specific enolase(NSE)and glial fibrillary acidic protein(GFAP), brain derived neural nutrition factor(BDNF)level changes in cerebral hemorrhage patients and the correlation with cognitive impairment. Method 100 cases of acute spontaneous cerebral hemorrhage were selected, and the corresponding treatments were given in the hospital, the seru NSE, GFAP and BDNF were detected. 100 patients were divided into cognitive impairment group(57cases)and non cognitive impairment group(43cases) according to whether there were cognitive impairment. Those three indicators were compared between different levels of cognitive injury(MMSE score)group, and the correlation between the three indicators and MMSE. Results The serum GFAP and NSE were significantly higher in the patients with cognitive impairment than those without cognitive impairment, and BDNF was significantly lower(t=7.039,t=2.247,t=4.847,P<0.01). The serum levels of NSE, GFAP and BDNF were significantly different in the mild injury group, moderate injury group and severe injury group (F=22.752, F=31.506, F=38.294, P<0.01). The levels of serum NSE and GFAP were positively correlated with MMSE scores(r=0.641, r=0.604, P<0.05), and BDNF was negatively correlated with MMSE scores(r=0.582, P<0.05). Conclusion The levels of serum NSE, GFAP and BDNF in patients with cerebral hemorrhage are closely related to cognitive dysfunction after stroke, which can be used to determine the severity of cognitive impairment in patients with cerebral hemorrhage.%目的:探讨脑出血患者血清神经元特异性烯醇化酶(NSE)、胶质纤维酸性蛋白(GFAP)、脑源性神经营养因子(BDNF)水平变化与认知障碍相关性。方法选择急性自发性脑出血患者100例,入院时均给予相应治疗,并进行 NSE、GFAP、BDNF 检测。将100例患者依据是否发生认知障碍分为认知障碍组(57例)和无认知障碍组(43例)。并

  1. Echinococcus granulosus fatty acid binding proteins subcellular localization.

    Science.gov (United States)

    Alvite, Gabriela; Esteves, Adriana

    2016-05-01

    Two fatty acid binding proteins, EgFABP1 and EgFABP2, were isolated from the parasitic platyhelminth Echinococcus granulosus. These proteins bind fatty acids and have particular relevance in flatworms since de novo fatty acids synthesis is absent. Therefore platyhelminthes depend on the capture and intracellular distribution of host's lipids and fatty acid binding proteins could participate in lipid distribution. To elucidate EgFABP's roles, we investigated their intracellular distribution in the larval stage by a proteomic approach. Our results demonstrated the presence of EgFABP1 isoforms in cytosolic, nuclear, mitochondrial and microsomal fractions, suggesting that these molecules could be involved in several cellular processes.

  2. Protein expression differs between neural progenitor cells from the adult rat brain subventricular zone and olfactory bulb

    Directory of Open Access Journals (Sweden)

    Bürgers Heinrich F

    2008-01-01

    Full Text Available Abstract Background Neural progenitor cells can be isolated from various regions of the adult mammalian brain, including the forebrain structures of the subventricular zone and the olfactory bulb. Currently it is unknown whether functional differences in these progenitor cell populations can already be found on the molecular level. Therefore, we compared protein expression profiles between progenitor cells isolated from the subventricular zone and the olfactory bulb using a proteomic approach based on two-dimensional gel electrophoresis and mass spectrometry. The subventricular zone and the olfactory bulb are connected by the Rostral Migratory Stream (RMS, in which glial fibrillary acidic protein (GFAP-positive cells guide neuroblasts. Recent literature suggested that these GFAP-positive cells possess neurogenic potential themselves. In the current study, we therefore compared the cultured neurospheres for the fraction of GFAP-positive cells and their morphology of over a prolonged period of time. Results We found significant differences in the protein expression patterns between subventricular zone and olfactory bulb neural progenitor cells. Of the differentially expressed protein spots, 105 were exclusively expressed in the subventricular zone, 23 showed a lower expression and 51 a higher expression in the olfactory bulb. The proteomic data showed that more proteins are differentially expressed in olfactory bulb progenitors with regard to proteins involved in differentiation and microenvironmental integration, as compared to the subventricular zone progenitors. Compared to 94% of all progenitors of the subventricular zone expressed GFAP, nearly none in the olfactory bulb cultures expressed GFAP. Both GFAP-positive subpopulations differed also in morphology, with the olfactory bulb cells showing more branching. No differences in growth characteristics such as doubling time, and passage lengths could be found over 26 consecutive passages in the

  3. PROTEINS, PEPTIDES AND AMINO ACIDS AS MARKERS OF BRONCHOPULMONARY DISEASES

    Directory of Open Access Journals (Sweden)

    V. I. Fyodorov

    2013-01-01

    Full Text Available The article is a review of current literature on a content of proteins, peptides and amino acids in human exhaled breath. The results of proteomics and metabolomics applying for selective detection of individual proteins, peptides and amino acids are described. The study of exhaled breath condensate and exhaled endogenous particles contained lung proteins are considered. The peculiarities of protein, peptide and amino acid content in exhaled breath at various respiratory diseases are described. It is shown that the detectable substances may be specific markers of individual diseases.

  4. Computational protein design quantifies structural constraints on amino acid covariation.

    Directory of Open Access Journals (Sweden)

    Noah Ollikainen

    Full Text Available Amino acid covariation, where the identities of amino acids at different sequence positions are correlated, is a hallmark of naturally occurring proteins. This covariation can arise from multiple factors, including selective pressures for maintaining protein structure, requirements imposed by a specific function, or from phylogenetic sampling bias. Here we employed flexible backbone computational protein design to quantify the extent to which protein structure has constrained amino acid covariation for 40 diverse protein domains. We find significant similarities between the amino acid covariation in alignments of natural protein sequences and sequences optimized for their structures by computational protein design methods. These results indicate that the structural constraints imposed by protein architecture play a dominant role in shaping amino acid covariation and that computational protein design methods can capture these effects. We also find that the similarity between natural and designed covariation is sensitive to the magnitude and mechanism of backbone flexibility used in computational protein design. Our results thus highlight the necessity of including backbone flexibility to correctly model precise details of correlated amino acid changes and give insights into the pressures underlying these correlations.

  5. Solution Structure and Backbone Dynamics of Human Liver Fatty Acid Binding Protein: Fatty Acid Binding Revisited

    OpenAIRE

    Cai, Jun; Lücke, Christian; Chen, Zhongjing; Qiao, Ye; Klimtchuk, Elena; Hamilton, James A.

    2012-01-01

    Liver fatty acid binding protein (L-FABP), a cytosolic protein most abundant in liver, is associated with intracellular transport of fatty acids, nuclear signaling, and regulation of intracellular lipolysis. Among the members of the intracellular lipid binding protein family, L-FABP is of particular interest as it can i), bind two fatty acid molecules simultaneously and ii), accommodate a variety of bulkier physiological ligands such as bilirubin and fatty acyl CoA. To better understand the p...

  6. Redução da expressão astrocitária de proteína glial fibrilar ácida em cães tratados com dexametasona Decrease in the astrocytic expression of glial fibrillary acidic protein in dogs treated with dexamethasone

    Directory of Open Access Journals (Sweden)

    Eduardo Fernandes Bondan

    2013-03-01

    Full Text Available A proteína glial fibrilar ácida (GFAP constitui o principal marcador dos astrócitos, as células gliais mais numerosas do tecido nervoso e que exibem receptores a diversos hormônios esteroidais, os quais exercem aparente influência sobre a expressão gênica das mesmas. O objetivo do presente estudo foi o de avaliar se a administração de dexametasona (DX em protocolos terapêuticos para cães seria capaz de afetar a expressão astrocitária dessa proteína. Para tal, amostras da ponte e da medula espinhal torácica de cães, tratados (n=6 ou não (n=6 com DX, foram submetidas à marcação imuno-histoquímica para a GFAP e a reatividade astrocitária foi determinada por colorimetria em um sistema computacional de análise de imagens. Diferença estatisticamente significativa foi constatada para as médias das áreas marcadas para GFAP na ponte de cães tratados e não-tratados com DX, assim como na medula espinhal torácica dos que haviam recebido previamente o corticoide ou não, com clara tendência, induzida pela droga, de redução da expressão astrocitária da proteína. Além disso, a expressão de GFAP na medula espinhal foi maior que na ponte, independentemente do emprego de DX ou não.Glial fibrillary acidic protein (GFAP is the most important marker of astrocytes, that are the major glial cells in the nervous tissue and exhibit receptors to several steroid hormones, which have an apparent influence in their genic expression. The aim of this study was to evaluate if dexamethasone (DX administration in therapeutic protocols to dogs would be capable of affecting the astrocytic expression of the protein. Samples from the pons and the spinal cord of dogs, treated (n=6 or not (n=6 with DX, were submitted to GFAP immunohistochemical staining and astrocytic reactivity was determined by colorimetry in a computer system for image analysis. Difference statistically significant was noted for the mean areas stained with GFAP in the pons of

  7. Amino acid sequences of proteins from Leptospira serovar pomona

    Directory of Open Access Journals (Sweden)

    Alves Selmo F

    2000-01-01

    Full Text Available This report describes a partial amino acid sequences from three putative outer envelope proteins from Leptospira serovar pomona. In order to obtain internal fragments for protein sequencing, enzymatic and chemical digestion was performed. The enzyme clostripain was used to digest the proteins 32 and 45 kDa. In situ digestion of 40 kDa molecular weight protein was accomplished using cyanogen bromide. The 32 kDa protein generated two fragments, one of 21 kDa and another of 10 kDa that yielded five residues. A fragment of 24 kDa that yielded nineteen residues of amino acids was obtained from 45 kDa protein. A fragment with a molecular weight of 20 kDa, yielding a twenty amino acids sequence from the 40 kDa protein.

  8. A Soluble, Folded Protein without Charged Amino Acid Residues

    DEFF Research Database (Denmark)

    Højgaard, Casper; Kofoed, Christian; Espersen, Roall

    2016-01-01

    Charges are considered an integral part of protein structure and function, enhancing solubility and providing specificity in molecular interactions. We wished to investigate whether charged amino acids are indeed required for protein biogenesis and whether a protein completely free of titratable...... side chains can maintain solubility, stability, and function. As a model, we used a cellulose-binding domain from Cellulomonas fimi, which, among proteins of more than 100 amino acids, presently is the least charged in the Protein Data Bank, with a total of only four titratable residues. We find...

  9. A Soluble, Folded Protein without Charged Amino Acid Residues

    DEFF Research Database (Denmark)

    Højgaard, Casper; Kofoed, Christian; Espersen, Roall;

    2016-01-01

    Charges are considered an integral part of protein structure and function, enhancing solubility and providing specificity in molecular interactions. We wished to investigate whether charged amino acids are indeed required for protein biogenesis and whether a protein completely free of titratable ...

  10. Electron microscopic features of brain edema in rodent cerebral malaria in relation to glial fibrillary acidic protein expression.

    Science.gov (United States)

    Ampawong, Sumate; Chaisri, Urai; Viriyavejakul, Parnpen; Nontprasert, Apichart; Grau, Georges E; Pongponratn, Emsri

    2014-01-01

    The mechanisms leading to cerebral malaria (CM) are not completely understood. Brain edema has been suggested as having an important role in experimental CM. In this study, CBA/CaH mice were infected with Plasmodium berghei ANKA blood-stage and when typical symptoms of CM developed on day 7, brain tissues were processed for electron-microscopic and immunohistochemical studies. The study demonstrated ultrastructural hallmarks of cerebral edema by perivascular edema and astroglial dilatation confirming existing evidence of vasogenic and cytogenic edema. This correlates closely with the clinical features of CM. An adaptive response of astrocytic activity, represented by increasing glial fibrillary acidic protein (GFAP) expression in the perivascular area and increasing numbers of large astrocyte clusters were predominately found in the CM mice. The presence of multivesicular and lamellar bodies indicates the severity of cerebral damage in experimental CM. Congestion of the microvessels with occluded white blood cells (WBCs), parasitized red blood cells (PRBCs) and platelets is also a crucial covariate role for CM pathogenesis.

  11. 40 CFR 79.67 - Glial fibrillary acidic protein assay.

    Science.gov (United States)

    2010-07-01

    ... describes the conduct of a radioimmunoassay for measurement of the amount of GFAP in the brain of vehicle...., the brain, is associated with astrocytic hypertrophy at the site of damage (see O'Callaghan, 1988 in... described by Jahn et al. (1984) in paragraph (e)(2) of this section. Briefly, brain tissue samples from...

  12. Interaction of milk whey protein with common phenolic acids

    Science.gov (United States)

    Zhang, Hao; Yu, Dandan; Sun, Jing; Guo, Huiyuan; Ding, Qingbo; Liu, Ruihai; Ren, Fazheng

    2014-01-01

    Phenolics-rich foods such as fruit juices and coffee are often consumed with milk. In this study, the interactions of α-lactalbumin and β-lactoglobulin with the phenolic acids (chlorogenic acid, caffeic acid, ferulic acid, and coumalic acid) were examined. Fluorescence, CD, and FTIR spectroscopies were used to analyze the binding modes, binding constants, and the effects of complexation on the conformation of whey protein. The results showed that binding constants of each whey protein-phenolic acid interaction ranged from 4 × 105 to 7 × 106 M-n and the number of binding sites n ranged from 1.28 ± 0.13 to 1.54 ± 0.34. Because of these interactions, the conformation of whey protein was altered, with a significant reduction in the amount of α-helix and an increase in the amounts of β-sheet and turn structures.

  13. SIFT: Predicting amino acid changes that affect protein function.

    Science.gov (United States)

    Ng, Pauline C; Henikoff, Steven

    2003-07-01

    Single nucleotide polymorphism (SNP) studies and random mutagenesis projects identify amino acid substitutions in protein-coding regions. Each substitution has the potential to affect protein function. SIFT (Sorting Intolerant From Tolerant) is a program that predicts whether an amino acid substitution affects protein function so that users can prioritize substitutions for further study. We have shown that SIFT can distinguish between functionally neutral and deleterious amino acid changes in mutagenesis studies and on human polymorphisms. SIFT is available at http://blocks.fhcrc.org/sift/SIFT.html.

  14. Informational Way to Protein Alphabet: Entropic Classification of Amino Acids

    CERN Document Server

    Gorban, A N; Popova, T

    2007-01-01

    What are proteins made from, as the working parts of the living cells protein machines? To answer this question, we need a technology to disassemble proteins onto elementary func-tional details and to prepare lumped description of such details. This lumped description might have a multiple material realization (in amino acids). Our hypothesis is that informational approach to this problem is possible. We propose a way of hierarchical classification that makes the primary structure of protein maximally non-random. The first steps of the suggested research program are realized: the method and the analysis of optimal informational protein binary alphabet. The general method is used to answer several specific questions, for example: (i) Is there a syntactic difference between Globular and Membrane proteins? (ii) Are proteins random sequences of amino acids (a long discussion)? For these questions, the answers are as follows: (i) There exists significant syntactic difference between Globular and Membrane proteins,...

  15. Analysis of proteins responsive to acetic acid in Acetobacter: molecular mechanisms conferring acetic acid resistance in acetic acid bacteria.

    Science.gov (United States)

    Nakano, Shigeru; Fukaya, Masahiro

    2008-06-30

    Acetic acid bacteria are used for industrial vinegar production because of their remarkable ability to oxidize ethanol and high resistance to acetic acid. Although several molecular machineries responsible for acetic acid resistance in acetic acid bacteria have been reported, the entire mechanism that confers acetic acid resistance has not been completely understood. One of the promising methods to elucidate the entire mechanism is global analysis of proteins responsive to acetic acid by two-dimensional gel electrophoresis. Recently, two proteins whose production was greatly enhanced by acetic acid in Acetobacter aceti were identified to be aconitase and a putative ABC-transporter, respectively; furthermore, overexpression or disruption of the genes encoding these proteins affected acetic acid resistance in A. aceti, indicating that these proteins are involved in acetic acid resistance. Overexpression of each gene increased acetic acid resistance in Acetobacter, which resulted in an improvement in the productivity of acetic acid fermentation. Taken together, the results of the proteomic analysis and those of previous studies indicate that acetic acid resistance in acetic acid bacteria is conferred by several mechanisms. These findings also provide a clue to breed a strain having high resistance to acetic acid for vinegar fermentation.

  16. Protein packing: dependence on protein size, secondary structure and amino acid composition.

    Science.gov (United States)

    Fleming, P J; Richards, F M

    2000-06-02

    We have used the occluded surface algorithm to estimate the packing of both buried and exposed amino acid residues in protein structures. This method works equally well for buried residues and solvent-exposed residues in contrast to the commonly used Voronoi method that works directly only on buried residues. The atomic packing of individual globular proteins may vary significantly from the average packing of a large data set of globular proteins. Here, we demonstrate that these variations in protein packing are due to a complex combination of protein size, secondary structure composition and amino acid composition. Differences in protein packing are conserved in protein families of similar structure despite significant sequence differences. This conclusion indicates that quality assessments of packing in protein structures should include a consideration of various parameters including the packing of known homologous proteins. Also, modeling of protein structures based on homologous templates should take into account the packing of the template protein structure. Copyright 2000 Academic Press.

  17. Intracerebroventricular administration of Shiga toxin type 2 altered the expression levels of neuronal nitric oxide synthase and glial fibrillary acidic protein in rat brains.

    Science.gov (United States)

    Boccoli, Javier; Loidl, C Fabián; Lopez-Costa, Juan José; Creydt, Virginia Pistone; Ibarra, Cristina; Goldstein, Jorge

    2008-09-16

    Shiga toxin (Stx) from enterohemorrhagic Escherichia coli (STEC) is the main cause of hemorrhagic colitis which may derive into Hemolytic Uremic Syndrome (HUS) and acute encephalopathy, one of the major risk factors for infant death caused by the toxin. We have previously demonstrated that intracerebroventricular administration of Stx2 causes neuronal death and glial cell damage in rat brains. In the present work, we observed that the intracerebroventricular administration of Stx2 increased the expression of glial fibrillary acidic protein (GFAP) leading to astrogliosis. Confocal microscopy showed reactive astrocytes in contact with Stx2-containing neurons. Immunocolocalization of increased GFAP and Stx2 in astrocytes was also observed. This insult in the brain was correlated with changes in the expression and activity of neuronal nitric oxide synthase (nNOS) by using the NADPH-diaphorase histochemical technique (NADPH-d HT). A significant decrease in NOS/NADPH-d-positive neurons and NOS/NADPH-d activity was observed in cerebral cortex and striatum, whereas an opposite effect was found in the hypothalamic paraventricular nucleus. We concluded that the i.c.v. administration of Stx2 promotes a typical pattern of brain injury showing reactive astrocytes and an alteration in the number and activity of nNOS/NADPH-d. According to the functional state of nNOS/NADPH-d and to brain cell morphology data, it could be inferred that the i.c.v. administration of Stx2 leads to either a neurodegenerative or a neuroprotective mechanism in the affected brain areas. The present animal model resembles the encephalopathy developed in Hemolytic Uremic Syndrome (HUS) patients by STEC intoxication.

  18. Protein evolution via amino acid and codon elimination

    DEFF Research Database (Denmark)

    Goltermann, Lise; Larsen, Marie Sofie Yoo; Banerjee, Rajat;

    2010-01-01

    BACKGROUND: Global residue-specific amino acid mutagenesis can provide important biological insight and generate proteins with altered properties, but at the risk of protein misfolding. Further, targeted libraries are usually restricted to a handful of amino acids because there is an exponential...... correlation between the number of residues randomized and the size of the resulting ensemble. Using GFP as the model protein, we present a strategy, termed protein evolution via amino acid and codon elimination, through which simplified, native-like polypeptides encoded by a reduced genetic code were obtained...... simultaneously), while retaining varying levels of activity. Combination of these substitutions to generate a Phe-free variant of GFP abolished fluorescence. Combinatorial re-introduction of five Phe residues, based on the activities of the respective single amino acid replacements, was sufficient to restore GFP...

  19. Modifications of proteins by polyunsaturated fatty acid peroxidation products

    DEFF Research Database (Denmark)

    Refsgaard, Hanne; Tsai, Lin; Stadtman, Earl

    2000-01-01

    The ability of unsaturated fatty acid methyl esters to modify amino acid residues in bovine serum albumin (BSA), glutamine synthetase, and insulin in the presence of a metal-catalyzed oxidation system [ascorbate/Fe(lll)/O-2] depends on the degree of unsaturation of the fatty acid. The fatty acid......-dependent generation of carbonyl groups and loss of lysine residues increased in the order methyl linoleate fatty acids were oxidized in the presence...... in the formation of protein carbonyls, These results are consistent with the proposition that metal-catalyzed oxidation of polyunsaturated fatty acids can contribute to the generation of protein carbonyls by direct interaction of lipid oxidation products (alpha,beta-unsaturated aldehydes) with lysine residues...

  20. Phthalic Acid Chemical Probes Synthesized for Protein-Protein Interaction Analysis

    Directory of Open Access Journals (Sweden)

    Chin-Jen Wu

    2013-06-01

    Full Text Available Plasticizers are additives that are used to increase the flexibility of plastic during manufacturing. However, in injection molding processes, plasticizers cannot be generated with monomers because they can peel off from the plastics into the surrounding environment, water, or food, or become attached to skin. Among the various plasticizers that are used, 1,2-benzenedicarboxylic acid (phthalic acid is a typical precursor to generate phthalates. In addition, phthalic acid is a metabolite of diethylhexyl phthalate (DEHP. According to Gene_Ontology gene/protein database, phthalates can cause genital diseases, cardiotoxicity, hepatotoxicity, nephrotoxicity, etc. In this study, a silanized linker (3-aminopropyl triethoxyslane, APTES was deposited on silicon dioxides (SiO2 particles and phthalate chemical probes were manufactured from phthalic acid and APTES–SiO2. These probes could be used for detecting proteins that targeted phthalic acid and for protein-protein interactions. The phthalic acid chemical probes we produced were incubated with epithelioid cell lysates of normal rat kidney (NRK-52E cells to detect the interactions between phthalic acid and NRK-52E extracted proteins. These chemical probes interacted with a number of chaperones such as protein disulfide-isomerase A6, heat shock proteins, and Serpin H1. Ingenuity Pathways Analysis (IPA software showed that these chemical probes were a practical technique for protein-protein interaction analysis.

  1. Representation of protein-sequence information by amino acid subalphabets

    DEFF Research Database (Denmark)

    Andersen, C.A.F.; Brunak, Søren

    2004-01-01

    -sequence information, using machine learning strategies, where the primary goal is the discovery of novel powerful representations for use in AI techniques. In the case of proteins and the 20 different amino acids they typically contain, it is also a secondary goal to discover how the current selection of amino acids...

  2. Acid stress response and protein induction in Campylobacter jejuni isolates with different acid tolerance

    Directory of Open Access Journals (Sweden)

    Birk Tina

    2012-08-01

    Full Text Available Abstract Background During the transmission route from poultry to the human host, the major foodborne pathogen C. jejuni may experience many types of stresses, including low pH caused by different acids. However, not all strains are equally sensitive to the stresses. The aim of this study was to investigate the response to acid stress of three sequenced C. jejuni strains with different acid tolerances using HCl and acetic acid. Results Two-dimensional gel electrophoresis was used for proteomic analysis and proteins were radioactively labelled with methionine to identify proteins only related to acid exposure. To allow added radioactive methionine to be incorporated into induced proteins, a modified chemically defined broth was developed with the minimal amount of methionine necessary for satisfactory growth of all strains. Protein spots were analyzed using image software and identification was done with MALDI-TOF-TOF. The most acid-sensitive isolate was C. jejuni 327, followed by NCTC 11168 and isolate 305 as the most tolerant. Overall, induction of five proteins was observed within the pI range investigated: 19 kDa periplasmic protein (p19, thioredoxin-disulfide (TrxB, a hypothetical protein Cj0706 (Cj0706, molybdenum cofactor biosynthesis protein (MogA, and bacterioferritin (Dps. Strain and acid type dependent differences in the level of response were observed. For strain NCTC 11168, the induced proteins and the regulator fur were analysed at the transcriptomic level using qRT-PCR. In this transcriptomic analysis, only up-regulation of trxB and p19 was observed. Conclusions A defined medium that supports the growth of a range of Campylobacter strains and suitable for proteomic analysis was developed. Mainly proteins normally involved in iron control and oxidative stress defence were induced during acid stress of C. jejuni. Both strain and acid type affected sensitivity and response.

  3. Predicting nucleic acid binding interfaces from structural models of proteins.

    Science.gov (United States)

    Dror, Iris; Shazman, Shula; Mukherjee, Srayanta; Zhang, Yang; Glaser, Fabian; Mandel-Gutfreund, Yael

    2012-02-01

    The function of DNA- and RNA-binding proteins can be inferred from the characterization and accurate prediction of their binding interfaces. However, the main pitfall of various structure-based methods for predicting nucleic acid binding function is that they are all limited to a relatively small number of proteins for which high-resolution three-dimensional structures are available. In this study, we developed a pipeline for extracting functional electrostatic patches from surfaces of protein structural models, obtained using the I-TASSER protein structure predictor. The largest positive patches are extracted from the protein surface using the patchfinder algorithm. We show that functional electrostatic patches extracted from an ensemble of structural models highly overlap the patches extracted from high-resolution structures. Furthermore, by testing our pipeline on a set of 55 known nucleic acid binding proteins for which I-TASSER produces high-quality models, we show that the method accurately identifies the nucleic acids binding interface on structural models of proteins. Employing a combined patch approach we show that patches extracted from an ensemble of models better predicts the real nucleic acid binding interfaces compared with patches extracted from independent models. Overall, these results suggest that combining information from a collection of low-resolution structural models could be a valuable approach for functional annotation. We suggest that our method will be further applicable for predicting other functional surfaces of proteins with unknown structure. Copyright © 2011 Wiley Periodicals, Inc.

  4. Mitotic apparatus: the selective extraction of protein with mild acid.

    Science.gov (United States)

    Bibring, T; Baxandall, J

    1968-07-26

    The treatment of isolated mitotic apparatus with mild (pH 3) hydrochloric acid results in the extraction of less than 10 percent of its protein, accompanied by the selective morphological disappearance of the microtubules. The same extraction can be shown to dissolve outer doublet microtubules from sperm flagella. A protein with points of similarity to the flagellar microtubule protein is the major component of the extract from mitotic apparatus.

  5. Modifications of proteins by polyunsaturated fatty acid peroxidation products

    Science.gov (United States)

    Refsgaard, Hanne H. F.; Tsai, Lin; Stadtman, Earl R.

    2000-01-01

    The ability of unsaturated fatty acid methyl esters to modify amino acid residues in bovine serum albumin (BSA), glutamine synthetase, and insulin in the presence of a metal-catalyzed oxidation system [ascorbate/Fe(III)/O2] depends on the degree of unsaturation of the fatty acid. The fatty acid-dependent generation of carbonyl groups and loss of lysine residues increased in the order methyl linoleate < methyl linolenate < methyl arachidonate. The amounts of alkyl hydroperoxides, malondialdehyde, and a number of other aldehydes that accumulated when polyunsaturated fatty acids were oxidized in the presence of BSA were significantly lower than that observed in the absence of BSA. Direct treatment of proteins with various lipid hydroperoxides led to a slight increase in the formation of protein carbonyl derivatives, whereas treatment with the hydroperoxides together with Fe(II) led to a substantial increase in the formation of protein carbonyls. These results are consistent with the proposition that metal-catalyzed oxidation of polyunsaturated fatty acids can contribute to the generation of protein carbonyls by direct interaction of lipid oxidation products (α,β-unsaturated aldehydes) with lysine residues (Michael addition reactions) and also by interactions with alkoxyl radicals obtained by Fe(II) cleavage of lipid hydroperoxides that are formed. In addition, saturated aldehydes derived from the polyunsaturated fatty acids likely react with lysine residues to form Schiff base adducts.

  6. Manipulating Fatty Acid Biosynthesis in Microalgae for Biofuel through Protein-Protein Interactions

    Science.gov (United States)

    Blatti, Jillian L.; Beld, Joris; Behnke, Craig A.; Mendez, Michael; Mayfield, Stephen P.; Burkart, Michael D.

    2012-01-01

    Microalgae are a promising feedstock for renewable fuels, and algal metabolic engineering can lead to crop improvement, thus accelerating the development of commercially viable biodiesel production from algae biomass. We demonstrate that protein-protein interactions between the fatty acid acyl carrier protein (ACP) and thioesterase (TE) govern fatty acid hydrolysis within the algal chloroplast. Using green microalga Chlamydomonas reinhardtii (Cr) as a model, a structural simulation of docking CrACP to CrTE identifies a protein-protein recognition surface between the two domains. A virtual screen reveals plant TEs with similar in silico binding to CrACP. Employing an activity-based crosslinking probe designed to selectively trap transient protein-protein interactions between the TE and ACP, we demonstrate in vitro that CrTE must functionally interact with CrACP to release fatty acids, while TEs of vascular plants show no mechanistic crosslinking to CrACP. This is recapitulated in vivo, where overproduction of the endogenous CrTE increased levels of short-chain fatty acids and engineering plant TEs into the C. reinhardtii chloroplast did not alter the fatty acid profile. These findings highlight the critical role of protein-protein interactions in manipulating fatty acid biosynthesis for algae biofuel engineering as illuminated by activity-based probes. PMID:23028438

  7. Manipulating fatty acid biosynthesis in microalgae for biofuel through protein-protein interactions.

    Directory of Open Access Journals (Sweden)

    Jillian L Blatti

    Full Text Available Microalgae are a promising feedstock for renewable fuels, and algal metabolic engineering can lead to crop improvement, thus accelerating the development of commercially viable biodiesel production from algae biomass. We demonstrate that protein-protein interactions between the fatty acid acyl carrier protein (ACP and thioesterase (TE govern fatty acid hydrolysis within the algal chloroplast. Using green microalga Chlamydomonas reinhardtii (Cr as a model, a structural simulation of docking CrACP to CrTE identifies a protein-protein recognition surface between the two domains. A virtual screen reveals plant TEs with similar in silico binding to CrACP. Employing an activity-based crosslinking probe designed to selectively trap transient protein-protein interactions between the TE and ACP, we demonstrate in vitro that CrTE must functionally interact with CrACP to release fatty acids, while TEs of vascular plants show no mechanistic crosslinking to CrACP. This is recapitulated in vivo, where overproduction of the endogenous CrTE increased levels of short-chain fatty acids and engineering plant TEs into the C. reinhardtii chloroplast did not alter the fatty acid profile. These findings highlight the critical role of protein-protein interactions in manipulating fatty acid biosynthesis for algae biofuel engineering as illuminated by activity-based probes.

  8. Macromolecular mimicry of nucleic acid and protein

    DEFF Research Database (Denmark)

    Nautrup Pedersen, Gitte; Nyborg, Jens; Clark, Brian F

    1999-01-01

    of the concept of macromolecular mimicry. Macromolecular mimicry has further been proposed among initiation and release factors, thereby adding a new element to the description of protein synthesis in bacteria. Such mimicry has also been observed in other biological processes such as autoimmunity, DNA repair......, and gene regulation, at both transcriptional and translational levels. Udgivelsesdato: 1999-Jul...

  9. Nucleic acid compositions and the encoding proteins

    Science.gov (United States)

    Preston, III, James F.; Chow, Virginia; Nong, Guang; Rice, John D.; St. John, Franz J.

    2014-09-02

    The subject invention provides at least one nucleic acid sequence encoding an aldouronate-utilization regulon isolated from Paenibacillus sp. strain JDR-2, a bacterium which efficiently utilizes xylan and metabolizes aldouronates (methylglucuronoxylosaccharides). The subject invention also provides a means for providing a coordinately regulated process in which xylan depolymerization and product assimilation are coupled in Paenibacillus sp. strain JDR-2 to provide a favorable system for the conversion of lignocellulosic biomass to biobased products. Additionally, the nucleic acid sequences encoding the aldouronate-utilization regulon can be used to transform other bacteria to form organisms capable of producing a desired product (e.g., ethanol, 1-butanol, acetoin, 2,3-butanediol, 1,3-propanediol, succinate, lactate, acetate, malate or alanine) from lignocellulosic biomass.

  10. Non-protein amino acids in peptide design

    Indian Academy of Sciences (India)

    S Aravinda; N Shamala; Rituparna S Roy; P Balaram

    2003-10-01

    An overview of the use of non-protein amino acids in the design of conformationally well-defined peptides, based on work from the author’s laboratory, is discussed. The crystal structures of several designed oligopeptides illustrate the use -aminoisobutyric acid (Aib) in the construction of helices, D-amino acids in the design of helix termination segments and DPro-Xxx segments for nucleating of -hairpin structures. - and -amino acid residues have been used to expand the range of designed polypeptide structures.

  11. Circuit topology of proteins and nucleic acids.

    Science.gov (United States)

    Mashaghi, Alireza; van Wijk, Roeland J; Tans, Sander J

    2014-09-02

    Folded biomolecules display a bewildering structural complexity and diversity. They have therefore been analyzed in terms of generic topological features. For instance, folded proteins may be knotted, have beta-strands arranged into a Greek-key motif, or display high contact order. In this perspective, we present a method to formally describe the topology of all folded linear chains and hence provide a general classification and analysis framework for a range of biomolecules. Moreover, by identifying the fundamental rules that intrachain contacts must obey, the method establishes the topological constraints of folded linear chains. We also briefly illustrate how this circuit topology notion can be applied to study the equivalence of folded chains, the engineering of artificial RNA structures and DNA origami, the topological structure of genomes, and the role of topology in protein folding.

  12. Reduction in the number of astrocytes and their projections is associated with increased synaptic protein density in the hypothalamus of poorly controlled diabetic rats.

    Science.gov (United States)

    Lechuga-Sancho, Alfonso M; Arroba, Ana I; Frago, Laura M; García-Cáceres, Cristina; de Célix, Arancha Delgado-Rubín; Argente, Jesús; Chowen, Julie A

    2006-11-01

    Processes under hypothalamic control, such as thermogenesis, feeding behavior, and pituitary hormone secretion, are disrupted in poorly controlled diabetes, but the underlying mechanisms are poorly understood. Because glial cells regulate neurosecretory neurons through modulation of synaptic inputs and function, we investigated the changes in hypothalamic glia in rats with streptozotocin-induced diabetes mellitus. Hypothalamic glial fibrillary acidic protein (GFAP) levels decreased significantly 6 wk after diabetes onset. This was coincident with decreased GFAP immunoreactive surface area, astrocyte number, and the extension of GFAP immunoreactive processes/astrocyte in the arcuate nucleus. Cell death, analyzed by terminal deoxyuridine 5-triphosphate nick-end labeling and ELISA, increased significantly at 4 wk of diabetes. Proliferation, measured by Western blot for proliferating cell nuclear antigen and immunostaining for phosphorylated histone H-3, decreased in the hypothalamus of diabetic rats throughout the study, becoming significantly reduced by 8 wk. Both proliferation and death affected astroctyes because both phosphorylated histone H-3- and terminal deoxyuridine 5-triphosphate nick-end labeling-labeled cells were GFAP positive. Western blot analysis revealed that postsynaptic density protein 95 and the presynaptic proteins synapsin I and synaptotagmin increased significantly at 8 wk of diabetes, suggesting increased hypothalamic synaptic density. Thus, in poorly controlled diabetic rats, there is a decrease in the number of hypothalamic astrocytes that is correlated with modifications in synaptic proteins and possibly synaptic inputs. These morphological changes in the arcuate nucleus could be involved in neurosecretory and metabolic changes seen in diabetic animals.

  13. Okadaic acid: the archetypal serine/threonine protein phosphatase inhibitor.

    Science.gov (United States)

    Dounay, A B; Forsyth, C J

    2002-11-01

    As the first recognized member of the "okadaic acid class" of phosphatase inhibitors, the marine natural product okadaic acid is perhaps the most well-known member of a diverse array of secondary metabolites that have emerged as valuable probes for studying the roles of various cellular protein serine/threonine phosphatases. This review provides a historical perspective on the role that okadaic acid has played in stimulating a broad spectrum of modern scientific research as a result of the natural product's ability to bind to and inhibit important classes of protein serine / threonine phosphatases. The relationships between the structure and biological activities of okadaic acid are briefly reviewed, as well as the structural information regarding the particular cellular receptors protein phosphatases 1 (PP1) and 2A. Laboratory syntheses of okadaic acid and its analogs are thoroughly reviewed. Finally, an interpretation of the critical contacts observed between okadaic acid and PP1 by X-ray crystallography is provided, and specific molecular recognition hypotheses that are testable via the synthesis and assay of non-natural analogs of okadaic acid are suggested.

  14. Acid stress response and protein induction in Campylobacter jejuni isolates with different acid tolerance

    DEFF Research Database (Denmark)

    Birk, Tina; Wik, Monica Takamiya; Lametsch, René

    2012-01-01

    BACKGROUND: During the transmission route from poultry to the human host, the major foodborne pathogen C. jejuni may experience many types of stresses, including low pH caused by different acids. However, not all strains are equally sensitive to the stresses. The aim of this study was to investig......BACKGROUND: During the transmission route from poultry to the human host, the major foodborne pathogen C. jejuni may experience many types of stresses, including low pH caused by different acids. However, not all strains are equally sensitive to the stresses. The aim of this study...... was to investigate the response to acid stress of three sequenced C. jejuni strains with different acid tolerances using HCl and acetic acid. RESULTS: Two-dimensional gel electrophoresis was used for proteomic analysis and proteins were radioactively labelled with methionine to identify proteins only related to acid...... with MALDI-TOF-TOF. The most acid-sensitive isolate was C. jejuni 327, followed by NCTC 11168 and isolate 305 as the most tolerant. Overall, induction of five proteins was observed within the pI range investigated: 19 kDa periplasmic protein (p19), thioredoxin-disulfide (TrxB), a hypothetical protein Cj0706...

  15. [Fractional and amino acid composition of krill proteins and the potential for obtaining protein preparations].

    Science.gov (United States)

    Orlova, T A; Churina, E E; Kuranova, L K

    1985-01-01

    Studies of the fractional composition of krill proteins demonstrated that the content of protein fractions changes depending on the time of krill catch. The highest amount of water-soluble proteins is contained by krill caught in December (64%), of salt-soluble by krill caught in June (12%), base-soluble by krill caught in May, September and February (34%). Krill protein contains from 50 to 60% of water- and salt-soluble fractions. Analysis of the amino acid composition of krill proteins showed that it does not differ essentially from that of adequate food proteins.

  16. Comparative proteomic analysis of differentially expressed proteins in β-aminobutyric acid enhanced Arabidopsis thaliana tolerance to simulated acid rain.

    Science.gov (United States)

    Liu, Tingwu; Jiang, Xinwu; Shi, Wuliang; Chen, Juan; Pei, Zhenming; Zheng, Hailei

    2011-05-01

    Acid rain is a worldwide environmental issue that has seriously destroyed forest ecosystems. As a highly effective and broad-spectrum plant resistance-inducing agent, β-aminobutyric acid could elevate the tolerance of Arabidopsis when subjected to simulated acid rain. Using comparative proteomic strategies, we analyzed 203 significantly varied proteins of which 175 proteins were identified responding to β-aminobutyric acid in the absence and presence of simulated acid rain. They could be divided into ten groups according to their biological functions. Among them, the majority was cell rescue, development and defense-related proteins, followed by transcription, protein synthesis, folding, modification and destination-associated proteins. Our conclusion is β-aminobutyric acid can lead to a large-scale primary metabolism change and simultaneously activate antioxidant system and salicylic acid, jasmonic acid, abscisic acid signaling pathways. In addition, β-aminobutyric acid can reinforce physical barriers to defend simulated acid rain stress.

  17. Marcação imunoistoquímica da expressão astrocitária de proteína glial fibrilar ácida e de vimentina no sistema nervoso central de cães com cinomose Immunohistochemical staining of the astrocytic expression of glial fibrillary acidic protein and vimentin in the central nervous system of dogs with canine distemper

    Directory of Open Access Journals (Sweden)

    Heloísa Orsini

    2007-12-01

    immunohistochemical staining of two astrocytic proteins - glial fibrillary acidic protein (GFAP and vimentin (VIM -, comparing samples of cerebellum and brainstem from eight dogs with canine distemper and from two healthy dogs, from different breeds and ages varying from 1 to 4 years old. Histological sections were submitted to the avidin-biotin-peroxidase indirect method of immmunohistochemical staining (ABC and the astrocytic reactivity, observed in light microscopy, was quantified in a computer system for image analysis. It was possible to notice, on most of the sections from sick animals, degenerative lesions that indicate demyelination. The immunostaining for GFAP and VIM was more intense on animals with canine distemper, specially around the ventricules and near degenerated sites. There was no significant difference between the immunostaining (GFAP and VIM of animals with canine distemper with and without inflammatory infiltrate of the cerebellar white matter. The increased immunoreactivity of astrocytes for GFAP and the VIM reexpression in injured areas indicate the astrocytic involvement on nervous tissue response to the demyelinating lesions induced by the canine distemper virus (CDV in the CNS.

  18. Protein and amino acid quality of meat and bone meal.

    Science.gov (United States)

    Parsons, C M; Castanon, F; Han, Y

    1997-02-01

    The in vivo protein quality of 14 meat and bone meals (MBM) was evaluated in three chick growth assays and a 48-h excreta collection assay using conventional and cecectomized roosters. In addition, in vitro evaluation of protein quality was assessed using pepsin N digestibility (0.2, 0.002, or 0.0002% pepsin), KOH protein solubility, and multi-enzyme pH change. Crude protein, lysine, and SAA in the MBM varied from 48 to 56, 2.32 to 3.01, and 1.0 to 2.13%, respectively. Protein efficiency ratio (weight gain:protein intake) estimated from feeding chicks diets containing 9% protein from a MBM ranged from 0.61 to 2.89 and averaged 1.78. Lysine bioavailability determined by slope-ratio chick assay ranged from 43 to 89%. True amino acid digestibility and TMEn values determined in cecectomized roosters were generally lower (P < 0.05) than those determined in conventional roosters. True digestibility of amino acids (percentage) also varied among MBM, with the mean (and range) for lysine, methionine, and cystine in cecectomized birds being 81 (73 to 88), 85 (77 to 91), and 58% (37 to 72%), respectively. Pepsin N digestibility values determined using 0.002 or 0.0002% pepsin were positively correlated (P < 0.05) with lysine digestibility. Pepsin N digestibility determined using 0.2% pepsin, KOH protein solubility, and multi-enzyme pH change were not significantly correlated with in vivo protein quality. Ash content was negatively correlated (-0.80, P < 0.05) with protein efficiency ratio. These results indicated that there is substantial variation in protein quality among commercial MBM and that pepsin N digestibility and ash content are correlated with some in vivo protein quality measurements.

  19. (-)-Hydroxycitric Acid Nourishes Protein Synthesis via Altering Metabolic Directions of Amino Acids in Male Rats.

    Science.gov (United States)

    Han, Ningning; Li, Longlong; Peng, Mengling; Ma, Haitian

    2016-08-01

    (-)-Hydroxycitric acid (HCA), a major active ingredient of Garcinia Cambogia extracts, had shown to suppress body weight gain and fat accumulation in animals and humans. While, the underlying mechanism of (-)-HCA has not fully understood. Thus, this study was aimed to investigate the effects of long-term supplement with (-)-HCA on body weight gain and variances of amino acid content in rats. Results showed that (-)-HCA treatment reduced body weight gain and increased feed conversion ratio in rats. The content of hepatic glycogen, muscle glycogen, and serum T4 , T3 , insulin, and Leptin were increased in (-)-HCA treatment groups. Protein content in liver and muscle were significantly increased in (-)-HCA treatment groups. Amino acid profile analysis indicated that most of amino acid contents in serum and liver, especially aromatic amino acid and branched amino acid, were higher in (-)-HCA treatment groups. However, most of the amino acid contents in muscle, especially aromatic amino acid and branched amino acid, were reduced in (-)-HCA treatment groups. These results indicated that (-)-HCA treatment could reduce body weight gain through promoting energy expenditure via regulation of thyroid hormone levels. In addition, (-)-HCA treatment could promote protein synthesis by altering the metabolic directions of amino acids. Copyright © 2016 John Wiley & Sons, Ltd.

  20. The interaction of amino acids, peptides, and proteins with DNA.

    Science.gov (United States)

    Solovyev, Andrey Y; Tarnovskaya, Svetlana I; Chernova, Irina A; Shataeva, Larisa K; Skorik, Yury A

    2015-01-01

    Amino acids that carry charges on their side groups can bind to double stranded DNA (dsDNA) and change the strength of the double helix. Measurement of the DNA melting temperature (Tm) confirmed that acidic amino acids (Glu, Asp) weaken the H-bonds between DNA strands, whereas basic amino acids (Arg, Lys) strengthen the interaction between the strands. A rank correlation exists between the amino acid isoelectric points and the observed changes in Tm. A similar dependence of the hyperchromic effect on the isoelectric point of a protein (pepsin, insulin, cortexin, and protamine) was observed for DNA-protein complexes at room temperature. Short peptides (KE, AEDG, and KEDP) containing a mixture of acidic and basic amino acid residues also affect Tm and the stability of the double helix. A model for binding Glu and Lys to dsDNA was explored by a docking simulation. The model shows that Glu, in an untwisted shape, binds to dsDNA in its major groove and disrupts three H-bonds between the strands, thereby destabilizing the double helix. Lys, in an untwisted shape, binds to the external side of the dsDNA and forms two bonds with O atoms of neighboring phosphodiester groups, thereby strengthening the DNA helix.

  1. 脑卒中患者血清胶质纤维酸性蛋白的变化观察%Observation of change of serum glial fibrillary acidic protein level in patients with acute hemorrhagic stroke

    Institute of Scientific and Technical Information of China (English)

    梁建伟; 崔桂萍; 杨萍; 张葳

    2011-01-01

    目的 测定出血性脑卒中患者血清中胶质纤维酸性蛋白(GFAP)的动态变化,探讨其与神经功能缺损程度评分(MESSS)、预后Barthel指数(BI)以及脑出血量和瘫痪程度的关系.方法 采用酶联免疫吸附试验(ELISA)检测出血性脑卒中患者发病后第1天、第3天和第14天血清GFAP水平.所有患者在相应的时间点进行MESSS评分,并在出院时评价BI.结果 患者发病后第1天、第3天和第14天血清GFAP水平分别为(9.22±3.65)、(8.18±3.52)、(8.45±3.77) ng/mL,均显著高于对照组[(4.62±1.56)ng/mL](P<0.01).第1天和第3天血清GFAP水平分别与相应时间的MESSS评分呈正相关[相关系数(r) =0.696,P<0.01;r=0.352,P<0.05].第1天和第3天血清GFAP水平分别与相应时间的出血量呈正相关(r=0.520,P <0.01;r =0.440,P<0.05).而第14天血清GFAP水平与出院时BI呈负相关(r=-0.431,P<0.05).第1天血清GFAP水平与瘫痪程度呈正相关(r =0.462,P<0.05).当临界值(Cut-off)为6.021 ng/mL时,ELISA的灵敏度为78.6%,特异度为80.0%.结论 出血性脑卒中患者血清GFAP水平显著升高,有望成为出血性脑卒中患者早期病情诊断和预后评估的指标.%Objective To determine the dynamic change of serum glial fibrillary acidic protein (GFAP) level in patients with acute hemorrhagic stroke, and investigate the relationships with modified Edinburgh-Scandinavian stroke scale ( MESSS) , Barihel index ( BI) , volume of haematoma and degree of paralysis. Methods The serum levels of GFAP in patients with acute hemorrhagic stroke at the 1st d, 3rd d and 14th d were determined by enzyme-linked immunosorbent assay ( ELISA). The status of all patients was evaluated by MESSS at the corresponding time points, and BI was evaluated at discharge from the hospital. Results For all patients observed, serum GFAP levels at the 1st d [(9. 22 ±3.65) ng/mL], 3rd d [ (8. 18 ±3. 52) ng/mL]and the 14th d [ (8. 45 ±3. 77)n^/mL] after onset of stroke

  2. Suppression of muscle protein turnover and amino acid degradation by dietary protein deficiency

    Science.gov (United States)

    Tawa, N. E. Jr; Goldberg, A. L.

    1992-01-01

    To define the adaptations that conserve amino acids and muscle protein when dietary protein intake is inadequate, rats (60-70 g final wt) were fed a normal or protein-deficient (PD) diet (18 or 1% lactalbumin), and their muscles were studied in vitro. After 7 days on the PD diet, both protein degradation and synthesis fell 30-40% in skeletal muscles and atria. This fall in proteolysis did not result from reduced amino acid supply to the muscle and preceded any clear decrease in plasma amino acids. Oxidation of branched-chain amino acids, glutamine and alanine synthesis, and uptake of alpha-aminoisobutyrate also fell by 30-50% in muscles and adipose tissue of PD rats. After 1 day on the PD diet, muscle protein synthesis and amino acid uptake decreased by 25-40%, and after 3 days proteolysis and leucine oxidation fell 30-45%. Upon refeeding with the normal diet, protein synthesis also rose more rapidly (+30% by 1 day) than proteolysis, which increased significantly after 3 days (+60%). These different time courses suggest distinct endocrine signals for these responses. The high rate of protein synthesis and low rate of proteolysis during the first 3 days of refeeding a normal diet to PD rats contributes to the rapid weight gain ("catch-up growth") of such animals.

  3. Induction of DNA damage by oxidised amino acids and proteins

    DEFF Research Database (Denmark)

    Luxford, Catherine; Dean, Roger T; Davies, Michael Jonathan

    2002-01-01

    Exposure of amino acids, peptides and proteins to radicals in the presence of O2 generates hydroperoxides in a dose-dependent manner. These hydroperoxides are stable in the absence of exogenous catalysts (e.g. heat, light, redox-active transition metal ions), but decompose rapidly in the presence...

  4. Amino acid nutrition beyond methionine and lysine for milk protein

    Science.gov (United States)

    Amino acids are involved in many important physiological processes affecting the production, health, and reproduction of high-producing dairy cows. Most research and recommendations for lactating dairy cows has focused on methionine and lysine for increasing milk protein yield. This is because these...

  5. Assessment of Kerch Bay environmental pollution using neuroglial proteins of ground fish

    Directory of Open Access Journals (Sweden)

    H. V. Sukharenko

    2014-03-01

    Full Text Available The modern ecology situation in waters of the Kerch Strait requires assessment of disturbances in biotopes and monitoring of the degree of impact of industrial pollutants on ecosystem. Deposit of oil products after the 2007 year ships’ accidents might have considerable impact on the water biocenosis area. The investigation of cytoskeleton marker of astrocytes glial fibrillary acidic protein (GFAP in brain of the bullhead (Neogobius fluviatilis, which is the typical representative of the commercial ground fish of the Kerch Strait, has been carried out. The results of comparative analysis of GFAP content in the brain of fish from the Kerch Bay near-shore waters and fish from conditionally clear area of Vorskla river shows the reliable (2.18 times increasing of GFAP in the area of industrial pollution. Rising GFAP content indicates the astrogliosis development as a result of metabolic disturbances which can be induced by higher content of oil products in the near-bottom biotopes of the Kerch Bay. Increase in lipid peroxidation level was observed in the brain of fish from the Kerch Bay. The results provided with regard to violations of the state of astrocyte cytoskeleton and oxidative stress in the brain of bullhead from the Kerch Bay prove the sublethal biology effect of industrial pollutants in hydrobionts from this area. Results of this investigation also indicate the necessity of continuous ecology monitoring and comprehensive study of hydrobiont populations in the industrial regions and ecological disaster zones.

  6. 脊髓源星形胶质细胞胶质原纤维酸性蛋白表达抑制最佳短发夹样RNA分子筛选及真核表达载体构建%Optimal screening of short hairpin and construction of its eukaryotic expression vector for glial fibrillary acidic protein

    Institute of Scientific and Technical Information of China (English)

    高明勇; 李振宇; 闫洪印

    2007-01-01

    BACKGROUND: The glial scar formation after spinal cord injury in mammals is the physical and chemical barriers for neural regeneration, and relieving or delaying glial scar formation can provide benefit conditions for the regeneration of injured spinal cord.OBJECTIVE: To design and screen short hairpin RNA (shRNA) interfere molecular targeting the gene coding region of glial fibrillary acidic protein (GFAP) in rat, and reconstruct the eukaryotic vector of shRNA.DESIGN: An observational animal experiment.SETTING: Department of Spine Surgery, Shenzhen Hospital affiliated to Southern Medical University.MATERIALS: Twenty-five Wistar rats of clean degree, either male or female, weighing 20-25 g, were used. DMEM/F12,lipofectamine2000, Trizol RNA separating kits); fetal bovine serum (Hyclone); BamH Ⅰ, HindⅢ, Pstl, Sail and T4 ligases;Plasmid mini preparation kit and DNA gel extraction kit.METHODS: The experiments were carried out in the Shenzhen Hospital affiliated to Southern Medical University from October 2005 to June 2006. Three pairs of shRNA template which composed of 19 bp reverse repeated motif of GFAP target sequence with 9 bp spacer were designed and synthesized, then they were inserted directionally into plasmid Psilencer 2.1 respectively to generate small interfering RNA (siRNA) eukaryotic expression vector. ShRNA molecules were transfected by liposome via ex vivo expression repressive model of GFAP of rat spinal astrocytes. The effects of expressive suppression were detected with real-time fluorescence quantitative reverse transcription-polymerase chain reaction (RT-PCR) and Western blot, and then the optimal shRNA eukaryotic vector of repressive expression of GFAP was screened.MAIN OUTCOME MEASURES: ① Interfering sequence specific shRNA template synthesis; ② Constructing specific recombinant plasmid eukaryotic expression vector. ③ Culturing rat spinal astrocytes in vitro; ④ Effects of expressive suppression on GFAP in primary astrocytes after RNA

  7. Protein and amino acid metabolism in skeletal muscle

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Guoyao.

    1989-01-01

    Isolated chick extensor digitorum communis (EDC) muscles and, in some experiments, rat skeletal muscles were used to study a number of aspects of protein and amino acid metabolism. (1) Chick EDC muscles synthesize and release large amounts of alanine and glutamine, which indirectly obtain their amino groups from branched-chain amino acids (BCAA). (2) Acetoacetate or DL-{beta}-hydroxybutyrate (4 mM) decrease (P < 0.01) alanine synthesis and BCAA transamination in EDC muscles from 24-h fasted chicks by decreasing (P < 0.01) intracellular concentrations of pyruvate due to inhibition of glycolysis. (3) Glutamine is extensively degraded in skeletal muscles from both chicks and rats, thus challenging the traditional view that glutamine oxidation is negligible in skeletal muscle. The cytosolic glutamine aminotransferases L and K in the rat and the mitochondrial phosphate-activated glutaminase in the chick play important roles in the conversion of glutamine to {alpha}-ketoglutarate for further oxidation. (4) Although methionine has been reported to be extensively transaminated in rat skeletal muscle preparations in the absence of other amino acids, transamination of methionine is absent or negligible in chick and rat skeletal muscles in the presence of physiological concentrations of amino acids. (5) Glutamine at 1.0-15 mM increases (P < 0.01) protein synthesis ({sup 3}H-phenylalanine incorporation), and at 10.0-15.0 mM decreases (P < 0.05) protein degradation ({sup 3}H-phenylalanine release from prelabelled protein in vivo) in EDC muscles from fed chicks as compared to muscles incubated in the absence of glutamine. (6) Acetoacetate or DL-{beta}-hydroxybutyrate (4 mM) has a small but significant inhibitory effect (P < 0.05) on the rate of protein synthesis, but has no effect (P > 0.05) on the rate of protein degradation in EDC muscles from fed chicks.

  8. Effect of Acupuncture on Contents of Tyrosine Hydroxylase and Glial Fibrillary Acidic Protein in Ventral Tegmental Area of Heroin Self-administrating Rats

    Institute of Scientific and Technical Information of China (English)

    ZHU Zhong-chun; HU Jun; XU Ping

    2006-01-01

    ,and electro-acupuncture group which were also withdrawn from heroin for 1 week (group E, n= 6) and for 2 weeks (group F, n= 6), during which time they were given electro-acupuncture treatment for 20 min daily and then returned to their individual home cages; in the meantime, another 6 rats were trained by nose-poking response with saline for 14 days as control (group A); Then the levels of tyrosine hydroxylase (TH) and glial fibrillary acidic protein (GFAP) in VTA, Nac, Amy, PFC, SN, Cpu were detected with immunohistochemistry method. Results: The leveks of TH and GFAP in VTA of the heroin self-administrating rats were obviously increased, and the levels of TH and GFAP in Nac were also decreased, and these changes were not found in SN, Cpu, Amy and PFC; Electro-acupuncture could promote the up-regulation of TH and GFAP in VTA and down-regulation of TH and GFAP in Nac to return to the normal level. Conclusions: The chronic heroin self-administration produced some biochemical adaptations in the rdated brain regions of the mesolimbic dopamine system and electroacupuncture could promote the repair of the "injured" DA neurons in VTA of heroin addicted rats and their functional recovery.

  9. Prediction of protein disorder on amino acid substitutions.

    Science.gov (United States)

    Anoosha, P; Sakthivel, R; Gromiha, M Michael

    2015-12-15

    Intrinsically disordered regions of proteins are known to have many functional roles in cell signaling and regulatory pathways. The altered expression of these proteins due to mutations is associated with various diseases. Currently, most of the available methods focus on predicting the disordered proteins or the disordered regions in a protein. On the other hand, methods developed for predicting protein disorder on mutation showed a poor performance with a maximum accuracy of 70%. Hence, in this work, we have developed a novel method to classify the disorder-related amino acid substitutions using amino acid properties, substitution matrices, and the effect of neighboring residues that showed an accuracy of 90.0% with a sensitivity and specificity of 94.9 and 80.6%, respectively, in 10-fold cross-validation. The method was evaluated with a test set of 20% data using 10 iterations, which showed an average accuracy of 88.9%. Furthermore, we systematically analyzed the features responsible for the better performance of our method and observed that neighboring residues play an important role in defining the disorder of a given residue in a protein sequence. We have developed a prediction server to identify disorder-related mutations, and it is available at http://www.iitm.ac.in/bioinfo/DIM_Pred/. Copyright © 2015 Elsevier Inc. All rights reserved.

  10. A conserved patch of hydrophobic amino acids modulates Myb activity by mediating protein-protein interactions.

    Science.gov (United States)

    Dukare, Sandeep; Klempnauer, Karl-Heinz

    2016-07-01

    The transcription factor c-Myb plays a key role in the control of proliferation and differentiation in hematopoietic progenitor cells and has been implicated in the development of leukemia and certain non-hematopoietic tumors. c-Myb activity is highly dependent on the interaction with the coactivator p300 which is mediated by the transactivation domain of c-Myb and the KIX domain of p300. We have previously observed that conservative valine-to-isoleucine amino acid substitutions in a conserved stretch of hydrophobic amino acids have a profound effect on Myb activity. Here, we have explored the function of the hydrophobic region as a mediator of protein-protein interactions. We show that the hydrophobic region facilitates Myb self-interaction and binding of the histone acetyl transferase Tip60, a previously identified Myb interacting protein. We show that these interactions are affected by the valine-to-isoleucine amino acid substitutions and suppress Myb activity by interfering with the interaction of Myb and the KIX domain of p300. Taken together, our work identifies the hydrophobic region in the Myb transactivation domain as a binding site for homo- and heteromeric protein interactions and leads to a picture of the c-Myb transactivation domain as a composite protein binding region that facilitates interdependent protein-protein interactions of Myb with regulatory proteins.

  11. Transcriptional regulation of muscle fatty acid-binding protein.

    Science.gov (United States)

    Carey, J O; Neufer, P D; Farrar, R P; Veerkamp, J H; Dohm, G L

    1994-03-15

    Heart fatty acid-binding protein (H-FABP) is present in a wide variety of tissues but is found in the highest concentration in cardiac and red skeletal muscle. It has been proposed that the expression of H-FABP correlates directly with the fatty acid-oxidative capacity of the tissue. In the present study, the expression of H-FABP was measured in red and white skeletal muscle under two conditions in which fatty acid utilization is known to be increased: streptozotocin-induced diabetes and fasting. Protein concentration, mRNA concentration and transcription rate were measured under both conditions. The level of both protein and mRNA increased approximately 2-fold under each condition. The transcription rate was higher in red skeletal muscle than in white muscle, was increased 2-fold during fasting, but was unchanged by streptozotocin-induced diabetes. In addition to supporting the hypothesis that H-FABP is induced during conditions of increased fatty acid utilization, these findings demonstrate that the regulation of H-FABP expression may or may not be at the level of transcription depending on the stimulus.

  12. Sequence and structural features of binding site residues in protein-protein complexes: comparison with protein-nucleic acid complexes

    Directory of Open Access Journals (Sweden)

    Selvaraj S

    2011-10-01

    Full Text Available Abstract Background Protein-protein interactions are important for several cellular processes. Understanding the mechanism of protein-protein recognition and predicting the binding sites in protein-protein complexes are long standing goals in molecular and computational biology. Methods We have developed an energy based approach for identifying the binding site residues in protein–protein complexes. The binding site residues have been analyzed with sequence and structure based parameters such as binding propensity, neighboring residues in the vicinity of binding sites, conservation score and conformational switching. Results We observed that the binding propensities of amino acid residues are specific for protein-protein complexes. Further, typical dipeptides and tripeptides showed high preference for binding, which is unique to protein-protein complexes. Most of the binding site residues are highly conserved among homologous sequences. Our analysis showed that 7% of residues changed their conformations upon protein-protein complex formation and it is 9.2% and 6.6% in the binding and non-binding sites, respectively. Specifically, the residues Glu, Lys, Leu and Ser changed their conformation from coil to helix/strand and from helix to coil/strand. Leu, Ser, Thr and Val prefer to change their conformation from strand to coil/helix. Conclusions The results obtained in this study will be helpful for understanding and predicting the binding sites in protein-protein complexes.

  13. In silico classification of proteins from acidic and neutral cytoplasms.

    Directory of Open Access Journals (Sweden)

    Yaping Fang

    Full Text Available Protein acidostability is a common problem in biopharmaceutical and other industries. However, it remains a great challenge to engineer proteins for enhanced acidostability because our knowledge of protein acidostabilization is still very limited. In this paper, we present a comparative study of proteins from bacteria with acidic (AP and neutral cytoplasms (NP using an integrated statistical and machine learning approach. We construct a set of 393 non-redundant AP-NP ortholog pairs and calculate a total of 889 sequence based features for these proteins. The pairwise alignments of these ortholog pairs are used to build a residue substitution propensity matrix between APs and NPs. We use Gini importance provided by the Random Forest algorithm to rank the relative importance of these features. A scoring function using the 10 most significant features is developed and optimized using a hill climbing algorithm. The accuracy of the score function is 86.01% in predicting AP-NP ortholog pairs and is 76.65% in predicting non-ortholog AP-NP pairs, suggesting that there are significant differences between APs and NPs which can be used to predict relative acidostability of proteins. The overall trends uncovered in the study can be used as general guidelines for designing acidostable proteins. To best of our knowledge, this work represents the first systematic comparative study of the acidostable proteins and their non-acidostable orthologs.

  14. Saturated fatty acids modulate autophagy's proteins in the hypothalamus.

    Science.gov (United States)

    Portovedo, Mariana; Ignacio-Souza, Letícia M; Bombassaro, Bruna; Coope, Andressa; Reginato, Andressa; Razolli, Daniela S; Torsoni, Márcio A; Torsoni, Adriana S; Leal, Raquel F; Velloso, Licio A; Milanski, Marciane

    2015-01-01

    Autophagy is an important process that regulates cellular homeostasis by degrading dysfunctional proteins, organelles and lipids. In this study, the hypothesis that obesity could lead to impairment in hypothalamic autophagy in mice was evaluated by examining the hypothalamic distribution and content of autophagic proteins in animal with obesity induced by 8 or 16 weeks high fat diet to induce obesity and in response to intracerebroventricular injections of palmitic acid. The results showed that chronic exposure to a high fat diet leads to an increased expression of inflammatory markers and downregulation of autophagic proteins. In obese mice, autophagic induction leads to the downregulation of proteins, such as JNK and Bax, which are involved in the stress pathways. In neuron cell-line, palmitate has a direct effect on autophagy even without inflammatory activity. Understanding the cellular and molecular bases of overnutrition is essential for identifying new diagnostic and therapeutic targets for obesity.

  15. Developmental changes in content of glial marker proteins in rats exposed to protein malnutrition.

    Science.gov (United States)

    Feoli, Ana Maria; Leite, Marina C; Tramontina, Ana Carolina; Tramontina, Francine; Posser, Thais; Rodrigues, Letícia; Swarowsky, Alessandra; Quincozes-Santos, André; Leal, Rodrigo B; Gottfried, Carmem; Perry, Marcos Luiz; Gonçalves, Carlos-Alberto

    2008-01-02

    Pre- and postnatal protein malnutrition (PMN) adversely affects the developing brain in numerous ways, but only a few studies have investigated specific glial parameters. This study aimed to evaluate specific glial changes of rats exposed to pre and postnatal PMN, based on glial fibrillary acidic protein (GFAP) and S100B immunocontents as well as glutamine synthetase (GS), in cerebral cortex, hippocampus, cerebellum and cerebrospinal fluid, on the 2nd, 15th and 60th postnatal days. We found increases in GFAP, S100B and GS in the cerebral cortex at birth, suggesting an astrogliosis. Hippocampus and cerebellum also exhibited this profile at birth. However, a significant interaction between age and diet in postnatal life was observed only in the S100B of the cerebral cortex. No changes in the content of GFAP and S100B and GS activity were found on the 60th postnatal day in malnourished rats. In contrast, following an increase in the levels of S100B in the cerebrospinal fluid, during the early developmental stages, levels remained elevated on the 60th postnatal day. Our data support the concept of astrogliosis at birth, induced by PMN, and involve extracellular-regulated kinase activation. Specific alterations in cerebral cortex emphasize the regional vulnerability of the brain to malnutrition; some alterations were observed only at birth (e.g. GFAP); others were observed on the 2nd and 15th post-natal days (e.g. ERK phosphorylation). Taken together, transient and persistent alterations (e.g. elevated extracellular levels of S100B) suggest some brain damage or a risk of brain diseases in rats exposed to PMN.

  16. Hypochlorous acid-mediated protein oxidation: how important are chloramine transfer reactions and protein tertiary structure?

    Science.gov (United States)

    Pattison, David I; Hawkins, Clare L; Davies, Michael J

    2007-08-28

    Hypochlorous acid (HOCl) is a powerful oxidant generated from H2O2 and Cl- by the heme enzyme myeloperoxidase, which is released from activated leukocytes. HOCl possesses potent antibacterial properties, but excessive production can lead to host tissue damage that occurs in numerous human pathologies. As proteins and amino acids are highly abundant in vivo and react rapidly with HOCl, they are likely to be major targets for HOCl. In this study, two small globular proteins, lysozyme and insulin, have been oxidized with increasing excesses of HOCl to determine whether the pattern of HOCl-mediated amino acid consumption is consistent with reported kinetic data for isolated amino acids and model compounds. Identical experiments have been carried out with mixtures of N-acetyl amino acids (to prevent reaction at the alpha-amino groups) that mimic the protein composition to examine the role of protein structure on reactivity. The results indicate that tertiary structure facilitates secondary chlorine transfer reactions of chloramines formed on His and Lys side chains. In light of these data, second-order rate constants for reactions of Lys side chain and Gly chloramines with Trp side chains and disulfide bonds have been determined, together with those for further oxidation of Met sulfoxide by HOCl and His side chain chloramines. Computational kinetic models incorporating these additional rate constants closely predict the experimentally observed amino acid consumption. These studies provide insight into the roles of chloramine formation and three-dimensional structure on the reactions of HOCl with isolated proteins and demonstrate that kinetic models can predict the outcome of HOCl-mediated protein oxidation.

  17. An amino acid depleted cell-free protein synthesis system for the incorporation of non-canonical amino acid analogs into proteins.

    Science.gov (United States)

    Singh-Blom, Amrita; Hughes, Randall A; Ellington, Andrew D

    2014-05-20

    Residue-specific incorporation of non-canonical amino acids into proteins is usually performed in vivo using amino acid auxotrophic strains and replacing the natural amino acid with an unnatural amino acid analog. Herein, we present an efficient amino acid depleted cell-free protein synthesis system that can be used to study residue-specific replacement of a natural amino acid by an unnatural amino acid analog. This system combines a simple methodology and high protein expression titers with a high-efficiency analog substitution into a target protein. To demonstrate the productivity and efficacy of a cell-free synthesis system for residue-specific incorporation of unnatural amino acids in vitro, we use this system to show that 5-fluorotryptophan and 6-fluorotryptophan substituted streptavidin retain the ability to bind biotin despite protein-wide replacement of a natural amino acid for the amino acid analog. We envisage this amino acid depleted cell-free synthesis system being an economical and convenient format for the high-throughput screening of a myriad of amino acid analogs with a variety of protein targets for the study and functional characterization of proteins substituted with unnatural amino acids when compared to the currently employed in vivo methodologies.

  18. Quantitation of glial fibrillary acidic protein in human brain tumours

    DEFF Research Database (Denmark)

    Rasmussen, S; Bock, E; Warecka, K

    1980-01-01

    The glial fibrillary acidic protein (GFA) content of 58 human brain tumours was determined by quantitative immunoelectrophoresis, using monospecific antibody against GFA. Astrocytomas, glioblastomas, oligodendrogliomas, spongioblastomas, ependymomas and medulloblastomas contained relatively high...... amounts of GFA, up to 85 times the concentration in parietal grey substance of normal human brain. GFA was not found in neurinomas, meningiomas, adenomas of the hypophysis, or in a single case of metastasis of adenocarcinoma. Non-glial tumours of craniopharyngioma and haemangioblastoma were infiltrated...

  19. The clinical significance of fatty acid binding proteins

    Directory of Open Access Journals (Sweden)

    Barbara Choromańska

    2011-11-01

    Full Text Available Excessive levels of free fatty acids are toxic to cells. The human body has evolved a defense mechanism in the form of small cytoplasmic proteins called fatty acid binding proteins (FABPs that bind long-chain fatty acids (LCFA, and then refer them to appropriate intracellular disposal sites (oxidation in mitochondria and peroxisomes or storage in the endoplasmic reticulum. So far, nine types of these proteins have been described, and their name refers to the place in which they were first identified or where they can be found in the greatest concentration. The most important FABPs were isolated from the liver (L-FABP, heart (H-FABP, intestine (I-FABP, brain (B-FABP, epidermis (E-FABP and adipocytes (A-FABP. Determination of H-FABP is used in the diagnosis of myocardial infarction, and L-FABP in kidney lesions of different etiologies. It is postulated that FABPs play an important role in the pathogenesis of metabolic diseases. Elevated levels of A-FABP have been found in the pericardial fat tissue and were associated with cardiac dysfunction in obese people. A rise in A-FABP has been observed in patients with type II diabetes. I-FABP is known as a marker of cell damage in the small intestine. Increased concentration of B-FABP has been associated with human brain tumors such as glioblastoma and astrocytoma, as well as with neurodegenerative diseases (Alzheimer’s, Parkinson’s and other disorders of cognitive function. The aim of this work was to present current data on the clinical significance of fatty acid binding proteins.

  20. Phytic acid reduction in soy protein improves zinc bioavailability

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, J.R.; Wong, M.S.; Burns, R.A.; Erdman, J.W. Jr. (Univ. of Illinois, Urbana (United States) Mead Johnson Research Center, Evansville, IN (United States))

    1991-03-15

    The objective of this study was to confirm previous studies that have suggested that reduction of phytic acid in soy improved zinc bioavailability (BV). Two commercially-produced soybean isolates containing either a normal phytic acid level or a reduced level were formulated into diets so as to provide 6 or 9 ppm zinc. Control diets were egg white protein-based and contained 3, 6 or 9 ppm zinc from zinc carbonate. Weanling male rats were fed these diets for 21 days and food intake and weight gain monitored. Slope ratio analysis of total tibia zinc content compared to total zinc intake revealed that zinc BV from reduced phytic acid soy isolate-containing diets was indistinguishable from control egg white diets. In contrast, zinc BV from normal soy isolate diets was significantly reduced compared to reduced phytic acid and control diets. These results coupled with other results indicate that phytic acid is the inhibitory factor in soybean products that results in reduced zinc BV.

  1. Longitudinal evolution of true protein, amino acids and bioactive proteins in breast milk: a developmental perspective.

    Science.gov (United States)

    Lönnerdal, Bo; Erdmann, Peter; Thakkar, Sagar K; Sauser, Julien; Destaillats, Frédéric

    2017-03-01

    The protein content of breast milk provides a foundation for estimating protein requirements of infants. Because it serves as a guideline for regulatory agencies issuing regulations for infant formula composition, it is critical that information on the protein content of breast milk is reliable. We have therefore carried out a meta-analysis of the protein and amino acid contents of breast milk and how they evolve during lactation. As several bioactive proteins are not completely digested in the infant and therefore represent "non-utilizable" protein, we evaluated the quantity, mechanism of action and digestive fate of several major breast milk proteins. A better knowledge of the development of the protein contents of breast milk and to what extent protein utilization changes with age of the infant will help improve understanding of protein needs in infancy. It is also essential when designing the composition of infant formulas, particularly when the formula uses a "staging" approach in which the composition of the formula is modified in stages to reflect changes in breast milk and changing requirements as the infant ages.

  2. Novel humic acid-bonded magnetite nanoparticles for protein immobilization

    Energy Technology Data Exchange (ETDEWEB)

    Bayrakci, Mevlut, E-mail: mevlutbayrakci@gmail.com [Ulukisla Vocational School, Nigde University, 51100 Ulukisla, Nigde (Turkey); Gezici, Orhan [Department of Chemistry, Nigde University, 51100 Nigde (Turkey); Bas, Salih Zeki; Ozmen, Mustafa; Maltas, Esra [Department of Chemistry, Selcuk University, 42031 Konya (Turkey)

    2014-09-01

    The present paper is the first report that introduces (i) a useful methodology for chemical immobilization of humic acid (HA) to aminopropyltriethoxysilane-functionalized magnetite iron oxide nanoparticles (APS-MNPs) and (ii) human serum albumin (HSA) binding to the obtained material (HA-APS-MNPs). The newly prepared magnetite nanoparticle was characterized by using Fourier transform infrared spectroscopy (FTIR), transmission electron microscopy (TEM), scanning electron microscopy (SEM), thermogravimetric analysis (TGA), and elemental analysis. Results indicated that surface modification of the bare magnetite nanoparticles (MNPs) with aminopropyltriethoxysilane (APS) and HA was successfully performed. The protein binding studies that were evaluated in batch mode exhibited that HA-APS-MNPs could be efficiently used as a substrate for the binding of HSA from aqueous solutions. Usually, recovery values higher than 90% were found to be feasible by HA-APS-MNPs, while that value was around 2% and 70% in the cases of MNPs and APS-MNPs, respectively. Hence, the capacity of MNPs was found to be significantly improved by immobilization of HA. Furthermore, thermal degradation of HA-APS-MNPs and HSA bonded HA-APS-MNPs was evaluated in terms of the Horowitz–Metzger equation in order to determine kinetic parameters for thermal decomposition. Activation energies calculated for HA-APS-MNPs (20.74 kJ mol{sup −1}) and HSA bonded HA-APS-MNPs (33.42 kJ mol{sup −1}) implied chemical immobilization of HA to APS-MNPs, and tight interactions between HA and HA-APS-MNPs. - Highlights: • A new magnetite nanoparticle based humic acid was prepared for the first time. • Protein binding studies of magnetite nanoparticle based humic acid were performed. • Kinetic parameters of protein and/or humic acid bonded nanoparticles were evaluated.

  3. The adult spinal cord harbors a population of GFAP-positive progenitors with limited self-renewal potential.

    Science.gov (United States)

    Fiorelli, Roberto; Cebrian-Silla, Arantxa; Garcia-Verdugo, Jose-Manuel; Raineteau, Olivier

    2013-12-01

    Adult neural stem cells (aNSCs) of the forebrain are GFAP-expressing cells that are intercalated within ependymal cells of the subventricular zone (SVZ). Cells showing NSCs characteristics in vitro can also be isolated from the periaqueductal region in the adult spinal cord (SC), but contradicting results exist concerning their glial versus ependymal identity. We used an inducible transgenic mouse line (hGFAP-CreERT2) to conditionally label GFAP-expressing cells in the adult SVZ and SC periaqueduct, and directly and systematically compared their self-renewal and multipotential properties in vitro. We demonstrate that a population of GFAP(+) cells that share the morphology and the antigenic properties of SVZ-NSCs mostly reside in the dorsal aspect of the central canal (CC) throughout the spinal cord. These cells are non-proliferative in the intact spinal cord, but incorporate the S-phase marker EdU following spinal cord injury. Multipotent, clonal YFP-expressing neurospheres (i.e., deriving from recombined GFAP-expressing cells) were successfully obtained from both the intact and injured spinal cord. These spheres however showed limited self-renewal properties when compared with SVZ-neurospheres, even after spinal cord injury. Altogether, these results demonstrate that significant differences exist in NSCs lineages between neurogenic and non-neurogenic regions of the adult CNS. Thus, although we confirm that a population of multipotent GFAP(+) cells co-exists alongside with multipotent ependymal cells within the adult SC, we identify these cells as multipotent progenitors showing limited self-renewal properties.

  4. Direct protein-protein interactions and substrate channeling between cellular retinoic acid binding proteins and CYP26B1.

    Science.gov (United States)

    Nelson, Cara H; Peng, Chi-Chi; Lutz, Justin D; Yeung, Catherine K; Zelter, Alex; Isoherranen, Nina

    2016-08-01

    Cellular retinoic acid binding proteins (CRABPs) bind all-trans-retinoic acid (atRA) tightly. This study aimed to determine whether atRA is channeled directly to cytochrome P450 (CYP) CYP26B1 by CRABPs, and whether CRABPs interact directly with CYP26B1. atRA bound to CRABPs (holo-CRABP) was efficiently metabolized by CYP26B1. Isotope dilution experiments showed that delivery of atRA to CYP26B1 in solution was similar with or without CRABP. Holo-CRABPs had higher affinity for CYP26B1 than free atRA, but both apo-CRABPs inhibited the formation of 4-OH-RA by CYP26B1. Similar protein-protein interactions between soluble binding proteins and CYPs may be important for other lipophilic CYP substrates.

  5. Astrocyte behavior and GFAP expression on Spirulina extract-incorporated PCL nanofiber.

    Science.gov (United States)

    Min, Seul Ki; Kim, Cho Rong; Jung, Sang Myung; Shin, Hwa Sung

    2013-12-01

    Nanomaterials are attractive for use in biological systems due to their ability to control the microenvironment of cells. Additionally, nanofibers can mimic fibrous characteristics of natural tissues. This study was conducted to assess astrocyte activity and infiltration behavior on Spirulina extract-embedded polycaprolactone (SP-PCL) nanofiber. Astrocytes moved along with the nanofiber, and developed an elongated and stellate shape, which is similar to those in the natural neural tissue. In addition, the expression of GFAP, a biomarker representing the activation of astrocytes, was gradually up-regulated with the increase of the concentration of Spirulina extract, indicating that Spirulina extract can control astrocyte activation. Overall, the results presented herein indicate that SP-PCL nanofiber could be used in astrocyte tissue engineering for neuronal regeneration. Copyright © 2013 Wiley Periodicals, Inc., a Wiley Company.

  6. The calcium-modulated proteins, S100A1 and S100B, as potential regulators of the dynamics of type III intermediate filaments

    Directory of Open Access Journals (Sweden)

    M. Garbuglia

    1999-10-01

    Full Text Available The Ca2+-modulated, dimeric proteins of the EF-hand (helix-loop-helix type, S100A1 and S100B, that have been shown to inhibit microtubule (MT protein assembly and to promote MT disassembly, interact with the type III intermediate filament (IF subunits, desmin and glial fibrillary acidic protein (GFAP, with a stoichiometry of 2 mol of IF subunit/mol of S100A1 or S100B dimer and an affinity of 0.5-1.0 µM in the presence of a few micromolar concentrations of Ca2+. Binding of S100A1 and S100B results in inhibition of desmin and GFAP assemblies into IFs and stimulation of the disassembly of preformed desmin and GFAP IFs. S100A1 and S100B interact with a stretch of residues in the N-terminal (head domain of desmin and GFAP, thereby blocking the head-to-tail process of IF elongation. The C-terminal extension of S100A1 (and, likely, S100B represents a critical part of the site that recognizes desmin and GFAP. S100B is localized to IFs within cells, suggesting that it might have a role in remodeling IFs upon elevation of cytosolic Ca2+ concentration by avoiding excess IF assembly and/or promoting IF disassembly in vivo. S100A1, that is not localized to IFs, might also play a role in the regulation of IF dynamics by binding to and sequestering unassembled IF subunits. Together, these observations suggest that S100A1 and S100B may be regarded as Ca2+-dependent regulators of the state of assembly of two important elements of the cytoskeleton, IFs and MTs, and, potentially, of MT- and IF-based activities.

  7. Retinoic acid binding protein in normal and neopolastic rat prostate.

    Science.gov (United States)

    Gesell, M S; Brandes, M J; Arnold, E A; Isaacs, J T; Ueda, H; Millan, J C; Brandes, D

    1982-01-01

    Sucrose density gradient analysis of cytosol from normal and neoplastic rat prostatic tissues exhibited a peak of (3H) retinoic acid binding in the 2S region, corresponding to the cytoplasmic retinoic acid binding protein (cRABP). In the Fisher-Copenhagen F1 rat, cRABP was present in the lateral lobe, but could not be detected in the ventral nor in the dorsal prostatic lobes. Four sublines of the R-3327 rat prostatic tumor contained similar levels of this binding protein. The absence of cRABP in the normal tissue of origin of the R-3327 tumor, the rat dorsal prostate, and reappearance in the neoplastic tissues follows a pattern described in other human and animal tumors. The occurrence of cRABP in the well-differentiated as well as in the anaplastic R-3327 tumors in which markers which reflect a state of differentiation and hormonal regulation, such as androgen receptor, 5 alpha reductase, and secretory acid phosphatase are either markedly reduced or absent, points to cRABP as a marker of malignant transformation.

  8. Variability of DNA structure and protein-nucleic acid reconginition

    Directory of Open Access Journals (Sweden)

    Shestopalova A. V.

    2010-09-01

    Full Text Available Revealing molecular mechanisms of sequence-specific recognition of DNA by proteins is one of the key tasks of biology. The current review presents the results of statistical analysis of the structural databases obtained by different scientific groups studying the conformational features of free and protein-bound DNA fragments that could be used for clarifying the mechanisms of protein-nucleic acid recognition. The analysis of the published data allowed us to make the following generalizations. The ability of DNA double helix to adopt alternative conformations, including the ones of sugarphosphate backbone, is an intrinsic characteristic of certain DNA sequences. Such conformational transitions are the potential sources of formation of unique geometry of the dinucleotide steps and/or individual nucleotides and lead to alteration of base stacking and/or changes of the assessable surface area of atoms, and can be the criteria of recognition of DNA by protein as well. Changes in the physical properties that depend on the DNA structure, i. e. the polar/unpolar profile and electrostatic potential of the grooves, can also be used by protein for DNA readout.

  9. d-Amino Acid Scan of Two Small Proteins.

    Science.gov (United States)

    Simon, Mark D; Maki, Yuta; Vinogradov, Alexander A; Zhang, Chi; Yu, Hongtao; Lin, Yu-Shan; Kajihara, Yasuhiro; Pentelute, Bradley L

    2016-09-21

    A "D-scan" of two small proteins, the disulfide-rich Ecballium elaterium trypsin inhibitor II (EETI-II) and a minimized Z domain of protein A (Z33), is reported. For each protein, the stereochemistry of one amino acid at a time was inverted to generate a series of diastereomers. In much the same way an alanine scan determines necessary residues for protein function, the D-scan elucidated the critical stereocenters of the 30-residue EETI-II and the 33-residue Z33. The folding properties and activity of each variant were investigated. A total of 24 out of 30 EETI-II D-scan analogues folded to give a three-disulfide product. Of the 24 variants that folded, half were high-affinity trypsin inhibitors, and three were as active as the wild type (WT). Of these 12 active variants, most were substantially less stable to reduction than WT EETI-II (WT first reduction potential -270.0 ± 1.5 mV, WT second reduction potential -307.2 ± 1.1 mV). Similarly, ten Z33 analogues retained high binding affinity to IgG (KD Z33 analogues were substantially less stable than the WT (ΔG(H2O, 263 K) = 2.4 ± 1.2 kcal/mol). Collectively, our findings show that the D-scan is powerful new strategy for studying how the stereochemistry of amino acids affects the structure and function of proteins.

  10. Analysis of Salicylic Acid Induced Proteins in Rice

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    An analysis using SDS-PAGE of acidic and basic protein fractions extracted from rice seedling treated with salicylic acid (SA) yielded several new proteins, some of which are similar in relative molecular mass to PR-1a,c, PR-2, 2e and PR-3d, 3e of tobacco.Direct assays for peroxidases and β-1,3-glucanases demonstrated that the activities of the two enzymes in the rice seedlings increased rapidly with time after SA treatment, reaching a maximum 6 days after treatment.Disease resistance tests showed that SA treated rice seedlings stunted the development of blight lesions and displayed higher resistance to rice blight pathogen (Xanthomonas oryzea pv.oryzea).The data suggest that the treatment with SA, even for plants with high endogenous SA levels such as rice, may induce the appearance of new proteins and the formation of disease resistance.The results contribute to the analysis of the SA role in rice systemic acquired resistance.

  11. Amino acid profiles and digestible indispensable amino acid scores of proteins from the prioritized key foods in Bangladesh.

    Science.gov (United States)

    Shaheen, Nazma; Islam, Saiful; Munmun, Sarah; Mohiduzzaman, Md; Longvah, Thingnganing

    2016-12-15

    Concentrations of standard amino acids were determined in the composite samples (representing 30 agro-ecological zones of Bangladesh) of six prioritized key dietary protein sources: Oryza sativa (rice), Triticum aestivum (wheat flour), Lens culinaris (lentils), Pangusius pangusius (pangas), Labeo rohita (rohu) and Oreochromis mossambicus (tilapia). Digestible indispensable amino acid scores (DIAAS) was calculated using published data on amino acids' digestibility to evaluate the protein quality of these foods. Indispensable amino acid (IAA) contents (mg IAA/g protein), found to be highest in pangas (430) and lowest in wheat (336), of all these analyzed foods exceeded the FAO recommended daily allowance (277mg IAA/g protein) and contributed on average 40% to total amino acid contents. Untruncated DIAAS values ranged from 51% (lysine) in wheat to 106% (histidine) in pangas and distinguished pangas, rohu, and tilapia containing 'excellent quality' protein (DIAAS>100%) with potential to complement lower quality protein of cereals, fruits, and vegetables.

  12. An Amino Acid Code for Irregular and Mixed Protein Packing

    Science.gov (United States)

    Joo, Hyun; Chavan, Archana; Fraga, Keith; Tsai, Jerry

    2015-01-01

    To advance our understanding of protein tertiary structure, the development of the knob-socket model is completed in an analysis of the packing in irregular coil and turn secondary structure packing as well as between mixed secondary structure. The knob-socket model simplifies packing based on repeated patterns of 2 motifs: a 3 residue socket for packing within 2° structure and a 4 residue knob-socket for 3° packing. For coil and turn secondary structure, knob-sockets allow identification of a correlation between amino acid composition and tertiary arrangements in space. Coil contributes almost as much as α-helices to tertiary packing. Irregular secondary structure involves 3 residue cliques of consecutive contacting residues or XYZ sockets. In irregular sockets, Gly, Pro, Asp and Ser are favored, while Cys, His, Met and Trp are not. For irregular knobs, the preference order is Arg, Asp, Pro, Asn, Thr, Leu, and Gly, while Cys, His, Met and Trp are not. In mixed packing, the knob amino acid preferences are a function of the socket that they are packing into, whereas the amino acid composition of the sockets does not depend on the secondary structure of the knob. A unique motif of a coil knob with an XYZ β-sheet socket may potentially function to inhibit β-sheet extension. In addition, analysis of the preferred crossing angles for strands within a β-sheet and mixed α-helices/β-sheets identifies canonical packing patterns useful in protein design. Lastly, the knob-socket model abstracts the complexity of protein tertiary structure into an intuitive packing surface topology map. PMID:26370334

  13. Acid-base chemistry of frustrated water at protein interfaces.

    Science.gov (United States)

    Fernández, Ariel

    2016-01-01

    Water molecules at a protein interface are often frustrated in hydrogen-bonding opportunities due to subnanoscale confinement. As shown, this condition makes them behave as a general base that may titrate side-chain ammonium and guanidinium cations. Frustration-based chemistry is captured by a quantum mechanical treatment of proton transference and shown to remove same-charge uncompensated anticontacts at the interface found in the crystallographic record and in other spectroscopic information on the aqueous interface. Such observations are untenable within classical arguments, as hydronium is a stronger acid than ammonium or guanidinium. Frustration enables a directed Grotthuss mechanism for proton transference stabilizing same-charge anticontacts.

  14. Reversible lysine modification on proteins by using functionalized boronic acids.

    Science.gov (United States)

    Cal, Pedro M S D; Frade, Raquel F M; Cordeiro, Carlos; Gois, Pedro M P

    2015-05-26

    Iminoboronates have been utilized to successfully install azide and alkyne bioorthogonal functions on proteins, which may then be further reacted with their bioorthogonal counterparts. These constructs were also used to add polyethylene glycol (PEG) to insulin, a modification which has been shown to be reversible in the presence of fructose. Finally, iminoboronates were used to assemble a folic acid/paclitaxel small-molecule/drug conjugate in situ with an IC50  value of 20.7 nM against NCI-H460 cancer cells and negligible cytotoxicity against the CRL-1502 noncancer cells. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Non-standard amino acid incorporation into proteins using Escherichia coli cell-free protein synthesis

    Directory of Open Access Journals (Sweden)

    Seok Hoon eHong

    2014-06-01

    Full Text Available Incorporating non-standard amino acids (NSAAs into proteins enables new chemical properties, new structures, and new functions. In recent years, improvements in cell-free protein synthesis (CFPS systems have opened the way to accurate and efficient incorporation of NSAAs into proteins. The driving force behind this development has been three-fold. First, a technical renaissance has enabled high-yielding (>1 g/L and long-lasting (>10 h in batch operation CFPS in systems derived from Escherichia coli. Second, the efficiency of orthogonal translation systems has improved. Third, the open nature of the CFPS platform has brought about an unprecedented level of control and freedom of design. Here, we review recent developments in CFPS platforms designed to precisely incorporate NSAAs. In the coming years, we anticipate that CFPS systems will impact efforts to elucidate structure/function relationships of proteins and to make biomaterials and sequence-defined biopolymers for medical and industrial applications.

  16. Phosphorylation of acidic ribosomal proteins from rabbit reticulocytes by a ribosome-associated casein kinase

    DEFF Research Database (Denmark)

    Issinger, O G

    1977-01-01

    Two acidic proteins from 80-S ribosomes were isolated and purified to homogeneity. The purified acidic proteins could be phosphorylated by casein kinase using [gamma-32P]ATP and [gamma-32P]GTP as a phosphoryl donor. The proteins became phosphorylated in situ, too. Sodium dodecyl sulfate polyacryl......Two acidic proteins from 80-S ribosomes were isolated and purified to homogeneity. The purified acidic proteins could be phosphorylated by casein kinase using [gamma-32P]ATP and [gamma-32P]GTP as a phosphoryl donor. The proteins became phosphorylated in situ, too. Sodium dodecyl sulfate...

  17. Utilization of Glial Fibrillary Acidic Protein and Galectin-3 in the ...

    African Journals Online (AJOL)

    2017-05-16

    May 16, 2017 ... in clinically suspected ischemic stroke patients with normal computerized cranial tomography ... of Galectin-3 and GFAP were increased in acute ischemic stroke patients. Utilization of ..... Bench-to-bedside review: Outcome.

  18. pH shift protein recovery with organic acids on texture and color of cooked gels.

    Science.gov (United States)

    Paker, Ilgin; Beamer, Sarah; Jaczynski, Jacek; Matak, Kristen E

    2015-01-01

    Isoelectric solubilization and precipitation (ISP) processing uses pH shifts to separate protein from fish frames, which may increase commercial interest for silver carp. Texture and color properties of gels made from silver carp protein recovered at different pH strategies and organic acid types were compared. ISP was applied to headed gutted silver carp using 10 mol L(-1) sodium hydroxide (NaOH) and either glacial acetic acid (AA) or a (1:1) formic and lactic acid combination (F&L). Protein gels were made with recovered protein and standard functional additives. Texture profile analysis and the Kramer shear test showed that protein gels made from protein solubilized at basic pH values were firmer, harder, more cohesive, gummier and chewier (P proteins solubilized under acidic conditions. Acidic solubilization led to whiter (P protein using organic acids show potential for use as a functional ingredient in restructured foods. © 2014 Society of Chemical Industry.

  19. Shedding light on proteins, nucleic acids, cells, humans and fish

    Science.gov (United States)

    Setlow, Richard B.

    2002-01-01

    I was trained as a physicist in graduate school. Hence, when I decided to go into the field of biophysics, it was natural that I concentrated on the effects of light on relatively simple biological systems, such as proteins. The wavelengths absorbed by the amino acid subunits of proteins are in the ultraviolet (UV). The wavelengths that affect the biological activities, the action spectra, also are in the UV, but are not necessarily parallel to the absorption spectra. Understanding these differences led me to investigate the action spectra for affecting nucleic acids, and the effects of UV on viruses and cells. The latter studies led me to the discovery of the important molecular nature of the damages affecting DNA (cyclobutane pyrimidine dimers) and to the discovery of nucleotide excision repair. Individuals with the genetic disease xeroderma pigmentosum (XP) are extraordinarily sensitive to sunlight-induced skin cancer. The finding, by James Cleaver, that their skin cells were defective in DNA repair strongly suggested that DNA damage was a key step in carcinogenesis. Such information was important for estimating the wavelengths in sunlight responsible for human skin cancer and for predicting the effects of ozone depletion on the incidence of non-melanoma skin cancer. It took experiments with backcross hybrid fish to call attention to the probable role of the longer UV wavelengths not absorbed by DNA in the induction of melanoma. These reflections trace the biophysicist's path from molecules to melanoma.

  20. Investigation of protein-fatty acid interactions in zein films

    Science.gov (United States)

    Wang, Qin

    Zein, the prolamin of corn, has shown potential as industrial biopolymer for packaging and agricultural uses. Previous researchers plasticized zein with oleic acid and extruded it into sheets and films. Such products showed reasonable tensile and water barrier properties. However, those properties were affected by film structure, which is believed controlled by the interaction between zein and plasticizer. The nature of those interactions is still not well understood. Protein-fatty acid interactions in zein resin films were investigated by surface plasmon resonance and atomic force microscopy in this work. Preliminary research was conducted to establish the effect of solvent systems on the plasticization of zein by oleic acid. Although slight differences were observed, it was concluded that the use of 75% ethanol or 75% 2-propanol produced similar results. Surface plasmon resonance was employed to investigate zein static and dynamic adsorption on surfaces of hydrophilic and hydrophobic self-assembled monolayers (SAMs) generated by 11-mercaptoundecanoic acid or 1-octanethiol, respectively, and representing the two end groups of oleic acid. Results indicated that zein was adsorbed to both surfaces but showed higher affinity for hydrophilic groups. The corresponding thickness of zein specific binding layer on hydrophilic and hydrophobic SAMs was around 4.7 nm and 4.6 nm. Zein exhibited higher affinity for hydrophilic than for hydrophobic SAMs evidenced from the higher initial adsorption rate and ultimate surface coverage at all zein concentrations. Flushing surface with buffer would leave an apparent monolayer of zein, which is 5 times higher for hydrophilic than hydrophobic SAMs. This observation suggested that zein may use different sides of its molecule to interact with hydrophobic or hydrophilic groups. The surface topography of zein deposits on both SAMs was examined by atomic force microscopy. It was found that zein formed distinct ring-shaped structures with

  1. The role of whey acidic protein four-disulfide-core proteins in respiratory health and disease.

    Science.gov (United States)

    Small, Donna M; Doherty, Declan F; Dougan, Caoifa M; Weldon, Sinéad; Taggart, Clifford C

    2017-04-01

    Members of the whey acidic protein (WAP) or WAP four-disulfide-core (WFDC) family of proteins are a relatively under-explored family of low molecular weight proteins. The two most prominent WFDC proteins, secretory leukocyte protease inhibitor (SLPI) and elafin (or the precursor, trappin-2), have been shown to possess multiple functions including anti-protease, anti-bacterial, anti-viral and anti-inflammatory properties. It is therefore of no surprise that both SLPI and elafin/trappin-2 have been developed as potential therapeutics. Given the abundance of SLPI and elafin/trappin-2 in the human lung, most work in the area of WFDC research has focused on the role of WFDC proteins in protecting the lung from proteolytic attack. In this review, we will outline the current evidence regarding the expanding role of WFDC protein function with a focus on WFDC activity in lung disease as well as emerging data regarding the function of some of the more recently described WFDC proteins.

  2. The folding type of a protein is relevant to the amino acid composition

    OpenAIRE

    Nakashima, Hiroshi; Nishikawa, Ken; Ooi, Tatsuo

    1986-01-01

    The folding types of 135 proteins, the three-dimensional structures of which are known, were analyzed in terms of the amino acid composition. The amino acid composition of a protein was expressed as a point in a multidimensional space spanned with 20 axes, on which the corresponding contents of 20 amino acids in the protein were represented. The distribution pattern of proteins in this composition space was examined in relation to five folding types, , ß, /ß, +ß, and irregular type. The resul...

  3. 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.

  4. Bile salt recognition by human liver fatty acid binding protein.

    Science.gov (United States)

    Favretto, Filippo; Santambrogio, Carlo; D'Onofrio, Mariapina; Molinari, Henriette; Grandori, Rita; Assfalg, Michael

    2015-04-01

    Fatty acid binding proteins (FABPs) act as intracellular carriers of lipid molecules, and play a role in global metabolism regulation. Liver FABP (L-FABP) is prominent among FABPs for its wide ligand repertoire, which includes long-chain fatty acids as well as bile acids (BAs). In this work, we performed a detailed molecular- and atomic-level analysis of the interactions established by human L-FABP with nine BAs to understand the binding specificity for this important class of cholesterol-derived metabolites. Protein-ligand complex formation was monitored using heteronuclear NMR, steady-state fluorescence spectroscopy, and mass spectrometry. BAs were found to interact with L-FABP with dissociation constants in the narrow range of 0.6-7 μm; however, the diverse substitution patterns of the sterol nucleus and the presence of side-chain conjugation resulted in complexes endowed with various degrees of conformational heterogeneity. Trihydroxylated BAs formed monomeric complexes in which single ligand molecules occupied similar internal binding sites, based on chemical-shift perturbation data. Analysis of NMR line shapes upon progressive addition of taurocholate indicated that the binding mechanism departed from a simple binary association equilibrium, and instead involved intermediates along the binding path. The co-linear chemical shift behavior observed for L-FABP complexes with cholate derivatives added insight into conformational dynamics in the presence of ligands. The observed spectroscopic features of L-FABP/BA complexes, discussed in relation to ligand chemistry, suggest possible molecular determinants of recognition, with implications regarding intracellular BA transport. Our findings suggest that human L-FABP is a poorly selective, universal BA binder.

  5. Proximate composition, fatty acid analysis and protein digestibility-corrected amino acid score of three Mediterranean cephalopods.

    Science.gov (United States)

    Zlatanos, Spiros; Laskaridis, Kostas; Feist, Christian; Sagredos, Angelos

    2006-10-01

    Proximate composition, fatty acid analysis and protein digestibility-corrected amino acid score (PDCAAS) in three commercially important cephalopods of the Mediterranean sea (cuttlefish, octopus and squid) were determined. The results of the proximate analysis showed that these species had very high protein:fat ratios similar to lean beef. Docosahexaenoic, palmitic and eicosipentaenoic acid were the most abundant fatty acids among analyzed species. The amount of n-3 fatty acids was higher than that of saturated, monounsaturated and n-6 fatty acids. Despite the fact that cephalopods contain small amounts of fat they were found quite rich in n-3 fatty acids. Finally, PDCAAS indicated that these organisms had a very good protein quality.

  6. Histogenesis and possible mechanism of chondroid changes in mixed tumour of the skin: immunohistochemical evaluation of bone morphogenetic protein, glycosaminoglycans, keratin, vimentin and neuronal markers.

    Science.gov (United States)

    Mori, M; Shrestha, P; Sakamoto, F; Yang, L J; Qin, C; Tsujimura, T

    1994-01-01

    The distribution of immunoreactivity of bone morphogenetic protein (BMP), the glycosaminoglycans chondroitin 4-sulphate (C4SPG), chondroitin 6-sulphate (C6SPG), dermatan sulphate (DSPG) and keratan sulphate proteoglycans (KSPG), cytokeratin (K8.12), vimentin, glial fibrillary acidic protein (GFAP), actin, desmin, S-100 protein and neuron-specific enolase (NSE) in mixed tumour of the skin was investigated using immunohistochemical methods using monoclonal (MoAb) and polyclonal antibodies (PoAb). A strong BMP immunoreactivity was found characteristically in outer tumour cells of tubuloductal structures and modified myoepithelial cells. Modified myoepithelial cells and chondroidally changed cells showed positive immunoreactivity for C4SPG, C6SPG and DSPG; and KSPG was more pronounced in the modified myoepithelial cells. Vimentin, S-100 protein, GFAP and NSE, but not actin and desmin, were distribute in the outer tumour cells and modified myoepithelial cells in chondroidally changed tissue. Two factors show that chondrogenesis in mixed tumour of the skin is associated with the modified myoepithelial cells through the activity of BMP and biosynthesis of glycosaminoglycans as matrix substance. First, outer or basal tumour cells in mixed tumour of the skin is characterized by the presence of positive immunoreactivity for BMP, KSPG, vimentin, cytokeratin K8.12, S-100 protein, GFAP and NSE, and second, there is a matrix of chondroidally changed tissue containing the reaction products of C4SPG, C6SPG, DSPF and KSPG.

  7. Thermophysical properties of starch and whey protein composite prepared in presence of organic acid and esters

    Science.gov (United States)

    Previously, we prepared starch and protein composite by reactive mixing in presence of various organic acids and found that use of these acid esters resulted in composites with good mechanical properties. In this study, concentration (% w/w) of acid citrates in the starch-protein composites were var...

  8. Defining meal requirements for protein to optimize metabolic roles of amino acids

    Science.gov (United States)

    Dietary protein provides essential amino acids (EAAs) for the synthesis of new proteins plus an array of other metabolic functions; many of these functions are sensitive to postprandial plasma and intracellular amino acid concentrations. Recent research has focused on amino acids as metabolic signal...

  9. Snake venom. The amino acid sequence of protein A from Dendroaspis polylepis polylepis (black mamba) venom.

    Science.gov (United States)

    Joubert, F J; Strydom, D J

    1980-12-01

    Protein A from Dendroaspis polylepis polylepis venom comprises 81 amino acids, including ten half-cystine residues. The complete primary structures of protein A and its variant A' were elucidated. The sequences of proteins A and A', which differ in a single position, show no homology with various neurotoxins and non-neurotoxic proteins and represent a new type of elapid venom protein.

  10. Site-specific incorporation of redox active amino acids into proteins

    Science.gov (United States)

    Alfonta; Lital , Schultz; Peter G. , Zhang; Zhiwen

    2010-10-12

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate redox active amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with redox active amino acids using these orthogonal pairs.

  11. Site-specific incorporation of redox active amino acids into proteins

    Science.gov (United States)

    Alfonta, Lital; Schultz, Peter G.; Zhang, Zhiwen

    2009-02-24

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate redox active amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with redox active amino acids using these orthogonal pairs.

  12. Site-specific incorporation of redox active amino acids into proteins

    Energy Technology Data Exchange (ETDEWEB)

    Alfonta, Lital [San Diego, CA; Schultz, Peter G [La Jolla, CA; Zhang, Zhiwen [San Diego, CA

    2012-02-14

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate redox active amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with redox active amino acids using these orthogonal pairs.

  13. Site-specific incorporation of redox active amino acids into proteins

    Energy Technology Data Exchange (ETDEWEB)

    Alfonta; Lital (San Diego, CA), Schultz; Peter G. (La Jolla, CA), Zhang; Zhiwen (San Diego, CA)

    2010-10-12

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate redox active amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with redox active amino acids using these orthogonal pairs.

  14. Site-specific incorporation of redox active amino acids into proteins

    Energy Technology Data Exchange (ETDEWEB)

    Alfonta, Lital (San Diego, CA); Schultz, Peter G. (La Jolla, CA); Zhang, Zhiwen (Austin, TX)

    2011-08-30

    Compositions and methods of producing components of protein biosynthetic machinery that include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, and orthogonal pairs of tRNAs/synthetases, which incorporate redox active amino acids into proteins are provided. Methods for identifying these orthogonal pairs are also provided along with methods of producing proteins with redox active amino acids using these orthogonal pairs.

  15. Membrane fractionation of herring marinade for separation and recovery of fats, proteins, amino acids, salt, acetic acid and water

    DEFF Research Database (Denmark)

    Fjerbæk Søtoft, Lene; Lizarazu, Juncal Martin; Razi Parjikolaei, Behnaz;

    2015-01-01

    containing sugars, amino acids and smaller peptides and a NF permeate containing salt and acetic acid ready for reuse. 42% of the spent marinade is recovered to substitute fresh water and chemicals. The Waste water amount is reduced 62.5%. Proteins are concentrated 30 times, while amino acids and smaller......In the production of marinated herring, nearly one ton of acidic saline marinade is produced per 1.5 tons herring fillet. This spent marinade contains highly valuable compounds such as proteins and amino acids. Membranes are suited to recover these substances. In this work, six membrane stages...... are employed: microfiltration (MF) (0.2 lm), ultrafiltration (UF) (50, 20, 10 and 1 kDa) and nanofiltration (NF). The most promising stages are 50 kDa UF and NF based on SDS–PAGE analyses and total amino acid concentration. The 50 kDa stage produces a protein concentrate (>17 kDa). NF produces a retentate...

  16. Interconnection between the protein solubility and amino acid and dipeptide compositions.

    Science.gov (United States)

    Niu, Xiaohui; Li, Nana; Chen, Dinyan; Wang, Zengzhen

    2013-01-01

    Obtaining soluble proteins in sufficient concentrations helps increase the overall success rate in various experimental studies. Protein solubility is an individual trait ultimately determined by its primary protein sequence. Exploring the interconnection between the protein solubility and the compositions of protein sequence is instrumental for setting priorities on targets in large scale proteomics projects. In this paper, amino acid composition (20 dimensions) and the dipeptide composition (400 dimensions) were extracted to form the total candidate feature pool (420 dimensions), and each feature was selected into the feature vectors one by one, which were sorted by the absolute value of the correlation coefficient. Finally, we evaluated and recorded the 420 results of Support Vector Machine (SVM) as the prediction engine. According to the results of SVM, the first 208 features were chosen from the 420 dimensions, which were considered as the efficient ones. By analyzing the composition of the former 208 features, we found that the protein solubility was significantly influenced by the occurrence frequencies of the acidic amino acids, basic amino acids, non-polar hydrophobic amino acids and the two polar neutral amino acids(C, Q) in the protein sequences. Additionally, we detected that the dipeptides composed by the acidic amino acids (D, E) and basic amino acids (K, R and H), especially the dipeptide composed by the acidic amino acids (D, E), had strong interconnection with the protein solubility.

  17. Phytic acid and phytase: implications for protein utilization by poultry.

    Science.gov (United States)

    Cowieson, A J; Acamovic, T; Bedford, M R

    2006-05-01

    The effect of the ingestion of myo-inositol hexaphosphate (IP6) and phytase (EC 3.1.3.26) on the digestibility of casein was investigated using growing broiler chickens. A total of 64 female Ross broilers were used in a precision feeding study. One group of 8 birds was fed a solution of glucose to estimate endogenous losses. Seven groups, each of 8 birds, were fed either casein, casein + 1,000 units of phytase activity (FTU), casein + 2,000 FTU, casein + 0.5 g of IP6, casein + 0.5 g of IP6 + 1,000 FTU, casein + 1 g of IP6, or casein + 1 g of IP6 + 1,000 FTU. The excretion of DM, amino acids, nitrogen, minerals, and phytate-phosphorus was determined over a 48-h period and nutrient digestibility coefficients were calculated. Casein was found to be highly digestible, with true coefficients of DM, N, and amino acid digestibility of between 0.85 and 1.0. However, the ingestion of IP6 reduced (P IP6 with phytase improved (P IP6 with no supplemental phytase. The excretion of endogenous minerals was increased (P IP6 and reduced (P IP6 with phytase. In the absence of exogenous phytase, the recovery of phytate-P in excreta was approximately 80%. However, the recovery of phytate-P was significantly reduced by the addition of exogenous phytase to the IP6/casein mixture. It can be concluded that the ingestion of IP6 reduces the digestibility coefficients of amino acids and the metabolizability of nitrogen of casein. This is likely to be mediated partially through increased endogenous losses. However, the addition of phytase can partially ameliorate the detrimental effects of IP6 on protein utilization.

  18. Nucleic acid induced unfolding of recombinant prion protein globular fragment is pH dependent.

    Science.gov (United States)

    Bera, Alakesh; Nandi, Pradip K

    2014-12-01

    Nucleic acid can catalyze the conversion of α-helical cellular prion protein to β-sheet rich Proteinase K resistant prion protein oligomers and amyloid polymers in vitro and in solution. Because unfolding of a protein molecule from its ordered α-helical structure is considered to be a necessary step for the structural conversion to its β-sheet rich isoform, we have studied the unfolding of the α-helical globular 121-231 fragment of mouse recombinant prion protein in the presence of different nucleic acids at neutral and acid pH. Nucleic acids, either single or double stranded, do not have any significant effect on the secondary structure of the protein fragment at neutral pH; however the protein secondary structure is modified by the nucleic acids at pH 5. Nucleic acids do not show any significant effect on the temperature induced unfolding of the globular prion protein domain at neutral pH which, however, undergoes a gross conformational change at pH 5 as evidenced from the lowering of the midpoint of thermal denaturation temperatures, Tm, of the protein. The extent of Tm decrease shows a dependence on the nature of nucleic acid. The interaction of nucleic acid with the nonpolar groups exposed from the protein interior at pH 5 probably contributes substantially to the unfolding process of the protein. © 2014 The Protein Society.

  19. Value of heart-type fatty acid-binding protein (H-FABP) for ...

    African Journals Online (AJOL)

    Key Words: heart-type fatty acid-binding protein, acute coronary syndrome, biomarker. ... facilitating the intracellular cytoplasmic transport of the fatty acids, and are highly expressed ..... nary syndrome: systematic review and critical appraisal.

  20. Salicylic acid enhances Staphylococcus aureus extracellular adhesin protein expression.

    Science.gov (United States)

    Alvarez, Lucía P; Barbagelata, María S; Cheung, Ambrose L; Sordelli, Daniel O; Buzzola, Fernanda R

    2011-11-01

    One of the virulence factors required by Staphylococcus aureus at the early stages of infection is Eap, a secreted adhesin that binds many host proteins and is upregulated by the two-component regulatory system saeRS. The S. aureus Newman strain harbors a mutation in saeS that is thought to be responsible for the high level of Eap expression in this strain. This study was designed to ascertain whether salicylic acid (SAL) affects the expression of Eap and the internalization of S. aureus into epithelial cells. The strain Newman treated with SAL exhibited increased levels of eap transcription and protein expression. Furthermore, SAL treatment increased the eap promoter activity. SAL treatment enhanced Eap expression in the Newman and in other S. aureus strains that do not carry the mutation in saeS. Internalization of S. aureus eap and sae mutants into the MAC-T epithelial cells was significantly decreased compared with the wild-type counterparts. In conclusion, we demonstrated that a low concentration of SAL increased S. aureus Eap expression possibly due to enhancement of sae. SAL may create the conditions for S. aureus persistence in the host, not only by decreasing the capsular polysaccharide expression as shown before, but also by enhancing Eap expression.

  1. Crystal growth of proteins, nucleic acids, and viruses in gels.

    Science.gov (United States)

    Lorber, Bernard; Sauter, Claude; Théobald-Dietrich, Anne; Moreno, Abel; Schellenberger, Pascale; Robert, Marie-Claire; Capelle, Bernard; Sanglier, Sarah; Potier, Noëlle; Giegé, Richard

    2009-11-01

    Medium-sized single crystals with perfect habits and no defect producing intense and well-resolved diffraction patterns are the dream of every protein crystallographer. Crystals of biological macromolecules possessing these characteristics can be prepared within a medium in which mass transport is restricted to diffusion. Chemical gels (like polysiloxane) and physical gels (such as agarose) provide such an environment and are therefore suitable for the crystallisation of biological macromolecules. Instructions for the preparation of each type of gel are given to urge crystal growers to apply diffusive media for enhancing crystallographic quality of their crystals. Examples of quality enhancement achieved with silica and agarose gels are given. Results obtained with other substances forming gel-like media (such as lipidic phases and cellulose derivatives) are presented. Finally, the use of gels in combination with capillary tubes for counter-diffusion experiments is discussed. Methods and techniques implemented with proteins can also be applied to nucleic acids and nucleoprotein assemblies such as viruses.

  2. Small acid soluble proteins for rapid spore identification.

    Energy Technology Data Exchange (ETDEWEB)

    Branda, Steven S.; Lane, Todd W.; VanderNoot, Victoria A.; Jokerst, Amanda S.

    2006-12-01

    This one year LDRD addressed the problem of rapid characterization of bacterial spores such as those from the genus Bacillus, the group that contains pathogenic spores such as B. anthracis. In this effort we addressed the feasibility of using a proteomics based approach to spore characterization using a subset of conserved spore proteins known as the small acid soluble proteins or SASPs. We proposed developing techniques that built on our previous expertise in microseparations to rapidly characterize or identify spores. An alternative SASP extraction method was developed that was amenable to both the subsequent fluorescent labeling required for laser-induced fluorescence detection and the low ionic strength requirements for isoelectric focusing. For the microseparations, both capillary isoelectric focusing and chip gel electrophoresis were employed. A variety of methods were evaluated to improve the molecular weight resolution for the SASPs, which are in a molecular weight range that is not well resolved by the current methods. Isoelectric focusing was optimized and employed to resolve the SASPs using UV absorbance detection. Proteomic signatures of native wild type Bacillus spores and clones genetically engineered to produce altered SASP patterns were assessed by slab gel electrophoresis, capillary isoelectric focusing with absorbance detection as well as microchip based gel electrophoresis employing sensitive laser-induced fluorescence detection.

  3. Determination of the amount of protein and amino acids extracted from the microbial protein (SCP) of lignocellulosic wastes.

    Science.gov (United States)

    Ahmadi, A R; Ghoorchian, H; Hajihosaini, R; Khanifar, J

    2010-04-15

    With the increasing world population, the use of lignocellulosic wastes for production of microbial protein as animal feed becomes a necessity of our time. In order to verify the most productive protein, the amount of protein and amino acid extracted from Single Cell Protein (SCP) needs to be determined by an effective method. In this study Microbial protein was produced by treatment of wheat straw with Pleurotus florida; with heat at 100 degrees C and NaOH 2% as substrate by solid state fermentation. Concentration of protein was 62.8% per 100 g of dried microbial protein. Then the extracted protein hydrolyzed with HCl 6 Normal for 48 h under 110 degrees C temperature condition. Then the amino acids analyzed by using A-200 Amino Nova analyzer. The results of this study indicated that the ratio of essential amino acids to total amino acids was 65.6%. The concentration of essnyial amino acids were: Lysine = 9.5, histidine = 19.8, threonine = 0.6, valine = 6.6, methionine = 2.1, isoleucine = 7.3, leucine = 6.8, phenylalanine = 4.3 and arginine = 8.3 g/100 g of extracted protein that indicated the obtained microbial protein can be a good or suitable substitute in the food program of animal feed.

  4. Analysis of protein-nucleic acid interactions by photochemical cross-linking and mass spectrometry

    DEFF Research Database (Denmark)

    Steen, Hanno; Jensen, Ole Nørregaard

    2002-01-01

    Photochemical cross-linking is a commonly used method for studying the molecular details of protein-nucleic acid interactions. Photochemical cross-linking aids in defining nucleic acid binding sites of proteins via subsequent identification of cross-linked protein domains and amino acid residues....... Mass spectrometry (MS) has emerged as a sensitive and efficient analytical technique for determination of such cross-linking sites in proteins. The present review of the field describes a number of MS-based approaches for the characterization of cross-linked protein-nucleic acid complexes...... and for sequencing of peptide-nucleic acid heteroconjugates. The combination of photochemical cross-linking and MS provides a fast screening method to gain insights into the overall structure and formation of protein-oligonucleotide complexes. Because the analytical methods are continuously refined and protein...

  5. Dietary Fat Content Effects on Concentrations of Liver and Intestinal Fatty Acid Binding Proteins

    OpenAIRE

    Murakami, Hiroshi; Sakai, Yasuo; Ohta, Kazutoshi; Hatakeyama, Katsuyoshi

    1998-01-01

    Two fatty acid binding proteins, liver and intestinal, have been identified in the rat intestine. Both are thought to be closely related to the absorption and metabolism of fatty acids in the intestinal epithelium. However, the underlying mechanism is not clearly understood. The purpose of this study was therefore to investigate the roles of these two fatty acid binding proteins in the intestinal absorption of fatty acids. Rats were fed diets varying in fat content for two or four weeks. Live...

  6. PROTEIN METABOLISM IN REGENERATING WOUND TISSUE: FUNCTION OF THE SULFUR AMINO ACIDS.

    Science.gov (United States)

    PROTEINS, *TISSUES(BIOLOGY), METABOLISM, TISSUES(BIOLOGY), REGENERATION(ENGINEERING), WOUNDS AND INJURIES, TISSUES(BIOLOGY), TRACER STUDIES, METHIONINE, COLLAGEN, TYROSINE, BIOSYNTHESIS, AMINO ACIDS .

  7. Amino acid composition of Lagenaria siceraria seed flour and protein fractions.

    Science.gov (United States)

    Ogunbusola, Moriyike Esther; Fagbemi, Tayo Nathaniel; Osundahunsi, Oluwatooyin Faramade

    2010-12-01

    Defatted seed flours of Lagenaria siceraria (calabash and bottle gourd) were fractionated into their major protein fractions. The amino acid composition of seed flours and their protein fractions were determined and the protein quality was evaluated. Glutamic acid (139-168 mg/g protein) was the most abundant amino acid followed by aspartic acid (89.0-116 mg/g protein) in both the seed flours and their protein fractions. The total essential amino acid ranged from 45.8 to 51.5%. The predicted protein efficiency ratio and the predicted biological value ranged from 2.4 to 2.9 and 8.7 to 44.0, respectively. Lysine and sulphur amino acids were mostly concentrated in the globulin fractions. The first and second limiting amino acids in seed flours and protein fractions were methionine and valine or threonine. The seed flours contained adequate essential amino acids required by growing school children and adults. The seed has potential as protein supplement in cereal based complementary diets or in the replacement of animal proteins in conventional foods.

  8. S100B expression defines a state in which GFAP-expressing cells lose their neural stem cell potential and acquire a more mature developmental stage

    Science.gov (United States)

    Raponi, Eric; Agenes, Fabien; Delphin, Christian; Assard, Nicole; Baudier, Jacques; Legraverend, Catherine; Deloulme, Jean-Christophe

    2007-01-01

    Summary During the post-natal development, GFAP-expressing cells in the parenchyma progressively lose their neural stem cell (NSC) potential, whereas this peculiar attribute is preserved in GFAP-expressing cells of adult germinal zones. Although the relationship between astrocytes localized in the parenchyma and those in the germinal zones is elusive, many reports suggest that GFAP-expressing cells contained in germinal zones are maintained in immature developmental stage. Starting from the observation that the pan-astrocytic marker S100B is not expressed in the GFAP-expressing cells of adult germinal zones, we first investigated the relationship between S100B expression and the developmental status of these astrocytic cells. We demonstrated that long after the loss of RC2 and gain of GFAP, the onset of S100B expression characterizes a mature developmental stage in mouse telencephalic GFAP-expressing cells, both in vitro and in vivo. Using a transgenic s100b-EGFP mouse-derived culture model, we next demonstrate that in vitro, S100B expression in GFAP-expressing cells coincides with the loss of their NSC potential. Through a series of ectopic and orthotopic grafting experiments, we found that in the adult subventricular zone, S100B expression is controlled by environmental cues. Furthermore, we showed that treatment with epidermal growth factor represses S100B expression in GFAP-expressing cells in vitro as well as in mouse forebrain. Altogether, our results indicate that the S100B expression defines a late developmental stage after which GFAP-expressing cells lose their NSC potential. PMID:17078026

  9. Acidic ribosomal proteins and histone H3 from Leishmania present a high rate of divergence

    Directory of Open Access Journals (Sweden)

    Ysabel Montoya

    2000-08-01

    Full Text Available Another additional peculiarity in Leishmania will be discussed about of the amino acid divergence rate of three structural proteins: acidic ribosomal P1 and P2b proteins, and histone H3 by using multiple sequence alignment and dendrograms. These structural proteins present a high rate of divergence regarding to their homologous protein in Trypanosoma cruzi. At this regard, L. (V. peruviana P1 and T. cruzi P1 showed 57.4% of divergence rate. Likewise, L. (V. braziliensis histone H3 and acidic ribosomal P2 protein exhibited 31.8% and 41.7% respectively of rate of divergence in comparison with their homologous in T. cruzi.

  10. 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.

  11. Identification of Dynamic Changes in Proteins Associated with the Cellular Cytoskeleton after Exposure to Okadaic Acid

    Directory of Open Access Journals (Sweden)

    Peter Roepstorff

    2013-05-01

    Full Text Available Exposure of cells to the diarrhetic shellfish poison, okadaic acid, leads to a dramatic reorganization of cytoskeletal architecture and loss of cell-cell contact. When cells are exposed to high concentrations of okadaic acid (100–500 nM, the morphological rearrangement is followed by apoptotic cell death. Okadaic acid inhibits the broad acting Ser/Thr protein phosphatases 1 and 2A, which results in hyperphosphorylation of a large number of proteins. Some of these hyperphosphorylated proteins are most likely key players in the reorganization of the cell morphology induced by okadaic acid. We wanted to identify these phosphoproteins and searched for them in the cellular lipid rafts, which have been found to contain proteins that regulate cytoskeletal dynamics and cell adhesion. By using stable isotope labeling by amino acids in cell culture cells treated with okadaic acid (400 nM could be combined with control cells before the isolation of lipid rafts. Protein phosphorylation events and translocations induced by okadaic acid were identified by mass spectrometry. Okadaic acid was shown to regulate the phosphorylation status and location of proteins associated with the actin cytoskeleton, microtubules and cell adhesion structures. A large number of these okadaic acid-regulated proteins have previously also been shown to be similarly regulated prior to cell proliferation and migration. Our results suggest that okadaic acid activates general cell signaling pathways that induce breakdown of the cortical actin cytoskeleton and cell detachment.

  12. Synthesis of alanyl nucleobase amino acids and their incorporation into proteins.

    Science.gov (United States)

    Talukder, Poulami; Dedkova, Larisa M; Ellington, Andrew D; Yakovchuk, Petro; Lim, Jaebum; Anslyn, Eric V; Hecht, Sidney M

    2016-09-15

    Proteins which bind to nucleic acids and regulate their structure and functions are numerous and exceptionally important. Such proteins employ a variety of strategies for recognition of the relevant structural elements in their nucleic acid substrates, some of which have been shown to involve rather subtle interactions which might have been difficult to design from first principles. In the present study, we have explored the preparation of proteins containing unnatural amino acids having nucleobase side chains. In principle, the introduction of multiple nucleobase amino acids into the nucleic acid binding domain of a protein should enable these modified proteins to interact with their nucleic acid substrates using Watson-Crick and other base pairing interactions. We describe the synthesis of five alanyl nucleobase amino acids protected in a fashion which enabled their attachment to a suppressor tRNA, and their incorporation into each of two proteins with acceptable efficiencies. The nucleobases studied included cytosine, uracil, thymine, adenine and guanine, i.e. the major nucleobase constituents of DNA and RNA. Dihydrofolate reductase was chosen as one model protein to enable direct comparison of the facility of incorporation of the nucleobase amino acids with numerous other unnatural amino acids studied previously. The Klenow fragment of DNA polymerase I was chosen as a representative DNA binding protein whose mode of action has been studied in detail. Copyright © 2016 Elsevier Ltd. All rights reserved.

  13. Brain injury-associated biomarkers of TGF-beta1, S100B, GFAP, NF-L, tTG, AbetaPP, and tau were concomitantly enhanced and the UPS was impaired during acute brain injury caused by Toxocara canis in mice

    Directory of Open Access Journals (Sweden)

    Ji Dar-Der

    2008-06-01

    Full Text Available Abstract Background Because the outcomes and sequelae after different types of brain injury (BI are variable and difficult to predict, investigations on whether enhanced expressions of BI-associated biomarkers (BIABs, including transforming growth factor β1 (TGF-β1, S100B, glial fibrillary acidic protein (GFAP, neurofilament light chain (NF-L, tissue transglutaminases (tTGs, β-amyloid precursor proteins (AβPP, and tau are present as well as whether impairment of the ubiquitin-proteasome system (UPS is present have been widely used to help delineate pathophysiological mechanisms in various BIs. Larvae of Toxocara canis can invade the brain and cause BI in humans and mice, leading to cerebral toxocariasis (CT. Because the parasitic burden is light in CT, it may be too cryptic to be detected in humans, making it difficult to clearly understand the pathogenesis of subtle BI in CT. Since the pathogenesis of murine toxocariasis is very similar to that in humans, it appears appropriate to use a murine model to investigate the pathogenesis of CT. Methods BIAB expressions and UPS function in the brains of mice inoculated with a single dose of 250 T. canis embryonated eggs was investigated from 3 days (dpi to 8 weeks post-infection (wpi by Western blotting and RT-PCR. Results Results revealed that at 4 and 8 wpi, T. canis larvae were found to have invaded areas around the choroid plexus but without eliciting leukocyte infiltration in brains of infected mice; nevertheless, astrogliosis, an indicator of BI, with 78.9~142.0-fold increases in GFAP expression was present. Meanwhile, markedly increased levels of other BIAB proteins including TGF-β1, S100B, NF-L, tTG, AβPP, and tau, with increases ranging 2.0~12.0-fold were found, although their corresponding mRNA expressions were not found to be present at 8 wpi. Concomitantly, UPS impairment was evidenced by the overexpression of conjugated ubiquitin and ubiquitin in the brain. Conclusion Further studies

  14. Microwave-assisted acid and base hydrolysis of intact proteins containing disulfide bonds for protein sequence analysis by mass spectrometry.

    Science.gov (United States)

    Reiz, Bela; Li, Liang

    2010-09-01

    Controlled hydrolysis of proteins to generate peptide ladders combined with mass spectrometric analysis of the resultant peptides can be used for protein sequencing. In this paper, two methods of improving the microwave-assisted protein hydrolysis process are described to enable rapid sequencing of proteins containing disulfide bonds and increase sequence coverage, respectively. It was demonstrated that proteins containing disulfide bonds could be sequenced by MS analysis by first performing hydrolysis for less than 2 min, followed by 1 h of reduction to release the peptides originally linked by disulfide bonds. It was shown that a strong base could be used as a catalyst for microwave-assisted protein hydrolysis, producing complementary sequence information to that generated by microwave-assisted acid hydrolysis. However, using either acid or base hydrolysis, amide bond breakages in small regions of the polypeptide chains of the model proteins (e.g., cytochrome c and lysozyme) were not detected. Dynamic light scattering measurement of the proteins solubilized in an acid or base indicated that protein-protein interaction or aggregation was not the cause of the failure to hydrolyze certain amide bonds. It was speculated that there were some unknown local structures that might play a role in preventing an acid or base from reacting with the peptide bonds therein.

  15. Sex Steroid Modulation of Fatty Acid Utilization and Fatty Acid Binding Protein Concentration in Rat Liver

    Science.gov (United States)

    Ockner, Robert K.; Lysenko, Nina; Manning, Joan A.; Monroe, Scott E.; Burnett, David A.

    1980-01-01

    The mechanism by which sex steroids influence very low density hepatic lipoprotein triglyceride production has not been fully elucidated. In previous studies we showed that [14C]oleate utilization and incorporation into triglycerides were greater in hepatocyte suspensions from adult female rats than from males. The sex differences were not related to activities of the enzymes of triglyceride biosynthesis, whereas fatty acid binding protein (FABP) concentration in liver cytosol was greater in females. These findings suggested that sex differences in lipoprotein could reflect a sex steroid influence on the availability of fatty acids for hepatocellular triglyceride biosynthesis. In the present studies, sex steroid effects on hepatocyte [14C]oleate utilization and FABP concentration were investigated directly. Hepatocytes from immature (30-d-old) rats exhibited no sex differences in [14C]oleate utilization. With maturation, total [14C]oleate utilization and triglyceride biosynthesis increased moderately in female cells and decreased markedly in male cells; the profound sex differences in adults were maximal by age 60 d. Fatty acid oxidation was little affected. Rats were castrated at age 30 d, and received estradiol, testosterone, or no hormone until age 60 d, when hepatocyte [14C]oleate utilization was studied. Castration virtually eliminated maturational changes and blunted the sex differences in adults. Estradiol or testosterone largely reproduced the appropriate adult pattern of [14C]oleate utilization regardless of the genotypic sex of the treated animal. In immature females and males, total cytosolic FABP concentrations were similar. In 60-d-old animals, there was a striking correlation among all groups (females, males, castrates, and hormone-treated) between mean cytosolic FABP concentration on the one hand, and mean total [14C]oleate utilization (r = 0.91) and incorporation into triglycerides (r = 0.94) on the other. In 30-d-old animals rates of [14C

  16. Effects of Gingko biloba leaf extract on the learning and memory and expression of glial fibrillary acidic protein in hippocampal astrocytes of type 2 diabetic rats%银杏叶提取物对2型糖尿病大鼠学习记忆及海马胶质纤维酸性蛋白表达的影响

    Institute of Scientific and Technical Information of China (English)

    林军; 韦力; 张锡流; 舒丽雁

    2006-01-01

    BACKGROUND: Studies have shown that Gingko biloba leaf extract (GbE) is effective in promoting the functions recovery of the brain that follows traumatic injury, in improving the dysfunctions of learning and memory of the elderly, and it is also effective in improving the plasticity in central nervous system (CNS). However, what is the effect on learning and memory functions of rats with type 2 diabetes mellitus?OBJECTIVE: To study the effects of GbE on the learning and memory dysfunction and glial fibrillary acidic protein (GFAP) expressions in hippocampus of diabetic rats.DESIGN: Complete-random design, controlled experimental study.SETTING: Department of Pharmacology, Guangxi Medical University.MATERIALS: A total of 84 male Wistar rats (180-220 g), 8 weeks old,SPF, were used in this study. GbE (containing 24.8% flavone glycosides and 6.2% diterpene lactone) was purchased from Guilin Xintejia Natural plants Pharmaceutical Factory, Guangxi Province, Lot No. 200405.METHODS: The experiment was completed at the Pharmacology Lab (Provincial Lab) of the Experimental Center of Guangxi Medical University from June 2004 to March 2005. ① A total of 70 rats were rendered diabetic by a single intraperitoneal injection of streptozotocin at a dose of 55 rmg/kg dissolved in citrate buffer (pH4.4) after 24 hours fasting. Tail vein blood glucose concentration was determined 4 days later using ONE TOUCH glucose meter. A total of 56 streptozotocin-treated rats with a blood glucose concentration of > 15 mmol/L were recognized as type 2 diabetic rats. ② These diabetic rats were randomly divided into model group, insulin group, high-dose GbE group, and low-dose GbE group. There were 14 rats in each group. There was no difference in the blood glucose concentration among the groups. Another 14 male rats with an intraperitoneal injection of citrate buffer solution were served as control group. After division, drugs were given. Insulin 10 μ/kg was injected subcutaneously every day

  17. Although it is rapidly metabolized in cultured rat hepatocytes, lauric acid is used for protein acylation.

    Science.gov (United States)

    Rioux, Vincent; Daval, Stéphanie; Guillou, Hervé; Jan, Sophie; Legrand, Philippe

    2003-01-01

    This study was designed to examine the metabolic fate of exogenous lauric acid in cultured rat hepatocytes, in terms of both lipid metabolism and acylation of proteins. Radiolabeled [14C]-lauric acid at 0.1 mM in the culture medium was rapidly taken up by the cells (94.8 +/- 2.2% of the initial radioactivity was cleared from the medium after a 4 h incubation) but its incorporation into cellular lipids was low (24.6 +/- 4.2% of initial radioactivity after 4 h), due to the high beta-oxidation of lauric acid in hepatocytes (38.7 +/- 4.4% after the same time). Among cellular lipids, lauric acid was preferentially incorporated into triglycerides (10.6 +/- 4.6% of initial radioactivity after 4 h). Lauric acid was also rapidly converted to palmitic acid by two successive elongations. Protein acylation was detected after metabolic labeling of the cells with [11,12-3H]-lauric acid. Two-dimensional electrophoresis separation of the cellular proteins and autoradiography evidenced the incorporation of radioactivity into 35 well-resolved proteins. Radiolabeling of several proteins resulted from covalent linkage to the precursor [11,12-3H]-lauric acid or to its elongation product, myristic acid. The covalent linkages between these proteins and lauric acid were broken by base hydrolysis, indicating that the linkage was of the thioester or ester-type. Endogenous myristic acid produced by lauric acid elongation was used for both protein N-myristoylation and protein S-acylation. Therefore, these results show for the first time that, although it is rapidly metabolized in hepatocytes, exogenous lauric acid is a substrate for the acylation of liver proteins.

  18. AMINO ACID METABOLISM IN COWS DURING THE TRANSITION PERIOD IN BALANCING DIET ON THE EXCHANGE PROTEIN AND DIGESTIBLE AMINO ACIDS

    Directory of Open Access Journals (Sweden)

    Ryadchikov V. G.

    2014-02-01

    Full Text Available Application of a factorial method for determining the needs in metabolic protein and essential amino acids, helps to deepen knowledge on physiology of protein and amino acid supply and allow to improve the standards for dairy cows during the transition period; in insufficient of metabolic protein and essential amino acids increased coefficients of their transformation into net protein and absorptive amino acids as a result of mobilization of body of cows; with an optimal protein nutrition their transformation in net milk protein, lysine and methionine accordingly amounted to 0.67, 0,83 and 0,82. The most significant changes in the concentration of methionine, proline, glutamate, glutamine, glycine were observed in cows before calving and immediately after birth, stabilization of their level starts with a 24 lactation day, that is connected with the peculiarities of the feeding behavior of the cows and the gradual intensification of the processes of metabolism and milk production. To control the status of protein metabolism we have offered benchmarks compositions of free amino acids in cows’ blood plasma phases: 21-0 days before calving, 0-21 and 22-120 days after calving

  19. Comparison of Bile Acids and Acetaminophen Protein Adducts in Children and Adolescents with Acetaminophen Toxicity.

    Science.gov (United States)

    James, Laura; Yan, Ke; Pence, Lisa; Simpson, Pippa; Bhattacharyya, Sudeepa; Gill, Pritmohinder; Letzig, Lynda; Kearns, Gregory; Beger, Richard

    2015-01-01

    Metabolomics approaches have enabled the study of new mechanisms of liver injury in experimental models of drug toxicity. Disruption of bile acid homeostasis is a known mechanism of drug induced liver injury. The relationship of individual bile acids to indicators of oxidative drug metabolism (acetaminophen protein adducts) and liver injury was examined in children with acetaminophen overdose, hospitalized children with low dose exposure to acetaminophen, and children with no recent exposure to acetaminophen. Nine bile acids were quantified through targeted metabolomic analysis in the serum samples of the three groups. Bile acids were compared to serum levels of acetaminophen protein adducts and alanine aminotransferase. Glycodeoxycholic acid, taurodeoxycholic acid, and glycochenodeoxycholic acid were significantly increased in children with acetaminophen overdose compared to healthy controls. Among patients with acetaminophen overdose, bile acids were higher in subjects with acetaminophen protein adduct values > 1.0 nmol/mL and modest correlations were noted for three bile acids and acetaminophen protein adducts as follows: taurodeoxycholic acid (R=0.604; pacetaminophen than in healthy children with no recent acetaminophen exposure. Compared to bile acids, acetaminophen protein adducts more accurately discriminated among children with acetaminophen overdose, children with low dose exposure to acetaminophen, and healthy control subjects. In children with acetaminophen overdose, elevations of conjugated bile acids were associated with specific indicators of acetaminophen metabolism and non-specific indicators of liver injury.

  20. Scale-free behaviour of amino acid pair interactions in folded proteins

    DEFF Research Database (Denmark)

    Petersen, Steffen B.; Neves-Petersen, Maria Teresa; Mortensen, Rasmus J.

    2012-01-01

    that they are in buried a-helices or b-strands, in a spatial distance of 3.8–4.3A° and in a sequence distance .4 residues. We speculate that the scale free organization of the amino acid pair interactions in the 8D protein structure combined with the clear dominance of pairs of Ala, Ile, Leu and Val is important......The protein structure is a cumulative result of interactions between amino acid residues interacting with each other through space and/or chemical bonds. Despite the large number of high resolution protein structures, the ‘‘protein structure code’’ has not been fully identified. Our manuscript...... presents a novel approach to protein structure analysis in order to identify rules for spatial packing of amino acid pairs in proteins. We have investigated 8706 high resolution non-redundant protein chains and quantified amino acid pair interactions in terms of solvent accessibility, spatial and sequence...

  1. Amino acid absorption and subsequent muscle protein accretion following graded intakes of whey protein in elderly men.

    Science.gov (United States)

    Pennings, Bart; Groen, Bart; de Lange, Anneke; Gijsen, Annemie P; Zorenc, Antoine H; Senden, Joan M G; van Loon, Luc J C

    2012-04-15

    Whey protein ingestion has been shown to effectively stimulate postprandial muscle protein accretion in older adults. However, the impact of the amount of whey protein ingested on protein digestion and absorption kinetics, whole body protein balance, and postprandial muscle protein accretion remains to be established. We aimed to fill this gap by including 33 healthy, older men (73 ± 2 yr) who were randomly assigned to ingest 10, 20, or 35 g of intrinsically l-[1-¹³C]phenylalanine-labeled whey protein (n = 11/treatment). Ingestion of labeled whey protein was combined with continuous intravenous l-[ring-²H₅]phenylalanine and l-[ring-²H₂]tyrosine infusion to assess the metabolic fate of whey protein-derived amino acids. Dietary protein digestion and absorption rapidly increased following ingestion of 10, 20, and 35 g whey protein, with the lowest and highest (peak) values observed following 10 and 35 g, respectively (P whey protein results in greater amino acid absorption and subsequent stimulation of de novo muscle protein synthesis compared with the ingestion of 10 or 20 g whey protein in healthy, older men.

  2. Toward amino acid typing for proteins in FFLUX.

    Science.gov (United States)

    Fletcher, Timothy L; Popelier, Paul L A

    2017-03-05

    Continuing the development of the FFLUX, a multipolar polarizable force field driven by machine learning, we present a modern approach to atom-typing and building transferable models for predicting atomic properties in proteins. Amino acid atomic charges in a peptide chain respond to the substitution of a neighboring residue and this response can be categorized in a manner similar to atom-typing. Using a machine learning method called kriging, we are able to build predictive models for an atom that is defined, not only by its local environment, but also by its neighboring residues, for a minimal additional computational cost. We found that prediction errors were up to 11 times lower when using a model specific to the correct group of neighboring residues, with a mean prediction of ∼0.0015 au. This finding suggests that atoms in a force field should be defined by more than just their immediate atomic neighbors. When comparing an atom in a single alanine to an analogous atom in a deca-alanine helix, the mean difference in charge is 0.026 au. Meanwhile, the same difference between a trialanine and a deca-alanine helix is only 0.012 au. When compared to deca-alanine models, the transferable models are up to 20 times faster to train, and require significantly less ab initio calculation, providing a practical route to modeling large biological systems. © 2016 The Authors. Journal of Computational Chemistry Published by Wiley Periodicals, Inc.

  3. The amino acid sequence of protein CM-3 from Dendroaspis polylepis polylepis (black mamba) venom.

    Science.gov (United States)

    Joubert, F J

    1985-01-01

    Protein CM-3 from Dendroaspis polylepis polylepis venom was purified by gel filtration and ion exchange chromatography. It comprises 65 amino acids including eight half-cystines. The complete amino acid sequence of protein CM-3 has been elucidated. The sequence (residues 1-50) resembles that of the N-terminal sequence of the subunits of a synergistic type protein and residues 51-65 that of the C-terminal sequence of an angusticeps type protein. Mixtures of protein CM-3 and angusticeps type proteins showed no apparent synergistic effect, in that their toxicity in combination was no greater than the sum of their individual toxicities.

  4. A biotin enrichment strategy identifies novel carbonylated amino acids in proteins from human plasma

    DEFF Research Database (Denmark)

    Havelund, Jesper F.; Wojdyla, Katarzyna; Davies, Michael J.

    2017-01-01

    Protein carbonylation is an irreversible protein oxidation correlated with oxidative stress, various diseases and ageing. Here we describe a peptide-centric approach for identification and characterisation of up to 14 different types of carbonylated amino acids in proteins. The modified residues...... in vitro metal ion-catalysed oxidation. Furthermore, we assigned 133 carbonylated sites in 36 proteins in native human plasma protein samples. The optimised workflow enabled detection of 10 hitherto undetected types of carbonylated amino acids in proteins: aldehyde and ketone modifications of leucine...

  5. Effects of antinutritional factors on protein digestibility and amino acid availability in foods.

    Science.gov (United States)

    Gilani, G Sarwar; Cockell, Kevin A; Sepehr, Estatira

    2005-01-01

    Digestibility of protein in traditional diets from developing countries such as India, Guatemala, and Brazil is considerably lower compared to that of protein in typical North American diets (54-78 versus 88-94%). The presence of less digestible protein fractions, high levels of insoluble fiber, and high concentrations of antinutritional factors in the diets of developing countries, which are based on less refined cereals and grain legumes as major sources of protein, are responsible for poor digestibility of protein. The effects of the presence of some of the important antinutritional factors on protein and amino digestibilities of food and feed products are reviewed in this chapter. Food and feed products may contain a number of antinutritional factors that may adversely affect protein digestibility and amino acid availability. Antinutritional factors may occur naturally, such as glucosinolates in mustard and rapeseed protein products, trypsin inhibitors and hemagglutinins in legumes, tannins in legumes and cereals, phytates in cereals and oilseeds, and gossypol in cottonseed protein products. Antinutritional factors may also be formed during heat/alkaline processing of protein products, yielding Maillard compounds, oxidized forms of sulfur amino acids, D-amino acids, and lysinoalanine (LAL, an unnatural amino acid derivative). The presence of high levels of dietary trypsin inhibitors from soybeans, kidney beans, or other grain legumes can cause substantial reductions in protein and amino acid digestibilities (up to 50%) in rats and pigs. Similarly, the presence of high levels of tannins in cereals, such as sorghum, and grain legumes, such as fababean (Vicia faba L.), can result in significantly reduced protein and amino acid digestibilities (up to 23%) in rats, poultry, and pigs. Studies involving phytase supplementation of production rations for swine or poultry have provided indirect evidence that normally encountered levels of phytates in cereals and legumes

  6. Four Proteins Synthesized in Response to Deoxyribonucleic Acid Damage in Micrococcus Radiodurans

    DEFF Research Database (Denmark)

    Hansen, M. T.

    1980-01-01

    or by starvation for thymine, failed to elicit the synthesis of either protein. Repair of deoxyribonucleic acid damage requires that a number of versatile and efficient processes by employed. It is proposed that the induced proteins participate in deoxyribonucleic acid repair in M. radiodurans. Mechanisms...

  7. Structure and rheological properties of acid-induced egg white protein gels

    NARCIS (Netherlands)

    Weijers, M.; Velde, van de F.; Stijnman, A.; Pijpekamp, van de A.; Visschers, R.W.

    2006-01-01

    This study compares the rheological properties of acid-induced gels prepared of industrial spray-dried egg white proteins (EWP) with the acid-induced gels prepared of ovalbumin (OA) and whey protein isolate (WPI). Also we aimed to form transparent gels of EWP by means of the cold-gelation process. W

  8. The phosphorylation status and cytoskeletal remodeling of striatal astrocytes treated with quinolinic acid

    Energy Technology Data Exchange (ETDEWEB)

    Pierozan, Paula; Ferreira, Fernanda; Ortiz de Lima, Bárbara; Gonçalves Fernandes, Carolina [Departamento de Bioquímica, Instituto de Ciências Básicas da Saúde, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS 90035-003 (Brazil); Totarelli Monteforte, Priscila; Castro Medaglia, Natalia de; Bincoletto, Claudia; Soubhi Smaili, Soraya [Departamento de Farmacologia, Universidade Federal de São Paulo (UNIFESP/EPM), São Paulo, SP (Brazil); Pessoa-Pureur, Regina, E-mail: rpureur@ufrgs.br [Departamento de Bioquímica, Instituto de Ciências Básicas da Saúde, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS 90035-003 (Brazil)

    2014-04-01

    Quinolinic acid (QUIN) is a glutamate agonist which markedly enhances the vulnerability of neural cells to excitotoxicity. QUIN is produced from the amino acid tryptophan through the kynurenine pathway (KP). Dysregulation of this pathway is associated with neurodegenerative conditions. In this study we treated striatal astrocytes in culture with QUIN and assayed the endogenous phosphorylating system associated with glial fibrillary acidic protein (GFAP) and vimentin as well as cytoskeletal remodeling. After 24 h incubation with 100 µM QUIN, cells were exposed to {sup 32}P-orthophosphate and/or protein kinase A (PKA), protein kinase dependent of Ca{sup 2+}/calmodulin II (PKCaMII) or protein kinase C (PKC) inhibitors, H89 (20 μM), KN93 (10 μM) and staurosporin (10 nM), respectively. Results showed that hyperphosphorylation was abrogated by PKA and PKC inhibitors but not by the PKCaMII inhibitor. The specific antagonists to ionotropic NMDA and non-NMDA (50 µM DL-AP5 and CNQX, respectively) glutamate receptors as well as to metabotropic glutamate receptor (mGLUR; 50 µM MCPG), mGLUR1 (100 µM MPEP) and mGLUR5 (10 µM 4C3HPG) prevented the hyperphosphorylation provoked by QUIN. Also, intra and extracellular Ca{sup 2+} quelators (1 mM EGTA; 10 µM BAPTA-AM, respectively) prevented QUIN-mediated effect, while Ca{sup 2+} influx through voltage-dependent Ca{sup 2+} channel type L (L-VDCC) (blocker: 10 µM verapamil) is not implicated in this effect. Morphological analysis showed dramatically altered actin cytoskeleton with concomitant change of morphology to fusiform and/or flattened cells with retracted cytoplasm and disruption of the GFAP meshwork, supporting misregulation of actin cytoskeleton. Both hyperphosphorylation and cytoskeletal remodeling were reversed 24 h after QUIN removal. Astrocytes are highly plastic cells and the vulnerability of astrocyte cytoskeleton may have important implications for understanding the neurotoxicity of QUIN in neurodegenerative

  9. Site specific incorporation of heavy atom-containing unnatural amino acids into proteins for structure determination

    Science.gov (United States)

    Xie, Jianming; Wang, Lei; Wu, Ning; Schultz, Peter G.

    2008-07-15

    Translation systems and other compositions including orthogonal aminoacyl tRNA-synthetases that preferentially charge an orthogonal tRNA with an iodinated or brominated amino acid are provided. Nucleic acids encoding such synthetases are also described, as are methods and kits for producing proteins including heavy atom-containing amino acids, e.g., brominated or iodinated amino acids. Methods of determining the structure of a protein, e.g., a protein into which a heavy atom has been site-specifically incorporated through use of an orthogonal tRNA/aminoacyl tRNA-synthetase pair, are also described.

  10. Analysis of the protein-protein interactions between the human acidic ribosomal P-proteins: evaluation by the two hybrid system

    DEFF Research Database (Denmark)

    Tchórzewski, M; Boldyreff, B; Issinger, O

    2000-01-01

    on the function of these proteins, we are the first to have precisely analyzed mutual interactions among human P-proteins, employing the two hybrid system. The human acidic ribosomal P-proteins, (P1 or P2,) were fused to the GAL4 binding domain (BD) as well as the activation domain (AD), and analyzed in yeast...

  11. Influence of dietary protein type and iron source on the absorption of amino acids and minerals.

    Science.gov (United States)

    Pérez-Llamas, F; Garaulet, M; Martínez, J A; Marín, J F; Larqué, E; Zamora, S

    2001-12-01

    The apparent digestibility coefficient (ADC) of amino acids and the balance of minerals (calcium, phosphorus, magnesium and iron) has been determined in rats fed four diets differing in the protein type (casein or soy protein) and iron source (ferrous sulphate or lactate) in order to study the possible interactions of these nutrients. The availability of amino acids, especially essential amino acids, was greater in the diet made with animal protein (casein). The iron source also affected the absorption of most amino acids in all the diets assayed with ferrous sulphate being greater. The balance of iron, magnesium and phosphorus was higher in the diets containing animal protein. The retention of calcium and magnesium was significantly greater when ferrous sulphate was used as iron source. These results demonstrate the important interaction between amino acids and minerals and between the minerals themselves, which must be carefully studied when selecting different types of protein or mineral sources in human or animal nutrition.

  12. Correlating dynamic amino acid properties with success rate of crystallization of proteins from Bacteroides vulgatus

    Energy Technology Data Exchange (ETDEWEB)

    Yan, Shaomin [State Key Laboratory of Non-food Biomass Enzyme Technology, National Engineering Research Center for Non-food Biorefinery, Guangxi Key Laboratory of Biorefinery, Guangxi Academy of Sciences, 98 Daling Road, Nanning, Guangxi, 530007 (China); Wu, Guang [State Key Laboratory of Non-food Biomass Enzyme Technology, National Engineering Research Center for Non-food Biorefinery, Guangxi Key Laboratory of Biorefinery, Guangxi Academy of Sciences, 98 Daling Road, Nanning, Guangxi, 530007 (China); DreamSciTech Consulting, 301, Building 12, Nanyou A-zone, Jiannan Road, Shenzhen, Guangdong, 518054 (China)

    2012-05-15

    To enhance the success rate of protein crystallization, many studies were conducted to determine the relationship between amino acid properties and the success rate of protein crystallization. Although those were successful, new efforts should be made to search for the new factors, which affect protein crystallization. In this study, two dynamic amino acid properties were used to correlate with the success rate of crystallization of proteins from Bacteroides vulgatus, because the amino acid properties used in previous studies were steady. As previously done, logistic regression and neural network were used to model that relationship, and the results were compared against those obtained from each of 532 amino acid properties, which severed as benchmark. The results demonstrated that dynamic amino acid properties should be taken into consideration of protein crystallization. (copyright 2012 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim) (orig.)

  13. Chemical aminoacylation of tRNAs with fluorinated amino acids for in vitro protein mutagenesis

    Directory of Open Access Journals (Sweden)

    Shijie Ye

    2010-04-01

    Full Text Available This article describes the chemical aminoacylation of the yeast phenylalanine suppressor tRNA with a series of amino acids bearing fluorinated side chains via the hybrid dinucleotide pdCpA and ligation to the corresponding truncated tRNA species. Aminoacyl-tRNAs can be used to synthesize biologically relevant proteins which contain fluorinated amino acids at specific sites by means of a cell-free translation system. Such engineered proteins are expected to contribute to our understanding of discrete fluorines’ interaction with canonical amino acids in a native protein environment and to enable the design of fluorinated proteins with arbitrary desired properties.

  14. Lipid recognition propensities of amino acids in membrane proteins from atomic resolution data

    Directory of Open Access Journals (Sweden)

    Morita Mizuki

    2011-12-01

    Full Text Available Abstract Background Protein-lipid interactions play essential roles in the conformational stability and biological functions of membrane proteins. However, few of the previous computational studies have taken into account the atomic details of protein-lipid interactions explicitly. Results To gain an insight into the molecular mechanisms of the recognition of lipid molecules by membrane proteins, we investigated amino acid propensities in membrane proteins for interacting with the head and tail groups of lipid molecules. We observed a common pattern of lipid tail-amino acid interactions in two different data sources, crystal structures and molecular dynamics simulations. These interactions are largely explained by general lipophilicity, whereas the preferences for lipid head groups vary among individual proteins. We also found that membrane and water-soluble proteins utilize essentially an identical set of amino acids for interacting with lipid head and tail groups. Conclusions We showed that the lipophilicity of amino acid residues determines the amino acid preferences for lipid tail groups in both membrane and water-soluble proteins, suggesting that tightly-bound lipid molecules and lipids in the annular shell interact with membrane proteins in a similar manner. In contrast, interactions between lipid head groups and amino acids showed a more variable pattern, apparently constrained by each protein's specific molecular function.

  15. Effects of gabapentin combined with morphine on expression of glial fibillary acid protein of spinal dorsal horn in rats with neuropathic pain%加巴喷丁联合吗啡对神经病理性疼痛大鼠脊髓胶质原纤维酸性蛋白表达的影响

    Institute of Scientific and Technical Information of China (English)

    刘红; 万朝权; 田可耘; 梅莉; 崔灿; 张晓晨

    2012-01-01

    目的:观察加巴喷丁联合吗啡镇痛对坐骨神经慢性压迫(CCI)大鼠脊髓背角胶质原纤维酸性蛋白(GFAP)表达的影响.方法:选择重量在180~220 g的成年雄性SD大鼠24只,随机分为4组(n=6):假手术组(S组)、模型组(M组)、预防性镇痛组(P组)和常规镇痛组(N组).M组、P组和N组采用CCI建立神经病理性疼痛模型,P组在术前1 d至手术后9 d使用加巴喷丁联合吗啡镇痛,N组在手术当日至手术后第9天加巴喷丁联合吗啡镇痛,S组和M组仅给予生理盐水.分别于手术后第3、5、7、9天测定各组50%机械刺激缩足阈值,手术后第10天采用免疫组化测定脊髓GFAP表达.结果:与M组相比,P组和N组50%缩足阈值升高,同时脊髓背角GFAP表达降低(M组:0.623 7±0.049 03,P组:0.461 6±0.038 90,N组:0.521 5±0.026 91).结论:加巴喷丁联合吗啡可以减轻CCI神经病理性疼痛和脊髓背角GFAP的表达.%Objective To observe the effects of gabapentin combined with morphine on expression of glial fibillary acid protein (GFAP) of spinal dorsal horn in rats model of chronic constriction injury (CCI) to sciatic nerve. Methods Twenty four male Sprague-Dawley rats were randomly divided into sham operation group (group S), model group (group M), preventive analgesia group (group P) and normal analgesia group (group N), and were subjected to CCI (group M, P, N) or sham surgery (group S). Gabapentin combined with morphine were given from pre-operative day 1 to postoperative day 9 in group P, and in group N they were given from operative day to postoperative day 9. The group M and group S were given normal saline. Mechanical withdraw threshold of the rats were measured with Von Frey hair on pre-operative day 1 and postoperative day 3, 5, 7, 9 respectively. The expression of GFAP in spinal dorsal horn was assessed by immunohistochemistry at postoperative day 10. Results After operation, mechanical withdraw threshold decreased markedly in group M. Compared

  16. Molecular mechanism of recombinant liver fatty acid binding protein's antioxidant activity

    OpenAIRE

    YAN, JING; Gong, Yuewen; She, Yi-Min; Wang, Guqi; Roberts, Michael S; Burczynski, Frank J.

    2009-01-01

    Hepatocytes expressing liver fatty acid binding protein (L-FABP) are known to be more resistant to oxidative stress than those devoid of this protein. The mechanism for the observed antioxidant activity is not known. We examined the antioxidant mechanism of a recombinant rat L-FABP in the presence of a hydrophilic (AAPH) or lipophilic (AMVN) free radical generator. Recombinant L-FABP amino acid sequence and its amino acid oxidative products following oxidation were identified by MALDI quadrup...

  17. Amino Acid Molecular Units: Building Primary and Secondary Protein Structures

    Directory of Open Access Journals (Sweden)

    Aparecido R. Silva

    2008-05-01

    Full Text Available In order to guarantee the learning quality and suitable knowledge  use  about structural biology, it is fundamental to  exist, since the beginning of  students’ formation, the possibility of clear visualization of biomolecule structures. Nevertheless, the didactic books can only bring  schematic  drawings; even more elaborated figures and graphic computation  do not permit the necessary interaction.  The representation of three-dimensional molecular structures with ludic models, built with representative units, have supplied to the students and teachers a successfully experience to  visualize such structures and correlate them to the real molecules.  The design and applicability of the representative units were discussed with researchers and teachers before mould implementation.  In this stage  it  will be presented the  developed  kit  containing the  representative  plastic parts of the main amino acids.  The kit can demonstrate the interaction among the amino acids  functional groups  (represented by colors, shapes,  sizes and  the peptidic bonds between them  facilitating the assembly and visuali zation of the primary and secondary protein structure.  The models were designed for  Ca,  amino,  carboxyl groups  and  hydrogen. The  lateral chains have  well defined models that represent their geometrical shape.  The completed kit set  will be presented in this meeting (patent requested.  In the last phase of the project will be realized  an effective evaluation  of the kit  as a facilitative didactic tool of the teaching/learning process in the Structural Molecular Biology area.

  18. Cellular fatty acid composition, protein profile and antimicrobial activity of Bacillus sp., isolated from fish gut

    Institute of Scientific and Technical Information of China (English)

    Pushparaj Sujith; Baskaran Rohini; Singaram Jayalakshmi

    2014-01-01

    Objective: To purify and partially characterize the antimicrobial compounds from bacteriaBacillus sp., isolated from fish gut. Methods: Protein and fatty acids were isolated from the bacteria and checked for the presence of antibacterial activity. Protein has been purified to apparent homogeneity from the supernatants of culture by means of ammonium sulphate precipitation followed by dialysis. Fourier transform infrared spectroscopy analyses were performed for proteins to identify the functional groups.Results:sulfate polyacrylamide gel electrophoresis. Fatty acids were extracted and subjected to gas chromatographic analysis.Conclusions:Protein showed an apparent molecular mass 56, 47 and 39 kDa on sodium dodecyl acids and proteins which holds promise for the development of new drugs. The antimicrobial activity of the bacteria might be due to the presence of fatty acids and proteins which holds promise for the development of new drugs.

  19. Values for digestible indispensable amino acid scores (DIAAS) for some dairy and plant proteins may better describe protein quality than values calculated using the concept for protein digestibility-corrected amino acid scores (PDCAAS).

    Science.gov (United States)

    Mathai, John K; Liu, Yanhong; Stein, Hans H

    2017-02-01

    An experiment was conducted to compare values for digestible indispensable amino acid scores (DIAAS) for four animal proteins and four plant proteins with values calculated as recommended for protein digestibility-corrected amino acid scores (PDCAAS), but determined in pigs instead of in rats. Values for standardised total tract digestibility (STTD) of crude protein (CP) and standardised ileal digestibility (SID) of amino acids (AA) were calculated for whey protein isolate (WPI), whey protein concentrate (WPC), milk protein concentrate (MPC), skimmed milk powder (SMP), pea protein concentrate (PPC), soya protein isolate (SPI), soya flour and whole-grain wheat. The PDCAAS-like values were calculated using the STTD of CP to estimate AA digestibility and values for DIAAS were calculated from values for SID of AA. Results indicated that values for SID of most indispensable AA in WPI, WPC and MPC were greater (Pproteins.

  20. Influence of Bleaching on Flavor of 34% Whey Protein Concentrate and Residual Benzoic Acid Concentration in Dried Whey Proteins

    Science.gov (United States)

    Previous studies have shown that bleaching negatively affects the flavor of 70% whey protein concentrate (WPC70), but bleaching effects on lower-protein products have not been established. Benzoyl peroxide (BP), a whey bleaching agent, degrades to benzoic acid (BA) and may elevate BA concentrations...

  1. Influence of protein source on amino acid uptake patterns and protein utilization in rainbow trout Oncorhynchus mykiss

    DEFF Research Database (Denmark)

    Rolland, Marine; Holm, Jørgen; Dalsgaard, Anne Johanne Tang

    Matrixes of different protein sources (fish and plant products) combined with the use of crystalline amino acids allow for formulation of diets that meet fish requirements with little or no effect on protein digestibility and/or feed intake. Despite this, a total or partial replacement of fish meal...... induces reduced growth performances that remain partly unexplained. The aim of the current study was to investigate the effect of exchanging the protein source on protein utilization. Marine (fish meal) and vegetable (pea protein) sources were used with or without supplementation of crystalline amino...

  2. Microwave-assisted Weak Acid Hydrolysis of Proteins

    Directory of Open Access Journals (Sweden)

    Miyeong Seo

    2012-06-01

    Full Text Available Myoglobin was hydrolyzed by microwave-assisted weak acid hydrolysis with 2% formic acid at 37 oC, 50 oC, and100 oC for 1 h. The most effective hydrolysis was observed at 100 oC. Hydrolysis products were investigated using matrixassistedlaser desorption/ionization time-of-flight mass spectrometry. Most cleavages predominantly occurred at the C-termini ofaspartyl residues. For comparison, weak acid hydrolysis was also performed in boiling water for 20, 40, 60, and 120 min. A 60-min weak acid hydrolysis in boiling water yielded similar results as a 60-min microwave-assisted weak acid hydrolysis at100 oC. These results strongly suggest that microwave irradiation has no notable enhancement effect on acid hydrolysis of proteinsand that temperature is the major factor that determines the effectiveness of weak acid hydrolysis.

  3. Comparison of Bile Acids and Acetaminophen Protein Adducts in Children and Adolescents with Acetaminophen Toxicity.

    Directory of Open Access Journals (Sweden)

    Laura James

    Full Text Available Metabolomics approaches have enabled the study of new mechanisms of liver injury in experimental models of drug toxicity. Disruption of bile acid homeostasis is a known mechanism of drug induced liver injury. The relationship of individual bile acids to indicators of oxidative drug metabolism (acetaminophen protein adducts and liver injury was examined in children with acetaminophen overdose, hospitalized children with low dose exposure to acetaminophen, and children with no recent exposure to acetaminophen. Nine bile acids were quantified through targeted metabolomic analysis in the serum samples of the three groups. Bile acids were compared to serum levels of acetaminophen protein adducts and alanine aminotransferase. Glycodeoxycholic acid, taurodeoxycholic acid, and glycochenodeoxycholic acid were significantly increased in children with acetaminophen overdose compared to healthy controls. Among patients with acetaminophen overdose, bile acids were higher in subjects with acetaminophen protein adduct values > 1.0 nmol/mL and modest correlations were noted for three bile acids and acetaminophen protein adducts as follows: taurodeoxycholic acid (R=0.604; p<0.001, glycodeoxycholic acid (R=0.581; p<0.001, and glycochenodeoxycholic acid (R=0.571; p<0.001. Variability in bile acids was greater among hospitalized children receiving low doses of acetaminophen than in healthy children with no recent acetaminophen exposure. Compared to bile acids, acetaminophen protein adducts more accurately discriminated among children with acetaminophen overdose, children with low dose exposure to acetaminophen, and healthy control subjects. In children with acetaminophen overdose, elevations of conjugated bile acids were associated with specific indicators of acetaminophen metabolism and non-specific indicators of liver injury.

  4. Optimizing Scoring Function of Protein-Nucleic Acid Interactions with Both Affinity and Specificity

    Science.gov (United States)

    Yan, Zhiqiang; Wang, Jin

    2013-01-01

    Protein-nucleic acid (protein-DNA and protein-RNA) recognition is fundamental to the regulation of gene expression. Determination of the structures of the protein-nucleic acid recognition and insight into their interactions at molecular level are vital to understanding the regulation function. Recently, quantitative computational approach has been becoming an alternative of experimental technique for predicting the structures and interactions of biomolecular recognition. However, the progress of protein-nucleic acid structure prediction, especially protein-RNA, is far behind that of the protein-ligand and protein-protein structure predictions due to the lack of reliable and accurate scoring function for quantifying the protein-nucleic acid interactions. In this work, we developed an accurate scoring function (named as SPA-PN, SPecificity and Affinity of the Protein-Nucleic acid interactions) for protein-nucleic acid interactions by incorporating both the specificity and affinity into the optimization strategy. Specificity and affinity are two requirements of highly efficient and specific biomolecular recognition. Previous quantitative descriptions of the biomolecular interactions considered the affinity, but often ignored the specificity owing to the challenge of specificity quantification. We applied our concept of intrinsic specificity to connect the conventional specificity, which circumvents the challenge of specificity quantification. In addition to the affinity optimization, we incorporated the quantified intrinsic specificity into the optimization strategy of SPA-PN. The testing results and comparisons with other scoring functions validated that SPA-PN performs well on both the prediction of binding affinity and identification of native conformation. In terms of its performance, SPA-PN can be widely used to predict the protein-nucleic acid structures and quantify their interactions. PMID:24098651

  5. Genetically programmed expression of proteins containing the unnatural amino acid phenylselenocysteine

    Science.gov (United States)

    Wang, Jiangyun; Schultz, Peter G.

    2010-09-07

    The invention relates to orthogonal pairs of tRNAs and aminoacyl-tRNA synthetases that can incorporate the unnatural amino acid phenylselenocysteine into proteins produced in eubacterial host cells such as E. coli. The invention provides, for example but not limited to, novel orthogonal aminoacyl-tRNA synthetases, polynucleotides encoding the novel synthetase molecules, methods for identifying and making the novel synthetases, methods for producing proteins containing the unnatural amino acid phenylselenocysteine and translation systems. The invention further provides methods for producing modified proteins (e.g., lipidated proteins) through targeted modification of the phenylselenocysteine residue in a protein.

  6. Constraints and consequences of the emergence of amino acid repeats in eukaryotic proteins.

    Science.gov (United States)

    Chavali, Sreenivas; Chavali, Pavithra L; Chalancon, Guilhem; de Groot, Natalia Sanchez; Gemayel, Rita; Latysheva, Natasha S; Ing-Simmons, Elizabeth; Verstrepen, Kevin J; Balaji, Santhanam; Babu, M Madan

    2017-09-01

    Proteins with amino acid homorepeats have the potential to be detrimental to cells and are often associated with human diseases. Why, then, are homorepeats prevalent in eukaryotic proteomes? In yeast, homorepeats are enriched in proteins that are essential and pleiotropic and that buffer environmental insults. The presence of homorepeats increases the functional versatility of proteins by mediating protein interactions and facilitating spatial organization in a repeat-dependent manner. During evolution, homorepeats are preferentially retained in proteins with stringent proteostasis, which might minimize repeat-associated detrimental effects such as unregulated phase separation and protein aggregation. Their presence facilitates rapid protein divergence through accumulation of amino acid substitutions, which often affect linear motifs and post-translational-modification sites. These substitutions may result in rewiring protein interaction and signaling networks. Thus, homorepeats are distinct modules that are often retained in stringently regulated proteins. Their presence facilitates rapid exploration of the genotype-phenotype landscape of a population, thereby contributing to adaptation and fitness.

  7. Crystal structure of axolotl (Ambystoma mexicanum) liver bile acid-binding protein bound to cholic and oleic acid.

    Science.gov (United States)

    Capaldi, Stefano; Guariento, Mara; Perduca, Massimiliano; Di Pietro, Santiago M; Santomé, José A; Monaco, Hugo L

    2006-07-01

    The family of the liver bile acid-binding proteins (L-BABPs), formerly called liver basic fatty acid-binding proteins (Lb-FABPs) shares fold and sequence similarity with the paralogous liver fatty acid-binding proteins (L-FABPs) but has a different stoichiometry and specificity of ligand binding. This article describes the first X-ray structure of a member of the L-BABP family, axolotl (Ambystoma mexicanum) L-BABP, bound to two different ligands: cholic and oleic acid. The protein binds one molecule of oleic acid in a position that is significantly different from that of either of the two molecules that bind to rat liver FABP. The stoichiometry of binding of cholate is of two ligands per protein molecule, as observed in chicken L-BABP. The cholate molecule that binds buried most deeply into the internal cavity overlaps well with the analogous bound to chicken L-BABP, whereas the second molecule, which interacts with the first only through hydrophobic contacts, is more external and exposed to the solvent.

  8. Effects of long-term protein restriction on meat quality, muscle amino acids, and amino acid transporters in pigs.

    Science.gov (United States)

    Yin, Jie; Li, Yuying; Zhu, Xiaotong; Liu, Gang; Huang, Xingguo; Fang, Rejun; Li, Tiejun; Yin, Yulong

    2017-10-02

    This study aimed to investigate the long-term effects of protein restriction from piglets to finishing pigs for 16 weeks on meat quality, muscle amino acids, and amino acid transporters. 39 piglets were randomly divided into three groups: a control (20-18-16% crude protein, CP) and two protein restricted group (17-15-13% CP and 14-12-10% CP). The results showed that severe protein restriction (14-12-10% CP) inhibited feed intake and body weight, while moderate protein restriction (17-15-13% CP) had little effect on growth performance in pigs. Meat quality (i.e. pH, color traits, marbling, water-holding capacity, and shearing force) were tested and the results exhibited that 14-12-10% CP treatment markedly improved muscle marbling score and increased yellowness (b*). pH value (45min) was significant higher in 17-15-13% CP group than that in other groups. In addition, protein restriction reduced muscle histone, arginine, valine, and isoleucine abundances and enhanced glycine andlysine concentrations compared with the control group, while the RT-PCR results showed that protein restriction downregulated amino acids transporters. Mechanistic target of rapamycin (mTOR) signaling pathway was inactivated in moderate protein restricted group (17-15-13% CP), while severe protein restriction with dietary 14-12-10% CP markedly enhanced mTOR phosphorylation. In conclusion, long-term protein restriction affected meat quality and muscle amino acid metabolism in pigs, which might be associated with mTOR signaling pathway.

  9. Effect of increased protein intake on renal acid load and renal hemodynamic responses

    NARCIS (Netherlands)

    Teunissen-Beekman, Karianna F.M.; Dopheide, Janneke; Geleijnse, Marianne; Bakker, Stephan J.L.; Brink, Elizabeth J.; Leeuw, de Peter W.; Baak, van Marleen A.

    2016-01-01

    Increased protein intake versus maltodextrin intake for 4 weeks lowers blood pressure. Concerns exist that high-protein diets reduce renal function. Effects of acute and 4-week protein intake versus maltodextrin intake on renal acid load, glomerular filtration rate and related parameters were com

  10. Effect of increased protein intake on renal acid load and renal hemodynamic responses

    NARCIS (Netherlands)

    Teunissen-Beekman, Karianna F M; Dopheide, Janneke; Geleijnse, Johanna M; Bakker, Stephan J L; Brink, Elizabeth J; de Leeuw, Peter W; van Baak, Marleen A

    2016-01-01

    Increased protein intake versus maltodextrin intake for 4 weeks lowers blood pressure. Concerns exist that high-protein diets reduce renal function. Effects of acute and 4-week protein intake versus maltodextrin intake on renal acid load, glomerular filtration rate and related parameters were compar

  11. Understanding the synergistic effect of arginine and glutamic acid mixtures on protein solubility.

    Science.gov (United States)

    Shukla, Diwakar; Trout, Bernhardt L

    2011-10-20

    Understanding protein solubility is a key part of physical chemistry. In particular, solution conditions can have a major effect, and the effect of multiple cosolutes is little understood. It has been shown that the simultaneous addition of L-arginine hydrochloride and L-glutamic acid enhances the maximum achievable solubility of several poorly soluble proteins up to 4-8 times (Golovanov et. al, J. Am. Chem. Soc., 2004, 126, 8933-8939) and reduces the intermolecular interactions between proteins. The observed solubility enhancement is negligible for arginine and glutamic acid solutions as compared to the equimolar mixtures. In this study, we have established the molecular mechanism behind this observed synergistic effect of arginine and glutamic acid mixtures using preferential interaction theory and molecular dynamics simulations of Drosophilia Su(dx) protein (ww34). It was found that the protein solubility enhancement is related to the relative increase in the number of arginine and glutamic acid molecules around the protein in the equimolar mixtures due to additional hydrogen bonding interactions between the excipients on the surface of the protein when both excipients are present. The presence of these additional molecules around the protein leads to enhanced crowding, which suppresses the protein association. These results highlight the role of additive-additive interaction in tuning the protein-protein interactions. Furthermore, this study reports a unique behavior of additive solutions, where the presence of one additive in solution affects the concentration of another on the protein surface.

  12. Analysis of the ligand binding properties of recombinant bovine liver-type fatty acid binding protein

    DEFF Research Database (Denmark)

    Rolf, B; Oudenampsen-Krüger, E; Börchers, T

    1995-01-01

    The coding part of the cDNA for bovine liver-type fatty acid binding protein (L-FABP) has been amplified by RT-PCR, cloned and used for the construction of an Escherichia coli (E. coli) expression system. The recombinant protein made up to 25% of the soluble E. coli proteins and could be isolated...

  13. Fatty Acid-Binding Protein in Small Intestine IDENTIFICATION, ISOLATION, AND EVIDENCE FOR ITS ROLE IN CELLULAR FATTY ACID TRANSPORT

    Science.gov (United States)

    Ockner, Robert K.; Manning, Joan A.

    1974-01-01

    A soluble fatty acid-binding protein (FABP), mol wt ∼ 12,000 is present in intestinal mucosa and other tissues that utilize fatty acids, including liver, myocardium, adipose, and kidney. This protein binds long chain fatty acids both in vivo and in vitro. FABP was isolated from rat intestine by gel filtration and isoelectric focusing. It showed a reaction of complete immunochemical identity with proteins in the 12,000 mol wt fatty acid-binding fractions of liver, myocardium, and adipose tissue supernates. (The presence of immunochemically nonidentical 12,000 mol wt FABP in these tissues is not excluded.) By quantitative radial immunodiffusion, supernatant FABP concentration in mucosa from proximal and middle thirds of jejuno-ileum significantly exceeded that in distal third, duodenum, and liver, expressed as micrograms per milligram soluble protein, micrograms per gram DNA, and micrograms per gram tissue. FABP concentration in villi was approximately three times greater than in crypts. Small quantities of FABP were present in washed nuclei-cell membrane, mitochondrial and microsomal fractions. However, the amount of FABP solubilized per milligram membrane protein was similar for all particulate fractions, and total membrane-associated FABP was only about 16% of supernatant FABP. Intestinal FABP concentration was significantly greater in animals maintained on high fat diets than on low fat; saturated and unsaturated fat diets did not differ greatly in this regard. The preponderance of FABP in villi from proximal and middle intestine, its ability to bind fatty acids in vivo as well as in vitro, and its response to changes in dietary fat intake support the concept that this protein participates in cellular fatty acid transport during fat absorption. Identical or closely related 12,000 mol wt proteins may serve similar functions in other tissues. Images PMID:4211161

  14. Influence of protein source on amino acid uptake patterns and protein utilization in rainbow trout Oncorhynchus mykiss

    DEFF Research Database (Denmark)

    Rolland, Marine; Holm, Jørgen; Dalsgaard, Anne Johanne Tang

    acids to the fishmeal diet level (see Table 1). Amino acid uptake patterns were assessed by the appearance of amino acids in the blood stream following the ingestion of a meal, while dietary protein utilization was evaluated by examining the metabolic response to digestion and ammonium and urea...... described time sampling. Fish were killed by a blow in the head and blood was collected from the caudal vein with heparinized syringes. Plasma and red blood cells content were stored separately at -80 for amino acid content analysis. The ammonium excretion profiles (Figure 1) will be correlated...... with the amino acid profile in the blood and oxygen consumption during digestion to investigate the effect on protein utilization for each treatment...

  15. Detection of D-amino acids in purified proteins synthesized in Escherichia coli.

    Science.gov (United States)

    Miyamoto, Tetsuya; Sekine, Masae; Ogawa, Tetsuhiro; Hidaka, Makoto; Homma, Hiroshi; Masaki, Haruhiko

    2010-05-01

    It has long been believed that amino acids comprising proteins of all living organisms are only of the L-configuration, except for Gly. However, peptidyl D-amino acids were observed in hydrolysates of soluble high molecular weight fractions extracted from cells or tissues of various organisms. This strongly suggests that significant amounts of D-amino acids are naturally present in usual proteins. Thus we analyzed the D-amino acid contents of His-tag-purified beta-galactosidase and human urocortin, which were synthesized by Escherichia coli grown in controlled synthetic media. After acidic hydrolysis for various times at 110 degrees C, samples were derivatized with 4-fluoro-7-nitro-2, 1, 3-benzoxadiazole (NBD-F) and separated on a reverse-phase column followed by a chiral column into D- and L-enantiomers. The contents of D-enantiomers of Ala, Leu, Phe, Val, Asp, and Glu were determined by plotting index D/(D + L) against the incubation time for hydrolysis and extrapolating the linear regression line to 0 h to eliminate the effect of racemization of amino acids during the incubation. Significant contents of D-amino acids were reproducibly detected, the D-amino acid profile being specific to an individual protein. This finding indicated the likelihood that D-amino acids are in fact present in the purified proteins. On the other hand, the D-amino acid contents of proteins were hardly influenced by the addition of D- or L-amino acids to the cultivation medium, whereas intracellular free D-amino acids sensitively varied according to the extracellular conditions. The origin of these D-amino acids detected in proteins was discussed.

  16. Amino acid composition, molecular weight distribution and gel electrophoresis of walnut (Juglans regia L.) proteins and protein fractionations.

    Science.gov (United States)

    Mao, Xiaoying; Hua, Yufei; Chen, Guogang

    2014-01-01

    As a by-product of oil production, walnut proteins are considered as an additional source of plant protein for human food. To make full use of the protein resource, a comprehensive understanding of composition and characteristics of walnut proteins are required. Walnut proteins have been fractionated and characterized in this study. Amino acid composition, molecular weight distribution and gel electrophoresis of walnut proteins and protein fractionations were analyzed. The proteins were sequentially separated into four fractions according to their solubility. Glutelin was the main component of the protein extract. The content of glutelin, albumin, globulin and prolamin was about 72.06%, 7.54%, 15.67% and 4.73% respectively. Glutelin, albumin and globulin have a balanced content of essential amino acids, except for methionine, with respect to the FAO pattern recommended for adults. SDS-PAGE patterns of albumin, globulin and glutelin showed several polypeptides with molecular weights 14.4 to 66.2 kDa. The pattern of walnut proteins in two-dimension electrophoresis (2-DE) showed that the isoelectric point was mainly in the range of 4.8-6.8. The results of size exclusion chromatogram indicated molecular weight of the major components of walnut proteins were between 3.54 and 81.76 kDa.

  17. Amino Acid Composition, Molecular Weight Distribution and Gel Electrophoresis of Walnut (Juglans regia L. Proteins and Protein Fractionations

    Directory of Open Access Journals (Sweden)

    Xiaoying Mao

    2014-01-01

    Full Text Available As a by-product of oil production, walnut proteins are considered as an additional source of plant protein for human food. To make full use of the protein resource, a comprehensive understanding of composition and characteristics of walnut proteins are required. Walnut proteins have been fractionated and characterized in this study. Amino acid composition, molecular weight distribution and gel electrophoresis of walnut proteins and protein fractionations were analyzed. The proteins were sequentially separated into four fractions according to their solubility. Glutelin was the main component of the protein extract. The content of glutelin, albumin, globulin and prolamin was about 72.06%, 7.54%, 15.67% and 4.73% respectively. Glutelin, albumin and globulin have a balanced content of essential amino acids, except for methionine, with respect to the FAO pattern recommended for adults. SDS-PAGE patterns of albumin, globulin and glutelin showed several polypeptides with molecular weights 14.4 to 66.2 kDa. The pattern of walnut proteins in two-dimension electrophoresis (2-DE showed that the isoelectric point was mainly in the range of 4.8–6.8. The results of size exclusion chromatogram indicated molecular weight of the major components of walnut proteins were between 3.54 and 81.76 kDa.

  18. Bile Acids Trigger GLP-1 Release Predominantly by Accessing Basolaterally Located G Protein-Coupled Bile Acid Receptors

    DEFF Research Database (Denmark)

    Brighton, Cheryl A.; Rievaj, Juraj; Kuhre, Rune E.;

    2015-01-01

    Bile acids are well-recognized stimuli of glucagon-like peptide-1 (GLP-1) secretion. This action has been attributed to activation of the G protein-coupled bile acid receptor GPBAR1 (TGR5), although other potential bile acid sensors include the nuclear farnesoid receptor and the apical sodium......-coupled bile acid transporter ASBT. The aim of this study was to identify pathways important for GLP-1 release and to determine whether bile acids target their receptors on GLP-1-secreting L-cells from the apical or basolateral compartment. Using transgenic mice expressing fluorescent sensors specifically in L...... to either TLCA or TDCA. We conclude that the action of bile acids on GLP-1 secretion is predominantly mediated by GPBAR1 located on the basolateral L-cell membrane, suggesting that stimulation of gut hormone secretion may include postabsorptive mechanisms....

  19. Neuromuscular electrical stimulation prior to presleep protein feeding stimulates the use of protein-derived amino acids for overnight muscle protein synthesis.

    Science.gov (United States)

    Dirks, Marlou L; Groen, Bart B L; Franssen, Rinske; van Kranenburg, Janneau; van Loon, Luc J C

    2017-01-01

    Short periods of muscle disuse result in substantial skeletal muscle atrophy. Recently, we showed that both neuromuscular electrical stimulation (NMES) as well as presleep dietary protein ingestion represent effective strategies to stimulate muscle protein synthesis rates. In this study, we test our hypothesis that NMES can augment the use of presleep protein-derived amino acids for overnight muscle protein synthesis in older men. Twenty healthy, older [69 ± 1 (SE) yr] men were subjected to 24 h of bed rest, starting at 8:00 AM. In the evening, volunteers were subjected to 70-min 1-legged NMES, while the other leg served as nonstimulated control (CON). Immediately following NMES, 40 g of intrinsically l-[1-(13)C]-phenylalanine labeled protein was ingested prior to sleep. Blood samples were taken throughout the night, and muscle biopsies were obtained from both legs in the evening and the following morning (8 h after protein ingestion) to assess dietary protein-derived l-[1-(13)C]-phenylalanine enrichments in myofibrillar protein. Plasma phenylalanine concentrations and plasma l-[1-(13)C]-phenylalanine enrichments increased significantly following protein ingestion and remained elevated for up to 6 h after protein ingestion (P stimulated compared with the control leg (0.0344 ± 0.0019 vs. 0.0297 ± 0.0016 MPE, respectively; P stimulates the use of dietary protein-derived amino acids for overnight muscle protein synthesis in older men. Neuromuscular electrical stimulation (NMES) as well as presleep dietary protein ingestion represent effective strategies to stimulate muscle protein synthesis rates. Here we demonstrate that in older men after a day of bed rest, the application of NMES prior to presleep protein feeding stimulates the use of dietary protein-derived amino acids for overnight muscle protein synthesis by 18% compared with presleep protein feeding only. Copyright © 2017 the American Physiological Society.

  20. Refsum disease diagnostic marker phytanic acid alters the physical state of membrane proteins of liver mitochondria.

    Science.gov (United States)

    Schönfeld, P; Struy, H

    1999-08-27

    Phytanic acid (3,7,11,15-tetramethylhexadecanoic acid), a branched chain fatty acid accumulating in Refsum disease to high levels throughout the body, induces uncoupling of rat liver mitochondria similar to non-branched fatty acids (e.g. palmitic acid), but the contribution of the ADP/ATP carrier or the aspartate/glutamate carrier in phytanic acid-induced uncoupling is of minor importance. Possible deleterious effects of phytanic acid on membrane-linked energy coupling processes were studied by ESR spectroscopy using rat liver mitochondria and a membrane preparation labeled with the lipid-specific spin probe 5-doxylstearic acid (5-DSA) or the protein-specific spin probe MAL-TEMPO (4-maleimido-2,2,6, 6-tetramethyl-piperidine-1-oxyl). The effects of phytanic acid on phospholipid molecular dynamics and on the physical state of membrane proteins were quantified by estimation of the order parameter or the ratio of the amplitudes of the weakly to strongly immobilized MAL-TEMPO binding sites (W/S ratio), respectively. It was found, that phytanic acid (1) increased the mobility of phospholipid molecules (indicated by a decrease in the order parameter) and (2) altered the conformational state and/or the segmental mobility of membrane proteins (indicated by a drastic decrease in the W/S ratio). Unsaturated fatty acids with multiple cis-double bonds (e.g. linolenic or arachidonic acid), but not non-branched FFA (ranging from chain length C10:0 to C18:0), also decrease the W/S ratio. It is hypothesized that the interaction of phytanic acid with transmembrane proteins might stimulate the proton permeability through the mitochondrial inner membrane according to a mechanism, different to a protein-supported fatty acid cycling.

  1. Effects of N-methyl-D-aspartate Receptor Antagonist MK-801 on Expression of Glial Fibrillary Acidic Protein in the Subventricular Zone of Neonatal Rat%NMDA受体拮抗剂MK-801对新生大鼠室管膜下区胶质纤维酸性蛋白表达的影响

    Institute of Scientific and Technical Information of China (English)

    李晓泉; 陈芳芳; 王伟; 来青伟; 耿德勤; 樊红彬

    2013-01-01

    Aim To investigate the effects of the N-methyl-D-aspartate(NMDA)-receptor antagonist MK801 on the expression of glial fibrillary acidic protein(GFAP) in the subventricular zone(SVZ) of neonatal rat.Methods 40 neonatal SD rats were assigned into two groups,a MK-801 group and a control group.Each group was respectively divided into four time points,including P7 d (postnatal 7 days),P14 d,P21 d and P28 d.Rats in the experiment group were intraperitoneally injected selective non-competitive NMDA receptor antagonist MK-801 (10 mg·kg1),while rats in the control group were intraperitoneally injected saline.Immunohistochemical staining was used to assay the number of GFAP-positive cells in the SVZ.Results ① In the control group,the number of GFAP-positive cells increased at P14 d,and the number of GFAP-positive cells reached maximum at P21 d,positive cells decreased significantly at 28 d.② Compared with the control group,the number of GFAP-positive cells reduced significantly at P 14 d (65.40 ± 6.11) and P21 d (239.60 ± 12.92),and the number of GFAP-positive cells increased significantly [P14 d(79.20 ± 5.26),P21 d(265.20 ± 7.40)](P<0.01),while the number of positive cells without obvious changes at P7 d and P28 d in the MK-801 group.Conclusion NMDA-receptor antagonist MK-801 could reduce the expression of GFAP in the SVZ,and inhibit the neural stem cells proliferation and differentiation.%目的 研究N-甲基-D-天冬氨酸(NMDA)受体拮抗剂MK-801对新生7d大鼠室管膜下区(SVZ)胶质纤维酸性蛋白(GFAP)表达的影响.方法 将40只新生SD大鼠分成对照组和MK-801组,各组按出生后口(P)时间点再随机分成4个亚组:P7d、P14d、P21 d、P28d组.新生大鼠均于生后第3天给药,MK-801组腹腔注射MK-801 10 mg·kg-1;对照组腹腔注射同量生理盐水.通过免疫组化学方法观察大鼠SVZ区GFAP阳性细胞数.结果 ①对照组GFAP阳性细胞数于P14 d开始增加,至P21 d达最大值;但P28 d时阳

  2. Effect of liver fatty acid binding protein on fatty acid movement between liposomes and rat liver microsomes.

    Science.gov (United States)

    McCormack, M; Brecher, P

    1987-01-01

    Although movement of fatty acids between bilayers can occur spontaneously, it has been postulated that intracellular movement is facilitated by a class of proteins named fatty acid binding proteins (FABP). In this study we have incorporated long chain fatty acids into multilamellar liposomes made of phosphatidylcholine, incubated them with rat liver microsomes containing an active acyl-CoA synthetase, and measured formation of acyl-CoA in the absence or presence of FABP purified from rat liver. FABP increased about 2-fold the accumulation of acyl-CoA when liposomes were the fatty acid donor. Using fatty acid incorporated into liposomes made either of egg yolk lecithin or of dipalmitoylphosphatidylcholine, it was found that the temperature dependence of acyl-CoA accumulation in the presence of FABP correlated with both the physical state of phospholipid molecules in the liposomes and the binding of fatty acid to FABP, suggesting that fatty acid must first desorb from the liposomes before FABP can have an effect. An FABP-fatty acid complex incubated with microsomes, in the absence of liposomes, resulted in greater acyl-CoA formation than when liposomes were present, suggesting that desorption of fatty acid from the membrane is rate-limiting in the accumulation of acyl-CoA by this system. Finally, an equilibrium dialysis cell separating liposomes from microsomes on opposite sides of a Nuclepore filter was used to show that liver FABP was required for the movement and activation of fatty acid between the compartments. These studies show that liver FABP interacts with fatty acid that desorbs from phospholipid bilayers, and promotes movement to a membrane-bound enzyme, suggesting that FABP may act intracellularly by increasing net desorption of fatty acid from cell membranes. PMID:3446187

  3. Acetic Acid Causes Endoplasmic Reticulum Stress and Induces the Unfolded Protein Response in Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Nozomi Kawazoe

    2017-06-01

    Full Text Available Since acetic acid inhibits the growth and fermentation ability of Saccharomyces cerevisiae, it is one of the practical hindrances to the efficient production of bioethanol from a lignocellulosic biomass. Although extensive information is available on yeast response to acetic acid stress, the involvement of endoplasmic reticulum (ER and unfolded protein response (UPR has not been addressed. We herein demonstrated that acetic acid causes ER stress and induces the UPR. The accumulation of misfolded proteins in the ER and activation of Ire1p and Hac1p, an ER-stress sensor and ER stress-responsive transcription factor, respectively, were induced by a treatment with acetic acid stress (>0.2% v/v. Other monocarboxylic acids such as propionic acid and sorbic acid, but not lactic acid, also induced the UPR. Additionally, ire1Δ and hac1Δ cells were more sensitive to acetic acid than wild-type cells, indicating that activation of the Ire1p-Hac1p pathway is required for maximum tolerance to acetic acid. Furthermore, the combination of mild acetic acid stress (0.1% acetic acid and mild ethanol stress (5% ethanol induced the UPR, whereas neither mild ethanol stress nor mild acetic acid stress individually activated Ire1p, suggesting that ER stress is easily induced in yeast cells during the fermentation process of lignocellulosic hydrolysates. It was possible to avoid the induction of ER stress caused by acetic acid and the combined stress by adjusting extracellular pH.

  4. Acetic Acid Causes Endoplasmic Reticulum Stress and Induces the Unfolded Protein Response in Saccharomyces cerevisiae.

    Science.gov (United States)

    Kawazoe, Nozomi; Kimata, Yukio; Izawa, Shingo

    2017-01-01

    Since acetic acid inhibits the growth and fermentation ability of Saccharomyces cerevisiae, it is one of the practical hindrances to the efficient production of bioethanol from a lignocellulosic biomass. Although extensive information is available on yeast response to acetic acid stress, the involvement of endoplasmic reticulum (ER) and unfolded protein response (UPR) has not been addressed. We herein demonstrated that acetic acid causes ER stress and induces the UPR. The accumulation of misfolded proteins in the ER and activation of Ire1p and Hac1p, an ER-stress sensor and ER stress-responsive transcription factor, respectively, were induced by a treatment with acetic acid stress (>0.2% v/v). Other monocarboxylic acids such as propionic acid and sorbic acid, but not lactic acid, also induced the UPR. Additionally, ire1Δ and hac1Δ cells were more sensitive to acetic acid than wild-type cells, indicating that activation of the Ire1p-Hac1p pathway is required for maximum tolerance to acetic acid. Furthermore, the combination of mild acetic acid stress (0.1% acetic acid) and mild ethanol stress (5% ethanol) induced the UPR, whereas neither mild ethanol stress nor mild acetic acid stress individually activated Ire1p, suggesting that ER stress is easily induced in yeast cells during the fermentation process of lignocellulosic hydrolysates. It was possible to avoid the induction of ER stress caused by acetic acid and the combined stress by adjusting extracellular pH.

  5. Predicting disordered regions in proteins using the profiles of amino acid indices

    Science.gov (United States)

    Han, Pengfei; Zhang, Xiuzhen; Feng, Zhi-Ping

    2009-01-01

    Background Intrinsically unstructured or disordered proteins are common and functionally important. Prediction of disordered regions in proteins can provide useful information for understanding protein function and for high-throughput determination of protein structures. Results In this paper, algorithms are presented to predict long and short disordered regions in proteins, namely the long disordered region prediction algorithm DRaai-L and the short disordered region prediction algorithm DRaai-S. These algorithms are developed based on the Random Forest machine learning model and the profiles of amino acid indices representing various physiochemical and biochemical properties of the 20 amino acids. Conclusion Experiments on DisProt3.6 and CASP7 demonstrate that some sets of the amino acid indices have strong association with the ordered and disordered status of residues. Our algorithms based on the profiles of these amino acid indices as input features to predict disordered regions in proteins outperform that based on amino acid composition and reduced amino acid composition, and also outperform many existing algorithms. Our studies suggest that the profiles of amino acid indices combined with the Random Forest learning model is an important complementary method for pinpointing disordered regions in proteins. PMID:19208144

  6. Acute supplementation of amino acids increases net protein accretion in IUGR fetal sheep.

    Science.gov (United States)

    Brown, Laura D; Rozance, Paul J; Thorn, Stephanie R; Friedman, Jacob E; Hay, William W

    2012-08-01

    Placental insufficiency decreases fetal amino acid uptake from the placenta, plasma insulin concentrations, and protein accretion, thus compromising normal fetal growth trajectory. We tested whether acute supplementation of amino acids or insulin into the fetus with intrauterine growth restriction (IUGR) would increase net fetal protein accretion rates. Late-gestation IUGR and control (CON) fetal sheep received acute, 3-h infusions of amino acids (with euinsulinemia), insulin (with euglycemia and euaminoacidemia), or saline. Fetal leucine metabolism was measured under steady-state conditions followed by a fetal muscle biopsy to quantify insulin signaling. In CON, increasing amino acid delivery rates to the fetus by 100% increased leucine oxidation rates by 100%. In IUGR, amino acid infusion completely suppressed fetal protein breakdown rates but increased leucine oxidation rate by only 25%, resulting in increased protein accretion rates by 150%. Acute insulin infusion, however, had very little effect on amino acid delivery rates, fetal leucine disposal rates, or fetal protein accretion rates in CON or IUGR fetuses despite robust signaling of the fetal skeletal muscle insulin-signaling cascade. These results indicate that, when amino acids are given directly into the fetal circulation independently of changes in insulin concentrations, IUGR fetal sheep have suppressed protein breakdown rates, thus increasing net fetal protein accretion.

  7. Crystal Structure of Okadaic Acid Binding Protein 2.1: A Sponge Protein Implicated in Cytotoxin Accumulation.

    Science.gov (United States)

    Ehara, Haruhiko; Makino, Marie; Kodama, Koichiro; Konoki, Keiichi; Ito, Takuhiro; Sekine, Shun-ichi; Fukuzawa, Seketsu; Yokoyama, Shigeyuki; Tachibana, Kazuo

    2015-07-06

    Okadaic acid (OA) is a marine polyether cytotoxin that was first isolated from the marine sponge Halichondria okadai. OA is a potent inhibitor of protein serine/threonine phosphatases (PP) 1 and 2A, and the structural basis of phosphatase inhibition has been well investigated. However, the role and mechanism of OA retention in the marine sponge have remained elusive. We have solved the crystal structure of okadaic acid binding protein 2.1 (OABP2.1) isolated from H. okadai; it has strong affinity for OA and limited sequence homology to other proteins. The structure revealed that OABP2.1 consists of two α-helical domains, with the OA molecule deeply buried inside the protein. In addition, the global fold of OABP2.1 was unexpectedly similar to that of aequorin, a jellyfish photoprotein. The presence of structural homologues suggested that, by using similar protein scaffolds, marine invertebrates have developed diverse survival systems adapted to their living environments.

  8. Tsetse salivary gland proteins 1 and 2 are high affinity nucleic acid binding proteins with residual nuclease activity.

    Directory of Open Access Journals (Sweden)

    Guy Caljon

    Full Text Available Analysis of the tsetse fly salivary gland EST database revealed the presence of a highly enriched cluster of putative endonuclease genes, including tsal1 and tsal2. Tsal proteins are the major components of tsetse fly (G. morsitans morsitans saliva where they are present as monomers as well as high molecular weight complexes with other saliva proteins. We demonstrate that the recombinant tsetse salivary gland proteins 1&2 (Tsal1&2 display DNA/RNA non-specific, high affinity nucleic acid binding with K(D values in the low nanomolar range and a non-exclusive preference for duplex. These Tsal proteins exert only a residual nuclease activity with a preference for dsDNA in a broad pH range. Knockdown of Tsal expression by in vivo RNA interference in the tsetse fly revealed a partially impaired blood digestion phenotype as evidenced by higher gut nucleic acid, hematin and protein contents.

  9. Enhanced Bio-hydrogen Production from Protein Wastewater by Altering Protein Structure and Amino Acids Acidification Type

    Science.gov (United States)

    Xiao, Naidong; Chen, Yinguang; Chen, Aihui; Feng, Leiyu

    2014-01-01

    Enhanced bio-hydrogen production from protein wastewater by altering protein structure and amino acids acidification type via pH control was investigated. The hydrogen production reached 205.2 mL/g-protein when protein wastewater was pretreated at pH 12 and then fermented at pH 10. The mechanism studies showed that pH 12 pretreatment significantly enhanced protein bio-hydrolysis during the subsequent fermentation stage as it caused the unfolding of protein, damaged the protein hydrogen bonding networks, and destroyed the disulfide bridges, which increased the susceptibility of protein to protease. Moreover, pH 10 fermentation produced more acetic but less propionic acid during the anaerobic fermentation of amino acids, which was consistent with the theory of fermentation type affecting hydrogen production. Further analyses of the critical enzymes, genes, and microorganisms indicated that the activity and abundance of hydrogen producing bacteria in the pH 10 fermentation reactor were greater than those in the control. PMID:24495932

  10. Enhanced Bio-hydrogen Production from Protein Wastewater by Altering Protein Structure and Amino Acids Acidification Type

    Science.gov (United States)

    Xiao, Naidong; Chen, Yinguang; Chen, Aihui; Feng, Leiyu

    2014-02-01

    Enhanced bio-hydrogen production from protein wastewater by altering protein structure and amino acids acidification type via pH control was investigated. The hydrogen production reached 205.2 mL/g-protein when protein wastewater was pretreated at pH 12 and then fermented at pH 10. The mechanism studies showed that pH 12 pretreatment significantly enhanced protein bio-hydrolysis during the subsequent fermentation stage as it caused the unfolding of protein, damaged the protein hydrogen bonding networks, and destroyed the disulfide bridges, which increased the susceptibility of protein to protease. Moreover, pH 10 fermentation produced more acetic but less propionic acid during the anaerobic fermentation of amino acids, which was consistent with the theory of fermentation type affecting hydrogen production. Further analyses of the critical enzymes, genes, and microorganisms indicated that the activity and abundance of hydrogen producing bacteria in the pH 10 fermentation reactor were greater than those in the control.

  11. How the folding rates of two- and multistate proteins depend on the amino acid properties.

    Science.gov (United States)

    Huang, Jitao T; Huang, Wei; Huang, Shanran R; Li, Xin

    2014-10-01

    Proteins fold by either two-state or multistate kinetic mechanism. We observe that amino acids play different roles in different mechanism. Many residues that are easy to form regular secondary structures (α helices, β sheets and turns) can promote the two-state folding reactions of small proteins. Most of hydrophilic residues can speed up the multistate folding reactions of large proteins. Folding rates of large proteins are equally responsive to the flexibility of partial amino acids. Other properties of amino acids (including volume, polarity, accessible surface, exposure degree, isoelectric point, and phase transfer energy) have contributed little to folding kinetics of the proteins. Cysteine is a special residue, it triggers two-state folding reaction and but inhibits multistate folding reaction. These findings not only provide a new insight into protein structure prediction, but also could be used to direct the point mutations that can change folding rate.

  12. Detection and quantification of protein adduction by electrophilic fatty acids: mitochondrial generation of fatty acid nitroalkene derivatives.

    Science.gov (United States)

    Schopfer, F J; Batthyany, C; Baker, P R S; Bonacci, G; Cole, M P; Rudolph, V; Groeger, A L; Rudolph, T K; Nadtochiy, S; Brookes, P S; Freeman, B A

    2009-05-01

    Nitroalkene fatty acid derivatives manifest a strong electrophilic nature, are clinically detectable, and induce multiple transcriptionally regulated anti-inflammatory responses. At present, the characterization and quantification of endogenous electrophilic lipids are compromised by their Michael addition with protein and small-molecule nucleophilic targets. Herein, we report a trans-nitroalkylation reaction of nitro-fatty acids with beta-mercaptoethanol (BME) and apply this reaction to the unbiased identification and quantification of reaction with nucleophilic targets. Trans-nitroalkylation yields are maximal at pH 7 to 8 and occur with physiological concentrations of target nucleophiles. This reaction is also amenable to sensitive mass spectrometry-based quantification of electrophilic fatty acid-protein adducts upon electrophoretic resolution of proteins. In-gel trans-nitroalkylation reactions also permit the identification of protein targets without the bias and lack of sensitivity of current proteomic approaches. Using this approach, it was observed that fatty acid nitroalkenes are rapidly metabolized in vivo by a nitroalkene reductase activity and mitochondrial beta-oxidation, yielding a variety of electrophilic and nonelectrophilic products that could be structurally characterized upon BME-based trans-nitroalkylation reaction. This strategy was applied to the detection and quantification of fatty acid nitration in mitochondria in response to oxidative inflammatory conditions induced by myocardial ischemia-reoxygenation.

  13. Metabolic Effects of Dietary Proteins, Amino Acids and The Other Amine Consisting Compounds on Cardiovascular System.

    OpenAIRE

    Elif Uğur; Reyhan Nergiz Ünal

    2017-01-01

    During the prevention and treatment of cardiovascular diseases, first cause of deaths in the world, diet has a vital role. While nutrition programs for the cardiovascular health generally focus on lipids and carbohydrates, effects of proteins are not well concerned. Thus this review is written in order to examine effect of proteins, amino acids, and the other amine consisting compounds on cardiovascular system. Because of that animal or plant derived proteins have different protein compositio...

  14. Compartmentation of hepatic fatty-acid-binding protein in liver cells and its effect on microsomal phosphatidic acid biosynthesis.

    Science.gov (United States)

    Bordewick, U; Heese, M; Börchers, T; Robenek, H; Spener, F

    1989-03-01

    Fatty-acid-binding proteins are known to occur in the cytosol of mammalian cells and to bind fatty acids and their CoA-esters. Application of the postembedding protein A-gold labeling method with antibody against the hepatic type fatty-acid-binding protein (hFABP) to cross-sections of liver cells and a newly developed gel-chromatographic immunofluorescence assay established qualitatively (1) that hFABP in mitochondria was confined to outer mitochondrial membranes, (2) the presence of this protein in microsomes and (3) that nuclei were also filled with hFABP. Quantitative data elaborated with a non-competitive ELISA confirmed these results. A significant difference to the distribution of cardiac FABP in heart muscle cells, where this type of protein was found in cytosol, matrix and nuclei, was observed (Börchers et al. (1989) Biochim. Biophys. Acta, in the press). hFABP-containing rat liver microsomes were incubated with long-chain acyl-CoAs in the presence of hFABP (isolated from rat liver cytosol) in a study on the acylation of sn-glycerol-3-phosphate and lysophosphatidic acid. Both acyltransferases were stimulated by addition of hFABP to the incubation medium. The morphological, immunochemical as well as kinetic data infer a direct interaction of hFABP with microsomal membranes in liver cells.

  15. Towards an understanding of Mesocestoides vogae fatty acid binding proteins' roles.

    Directory of Open Access Journals (Sweden)

    Gabriela Alvite

    Full Text Available Two fatty acid binding proteins, MvFABPa and MvFABPb were identified in the parasite Mesocestoides vogae (Platyhelmithes, Cestoda. Fatty acid binding proteins are small intracellular proteins whose members exhibit great diversity. Proteins of this family have been identified in many organisms, of which Platyhelminthes are among the most primitive. These proteins have particular relevance in flatworms since de novo synthesis of fatty acids is absent. Fatty acids should be captured from the media needing an efficient transport system to uptake and distribute these molecules. While HLBPs could be involved in the shuttle of fatty acids to the surrounding host tissues and convey them into the parasite, FABPs could be responsible for the intracellular trafficking. In an effort to understand the role of MvFABPs in fatty acid transport of M. vogae larvae, we analysed the intracellular localization of both MvFABPs and the co-localization with in vivo uptake of fatty acid analogue BODIPY FL C16. Immunohistochemical studies on larvae sections using specific antibodies, showed a diffuse cytoplasmic distribution of each protein with some expression in nuclei and mitochondria. MvFABPs distribution was confirmed by mass spectrometry identification from 2D-electrophoresis of larvae subcellular fractions. This work is the first report showing intracellular distribution of MvFABPs as well as the co-localization of these proteins with the BODIPY FL C16 incorporated from the media. Our results suggest that fatty acid binding proteins could target fatty acids to cellular compartments including nuclei. In this sense, M. vogae FABPs could participate in several cellular processes fulfilling most of the functions attributed to vertebrate's counterparts.

  16. Metabolic Effects of Dietary Proteins, Amino Acids and The Other Amine Consisting Compounds on Cardiovascular System.

    Directory of Open Access Journals (Sweden)

    Elif Uğur

    2017-01-01

    Full Text Available During the prevention and treatment of cardiovascular diseases, first cause of deaths in the world, diet has a vital role. While nutrition programs for the cardiovascular health generally focus on lipids and carbohydrates, effects of proteins are not well concerned. Thus this review is written in order to examine effect of proteins, amino acids, and the other amine consisting compounds on cardiovascular system. Because of that animal or plant derived proteins have different protein composition in different foods such as dairy products, egg, meat, chicken, fish, pulse and grains, their effects on blood pressure and regulation of lipid profile are unlike. In parallel amino acids made up proteins have different effect on cardiovascular system. From this point, sulfur containing amino acids, branched chain amino acids, aromatic amino acids, arginine, ornithine, citrulline, glycine, and glutamine may affect cardiovascular system in different metabolic pathways. In this context, one carbon metabolism, synthesis of hormone, stimulation of signaling pathways and effects of intermediate and final products that formed as a result of amino acids metabolism is determined. Despite the protein and amino acids, some other amine consisting compounds in diet include trimethylamine N-oxide, heterocyclic aromatic amines, polycyclic aromatic hydrocarbons and products of Maillard reaction. These amine consisting compounds generally increase the risk for cardiovascular diseases by stimulating oxidative stress, inflammation, and formation of atherosclerotic plaque.

  17. Natural vs. random protein sequences: Discovering combinatorics properties on amino acid words.

    Science.gov (United States)

    Santoni, Daniele; Felici, Giovanni; Vergni, Davide

    2016-02-21

    Casual mutations and natural selection have driven the evolution of protein amino acid sequences that we observe at present in nature. The question about which is the dominant force of proteins evolution is still lacking of an unambiguous answer. Casual mutations tend to randomize protein sequences while, in order to have the correct functionality, one expects that selection mechanisms impose rigid constraints on amino acid sequences. Moreover, one also has to consider that the space of all possible amino acid sequences is so astonishingly large that it could be reasonable to have a well tuned amino acid sequence indistinguishable from a random one. In order to study the possibility to discriminate between random and natural amino acid sequences, we introduce different measures of association between pairs of amino acids in a sequence, and apply them to a dataset of 1047 natural protein sequences and 10,470 random sequences, carefully generated in order to preserve the relative length and amino acid distribution of the natural proteins. We analyze the multidimensional measures with machine learning techniques and show that, to a reasonable extent, natural protein sequences can be differentiated from random ones.

  18. One-Pot Procedure for Recovery of Gallic Acid from Wastewater and Encapsulation within Protein Particles.

    Science.gov (United States)

    Nourbakhsh, Himan; Madadlou, Ashkan; Emam-Djomeh, Zahra; Wang, Yi-Cheng; Gunasekaran, Sundaram; Mousavi, Mohammad E

    2016-02-24

    A whey protein isolate solution was heat-denatured and treated with the enzyme transglutaminase, which cross-linked ≈26% of the amino groups and increased the magnitude of the ζ-potential value. The protein solution was microemulsified, and then the resulting water-in-oil microemulsion was dispersed within a gallic acid-rich model wastewater. Gallic acid extraction by the outlined microemulsion liquid membrane (MLM) from the exterior aqueous phase (wastewater) and accumulation within the internal aqueous nanodroplets induced protein cold-set gelation and resulted in the formation of gallic acid-enveloping nanoparticles. Measurements with a strain-controlled rheometer indicated a progressive increase in the MLM viscosity during gallic acid recovery corresponding to particle formation. The mean hydrodynamic size of the nanoparticles made from the heat-denatured and preheated enzymatically cross-linked proteins was 137 and 122 nm, respectively. The enzymatic cross-linking of whey proteins led to a higher gallic acid recovery yield and increased the glass transition enthalpy and temperature. A similar impact on glass transition indices was observed by the gallic acid-induced nanoparticulation of proteins. Scanning electron microscopy showed the existence of numerous jammed/fused nanoparticles. It was suggested on the basis of the results of Fourier transform infrared spectroscopy that the in situ nanoparticulation of proteins shifted the C-N stretching and C-H bending peaks to higher wavenumbers. X-ray diffraction results proposed a decreased β-sheet content for proteins because of the acid-induced particulation. The nanoparticles made from the enzymatically cross-linked protein were more stable against the in vitro gastrointestinal digestion and retained almost 19% of the entrapped gallic acid after 300 min sequential gastric and intestinal digestions.

  19. Release of periplasmic proteins of Brucella suis upon acidic shock involves the outer membrane protein Omp25.

    Science.gov (United States)

    Boigegrain, Rose-Anne; Salhi, Imed; Alvarez-Martinez, Maria-Teresa; Machold, Jan; Fedon, Yann; Arpagaus, Martine; Weise, Christoph; Rittig, Michael; Rouot, Bruno

    2004-10-01

    The survival and replication of Brucella in macrophages is initially triggered by a low intraphagosomal pH. In order to identify proteins released by Brucella during this early acidification step, we analyzed Brucella suis conditioned medium at various pH levels. No significant proteins were released at pH 4.0 in minimal medium or citrate buffer, whereas in acetate buffer, B. suis released a substantial amount of soluble proteins. Comparison of 13 N-terminal amino acid sequences determined by Edman degradation with their corresponding genomic sequences revealed that all of these proteins possessed a signal peptide indicative of their periplasmic location. Ten proteins are putative substrate binding proteins, including a homologue of the nopaline binding protein of Agrobacterium tumefaciens. The absence of this homologue in Brucella melitensis was due to the deletion of a 7.7-kb DNA fragment in its genome. We also characterized for the first time a hypothetical 9.8-kDa basic protein composed of five amino acid repeats. In B. suis, this protein contained 9 repeats, while 12 were present in the B. melitensis orthologue. B. suis in acetate buffer depended on neither the virB type IV secretory system nor the omp31 gene product. However, the integrity of the omp25 gene was required for release at acidic pH, while the absence of omp25b or omp25c displayed smaller effects. Together, these results suggest that Omp25 is involved in the membrane permeability of Brucella in acidic medium.

  20. [Analysis of proteins, amino acids and inorganic elements in Holotrichia diomphalia from different areas].

    Science.gov (United States)

    Cao, Wei; Liu, Dan; Zhang, Yi-Kai; Wang, Xiao-Yu; Chang, Yan-Rong; Yang, Qian; Wang, Si-Wang

    2010-10-01

    To analyze the content of proteins,amino acids and inorganic elements of Holotrichia diomphalia in different growing areas as the references for quality evaluation and reasonable application of them. The contents of proteins were determined using semi-micro Kjeldahl method. The contents of seventeen amino acids and inorganic elements were determined with amino acid analyzer and atomic absorption spectrometer and elemental analyzer, respectively. The contents of protein were 33.4%-44.4%, and that in Jiangxi were the highest in five different areas. There were seventeen kinds of amino acids in Holotrichia diomphalia. Among them, seven amino acids were essential to human life. The content of glutamic acid was the highest in seventeen amino acids. In inorganic elements, the content of Mg, Ca was higher in macroelements and Fe, Zn was higher in microelements. There are many kinds of necessary amino acids and inorganic elements for man kind in Holotrichia diomphalia. The contents of proteins, amino acids and inorganic elements have some difference in Holotrichia diomphalia of different growing areas.

  1. Dietary protein, physiological condition and metabolic amino acid utilisation.

    NARCIS (Netherlands)

    Weijs, P.J.M.

    1993-01-01

    This thesis describes the investigated effects of the level of dietary protein intake and the physiological condition of the animal on the percental oxidation of leucine. This measure reflects which part of the free leucine pool was used for protein and energy metabolism. The employed technique cons

  2. Hypochlorite-induced oxidation of amino acids, peptides and proteins

    DEFF Research Database (Denmark)

    Hawkins, C L; Pattison, D I; Davies, Michael Jonathan

    2003-01-01

    Activated phagocytes generate the potent oxidant hypochlorite (HOCl) via the release of the enzyme myeloperoxidase and hydrogen peroxide. HOCl is known to react with a number of biological targets including proteins, DNA, lipids and cholesterol. Proteins are likely to be major targets for reaction...

  3. Dietary protein, physiological condition and metabolic amino acid utilisation

    NARCIS (Netherlands)

    Weijs, P.J.M.

    1993-01-01

    This thesis describes the investigated effects of the level of dietary protein intake and the physiological condition of the animal on the percental oxidation of leucine. This measure reflects which part of the free leucine pool was used for protein and energy metabolism. The employed

  4. The pathogen-related yeast protein Pry1, a member of the CAP protein superfamily, is a fatty acid-binding protein.

    Science.gov (United States)

    Darwiche, Rabih; Mène-Saffrané, Laurent; Gfeller, David; Asojo, Oluwatoyin A; Schneiter, Roger

    2017-05-19

    Members of the CAP superfamily (cysteine-rich secretory proteins, antigen 5, and pathogenesis-related 1 proteins), also known as SCP superfamily (sperm-coating proteins), have been implicated in many physiological processes, including immune defenses, venom toxicity, and sperm maturation. Their mode of action, however, remains poorly understood. Three proteins of the CAP superfamily, Pry1, -2, and -3 (pathogen related in yeast), are encoded in the Saccharomyces cerevisiae genome. We have shown previously that Pry1 binds cholesterol in vitro and that Pry function is required for sterol secretion in yeast cells, indicating that members of this superfamily may generally bind sterols or related small hydrophobic compounds. On the other hand, tablysin-15, a CAP protein from the horsefly Tabanus yao, has been shown to bind leukotrienes and free fatty acids in vitro Therefore, here we assessed whether the yeast Pry1 protein binds fatty acids. Computational modeling and site-directed mutagenesis indicated that the mode of fatty acid binding is conserved between tablysin-15 and Pry1. Pry1 bound fatty acids with micromolar affinity in vitro, and its function was essential for fatty acid export in cells lacking the acyl-CoA synthetases Faa1 and Faa4. Fatty acid binding of Pry1 is independent of its capacity to bind sterols, and the two sterol- and fatty acid-binding sites are nonoverlapping. These results indicate that some CAP family members, such as Pry1, can bind different lipids, particularly sterols and fatty acids, at distinct binding sites, suggesting that the CAP domain may serve as a stable, secreted protein domain that can accommodate multiple ligand-binding sites. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  5. Role of fatty acid binding proteins and long chain fatty acids in modulating nuclear receptors and gene transcription.

    Science.gov (United States)

    Schroeder, Friedhelm; Petrescu, Anca D; Huang, Huan; Atshaves, Barbara P; McIntosh, Avery L; Martin, Gregory G; Hostetler, Heather A; Vespa, Aude; Landrock, Danilo; Landrock, Kerstin K; Payne, H Ross; Kier, Ann B

    2008-01-01

    Abnormal energy regulation may significantly contribute to the pathogenesis of obesity, diabetes mellitus, cardiovascular disease, and cancer. For rapid control of energy homeostasis, allosteric and posttranslational events activate or alter activity of key metabolic enzymes. For longer impact, transcriptional regulation is more effective, especially in response to nutrients such as long chain fatty acids (LCFA). Recent advances provide insights into how poorly water-soluble lipid nutrients [LCFA; retinoic acid (RA)] and their metabolites (long chain fatty acyl Coenzyme A, LCFA-CoA) reach nuclei, bind their cognate ligand-activated receptors, and regulate transcription for signaling lipid and glucose catabolism or storage: (i) while serum and cytoplasmic LCFA levels are in the 200 mircroM-mM range, real-time imaging recently revealed that LCFA and LCFA-CoA are also located within nuclei (nM range); (ii) sensitive fluorescence binding assays show that LCFA-activated nuclear receptors [peroxisome proliferator-activated receptor-alpha (PPARalpha) and hepatocyte nuclear factor 4alpha (HNF4alpha)] exhibit high affinity (low nM KdS) for LCFA (PPARalpha) and/or LCFA-CoA (PPARalpha, HNF4alpha)-in the same range as nuclear levels of these ligands; (iii) live and fixed cell immunolabeling and imaging revealed that some cytoplasmic lipid binding proteins [liver fatty acid binding protein (L-FABP), acyl CoA binding protein (ACBP), cellular retinoic acid binding protein-2 (CRABP-2)] enter nuclei, bind nuclear receptors (PPARalpha, HNF4alpha, CRABP-2), and activate transcription of genes in fatty acid and glucose metabolism; and (iv) studies with gene ablated mice provided physiological relevance of LCFA and LCFA-CoA binding proteins in nuclear signaling. This led to the hypothesis that cytoplasmic lipid binding proteins transfer and channel lipidic ligands into nuclei for initiating nuclear receptor transcriptional activity to provide new lipid nutrient signaling pathways that

  6. AMINO-ACIDS FOR THE MEASUREMENT OF PROTEIN-SYNTHESIS INVIVO BY PET

    NARCIS (Netherlands)

    VAALBURG, W; COENEN, HH; CROUZEL, C; ELSINGA, PH; LANGSTROM, B; LEMAIRE, C; MEYER, GJ

    Aspects of the suitability of various labelled amino acids for measuring their local incorporation into proteins by positron emission tomography, especially radiochemical aspects including choice of radionuclide, position of label and ease of preparation, are discussed.

  7. Lauric acid dependent enhancement in hepatic SCPx protein requires an insulin deficient environment.

    Science.gov (United States)

    Lopez, Dayami; Niesen, Melissa; Bedi, Mohini; McLean, Mark P

    2008-02-01

    Sterol carrier protein X (SCPx) is a peroxisomal protein with both lipid transfer and thiolase activity. Treating with the fatty acid, lauric acid, induced SCPx mRNA levels in rat liver and in rat hepatoma H4IIE cells but enhanced protein levels of SCPx and the thiolase produced as a post-translational modification of SCPx were only seen in H4IIE cells. Further investigation revealed that the presence of insulin can mask lauric acid effects on the SCPx gene especially at the protein level. These data are in agreement with the findings that diabetes, a medical condition characterized by high levels of fatty acids in an insulin deficient environment, enhances the hepatic expression of SCPx.

  8. Effect of disintegration wave grinding on fractional protein and amino acid composition of chickpea

    Directory of Open Access Journals (Sweden)

    G. O. Magomedov

    2013-01-01

    Full Text Available The study of fractional changes and amino acid composition of proteins in the application of chickpea disintegration wave grinding. Comparative analysis of six varieties of chickpea before and after grinding.

  9. Study of stationary phase metabolism via isotopomer analysis of amino acids from an isolated protein.

    Science.gov (United States)

    Shaikh, Afshan S; Tang, Yinjie J; Mukhopadhyay, Aindrila; Martín, Héctor García; Gin, Jennifer; Benke, Peter I; Keasling, Jay D

    2010-01-01

    Microbial production of many commercially important secondary metabolites occurs during stationary phase, and methods to measure metabolic flux during this growth phase would be valuable. Metabolic flux analysis is often based on isotopomer information from proteinogenic amino acids. As such, flux analysis primarily reflects the metabolism pertinent to the growth phase during which most proteins are synthesized. To investigate central metabolism and amino acids synthesis activity during stationary phase, addition of fully (13)C-labeled glucose followed by induction of green fluorescent protein (GFP) expression during stationary phase was used. Our results indicate that Escherichia coli was able to produce new proteins (i.e., GFP) in the stationary phase, and the amino acids in GFP were mostly from degraded proteins synthesized during the exponential growth phase. Among amino acid biosynthetic pathways, only those for serine, alanine, glutamate/glutamine, and aspartate/asparagine had significant activity during the stationary phase.

  10. Study of Stationary Phase Metabolism Via Isotopomer Analysis of Amino Acids from an Isolated Protein

    Energy Technology Data Exchange (ETDEWEB)

    Shaikh, AfshanS.; Tang, YinjieJ.; Mukhopadhyay, Aindrila; Martin, Hector Garcia; Gin, Jennifer; Benke, Peter; Keasling, Jay D.

    2009-09-14

    Microbial production of many commercially important secondary metabolites occurs during stationary phase, and methods to measure metabolic flux during this growth phase would be valuable. Metabolic flux analysis is often based on isotopomer information from proteinogenic amino acids. As such, flux analysis primarily reflects the metabolism pertinent to the growth phase during which most proteins are synthesized. To investigate central metabolism and amino acids synthesis activity during stationary phase, addition of fully 13C-labeled glucose followed by induction of green fluorescent protein (GFP) expression during stationary phase was used. Our results indicate that Escherichia coli was able to produce new proteins (i.e., GFP) in the stationary phase, and the amino acids in GFP were mostly from degraded proteins synthesized during the exponential growth phase. Among amino acid biosynthetic pathways, only those for serine, alanine, glutamate/glutamine, and aspartate/asparagine had significant activity during the stationary phase.

  11. A general method for site-specific incorporation of unnatural amino acids into proteins

    Energy Technology Data Exchange (ETDEWEB)

    Noren, C.J.; Anthony-Cahill, S.J; Griffith, M.C.; Schultz, P.G. (Lawrence Berkeley Lab., CA (USA))

    1989-04-14

    A new method has been developed that makes it possible to site-specifically incorporate unnatural amino acids into proteins. Synthetic amino acids were incorporated into the enzyme {beta}-lactamase by the use of a chemically acylated suppressor transfer RNA that inserted the amino acid in response to a stop codon substituted for the codon encoding residue of interest. Peptide mapping localized the inserted amino acid to a single peptide, and enough enzyme could be generated for purification to homogeneity. The catalytic properties of several mutants at the conserved Phe{sup 66} were characterized. The ability to selectively replace amino acids in a protein with a wide variety of structural and electronic variants should provide a more detailed understanding of protein structure and function. 45 refs., 7 figs., 1 tab.

  12. Engineering acyl carrier protein to enhance production of shortened fatty acids.

    Science.gov (United States)

    Liu, Xueliang; Hicks, Wade M; Silver, Pamela A; Way, Jeffrey C

    2016-01-01

    The acyl carrier protein (ACP) is an essential and ubiquitous component of microbial synthesis of fatty acids, the natural precursor to biofuels. Natural fatty acids usually contain long chains of 16 or more carbon atoms. Shorter carbon chains, with increased fuel volatility, are desired for internal combustion engines. Engineering the length specificity of key proteins in fatty acid metabolism, such as ACP, may enable microbial synthesis of these shorter chain fatty acids. We constructed a homology model of the Synechococcus elongatus ACP, showing a hydrophobic pocket harboring the growing acyl chain. Amino acids within the pocket were mutated to increase steric hindrance to the acyl chain. Certain mutant ACPs, when over-expressed in Escherichia coli, increased the proportion of shorter chain lipids; I75 W and I75Y showed the strongest effects. Expression of I75 W and I75Y mutant ACPs also increased production of lauric acid in E. coli that expressed the C12-specific acyl-ACP thioesterase from Cuphea palustris. We engineered the specificity of the ACP, an essential protein of fatty acid metabolism, to alter the E. coli lipid pool and enhance production of medium-chain fatty acids as biofuel precursors. These results indicate that modification of ACP itself could be combined with enzymes affecting length specificity in fatty acid synthesis to enhance production of commodity chemicals based on fatty acids.

  13. Conservation of Shannon's redundancy for proteins. [information theory applied to amino acid sequences

    Science.gov (United States)

    Gatlin, L. L.

    1974-01-01

    Concepts of information theory are applied to examine various proteins in terms of their redundancy in natural originators such as animals and plants. The Monte Carlo method is used to derive information parameters for random protein sequences. Real protein sequence parameters are compared with the standard parameters of protein sequences having a specific length. The tendency of a chain to contain some amino acids more frequently than others and the tendency of a chain to contain certain amino acid pairs more frequently than other pairs are used as randomness measures of individual protein sequences. Non-periodic proteins are generally found to have random Shannon redundancies except in cases of constraints due to short chain length and genetic codes. Redundant characteristics of highly periodic proteins are discussed. A degree of periodicity parameter is derived.

  14. Cellular fatty acid composition, protein profile and antimicrobial activity of Bacillus sp., isolated from fish gut

    Directory of Open Access Journals (Sweden)

    Pushparaj Sujith

    2014-01-01

    Full Text Available Objective: To purify and partially characterize the antimicrobial compounds from bacteria Bacillus sp., isolated from fish gut. Methods: Protein and fatty acids were isolated from the bacteria and checked for the presence of antibacterial activity. Protein has been purified to apparent homogeneity from the supernatants of culture by means of ammonium sulphate precipitation followed by dialysis. Fourier transform infrared spectroscopy analyses were performed for proteins to identify the functional groups. Results: Protein showed an apparent molecular mass 56, 47 and 39 kDa on sodium dodecyl sulfate polyacrylamide gel electrophoresis. Fatty acids were extracted and subjected to gas chromatographic analysis. Conclusions: The antimicrobial activity of the bacteria might be due to the presence of fatty acids and proteins which holds promise for the development of new drugs.

  15. Differential Effects of High-Protein Diets Derived from Soy and Casein on Blood-Brain Barrier Integrity in Wild-type Mice.

    Science.gov (United States)

    Snelson, Matthew; Mamo, John C L; Lam, Virginie; Giles, Corey; Takechi, Ryusuke

    2017-01-01

    A number of studies report that a diet high in protein influences cognitive performance, but the results are inconsistent. Studies demonstrated that protein from different food sources has differential effects on cognition. It is increasingly recognized that the integrity of cerebrovascular blood-brain barrier (BBB) is pivotal for central nervous system function. However, to date, no studies have reported the effects of high-protein diets on BBB integrity. Therefore, in this study, the effects of diets enriched in casein or soy protein on BBB permeability were investigated. Immunomicroscopy analyses of cerebral parenchymal immunoglobulin G extravasation indicated significant BBB disruption in the cortex of young adult mice maintained on high-casein diet for 12 weeks, while no signs of BBB dysfunction were observed in mice fed with control or high-soy protein diet. Moreover, cortical expression of glial fibrillary acidic protein (GFAP) was significantly greater in mice fed the high-casein diet compared to control mice, indicating heightened astrocyte activation, whereas mice maintained on a soy-enriched diet showed no increase of GFAP abundance. Plasma concentrations of homocysteine were markedly greater in mice maintained on a high-casein diet in comparison to control mice. Collectively, these findings suggest that a diet enriched in casein but not soy protein may induce astrocyte activation through exaggerated BBB permeability by increased plasma homocysteine. The outcomes indicate the differential effects of protein sources on BBB and neuroinflammation, which may provide an important implication for dietary guidelines for protein supplementation.

  16. Bacterial periplasmic sialic acid-binding proteins exhibit a conserved binding site

    Energy Technology Data Exchange (ETDEWEB)

    Gangi Setty, Thanuja [Institute for Stem Cell Biology and Regenerative Medicine, NCBS Campus, GKVK Post, Bangalore, Karnataka 560 065 (India); Cho, Christine [Carver College of Medicine, University of Iowa, Iowa City, IA 52242-1109 (United States); Govindappa, Sowmya [Institute for Stem Cell Biology and Regenerative Medicine, NCBS Campus, GKVK Post, Bangalore, Karnataka 560 065 (India); Apicella, Michael A. [Carver College of Medicine, University of Iowa, Iowa City, IA 52242-1109 (United States); Ramaswamy, S., E-mail: ramas@instem.res.in [Institute for Stem Cell Biology and Regenerative Medicine, NCBS Campus, GKVK Post, Bangalore, Karnataka 560 065 (India)

    2014-07-01

    Structure–function studies of sialic acid-binding proteins from F. nucleatum, P. multocida, V. cholerae and H. influenzae reveal a conserved network of hydrogen bonds involved in conformational change on ligand binding. Sialic acids are a family of related nine-carbon sugar acids that play important roles in both eukaryotes and prokaryotes. These sialic acids are incorporated/decorated onto lipooligosaccharides as terminal sugars in multiple bacteria to evade the host immune system. Many pathogenic bacteria scavenge sialic acids from their host and use them for molecular mimicry. The first step of this process is the transport of sialic acid to the cytoplasm, which often takes place using a tripartite ATP-independent transport system consisting of a periplasmic binding protein and a membrane transporter. In this paper, the structural characterization of periplasmic binding proteins from the pathogenic bacteria Fusobacterium nucleatum, Pasteurella multocida and Vibrio cholerae and their thermodynamic characterization are reported. The binding affinities of several mutations in the Neu5Ac binding site of the Haemophilus influenzae protein are also reported. The structure and the thermodynamics of the binding of sugars suggest that all of these proteins have a very well conserved binding pocket and similar binding affinities. A significant conformational change occurs when these proteins bind the sugar. While the C1 carboxylate has been identified as the primary binding site, a second conserved hydrogen-bonding network is involved in the initiation and stabilization of the conformational states.

  17. Designing medical foods for inherited metabolic disorders: why intact protein is superior to amino acids.

    Science.gov (United States)

    Ney, Denise Marie; Etzel, Mark Raymond

    2017-04-01

    Phenylketonuria and tyrosinemia are inherited metabolic disorders characterized by high blood levels of phenylalanine (Phe) or tyrosine (Tyr), due to mutations in genes affecting Phe and Tyr metabolism, respectively. The primary management is a lifelong diet restricted in protein from natural foods in combination with medical foods comprised mixtures of synthetic amino acids. Compliance is often poor after childhood leading to neuropsychological sequela. Glycomacropeptide, an intact 64 amino acid glycophosphopeptide isolated from cheese whey, provides a new paradigm for the management of phenylketonuria and tyrosinemia because glycomacropeptide contains no Phe and Tyr in its pure form, and is also a prebiotic. Medical foods made from glycomacropeptide have been used successfully for the management of phenylketonuria and tyrosinemia. Preclinical and clinical studies demonstrate that intact protein from glycomacropeptide provides a more acceptable and physiologic source of defined protein compared to amino acids in medical foods. For example, harmful gut bacteria were reduced, beneficial short chain fatty acids increased, renal workload decreased, protein utilization increased, and bone fragility decreased using intact protein versus amino acids. Advances in biotechnology will propel the transition from synthetic amino acids to intact proteins for the management of inherited metabolic disorders. Copyright © 2016 Elsevier Ltd. All rights reserved.

  18. Evolution of the genetic code by incorporation of amino acids that improved or changed protein function.

    Science.gov (United States)

    Francis, Brian R

    2013-10-01

    Fifty years have passed since the genetic code was deciphered, but how the genetic code came into being has not been satisfactorily addressed. It is now widely accepted that the earliest genetic code did not encode all 20 amino acids found in the universal genetic code as some amino acids have complex biosynthetic pathways and likely were not available from the environment. Therefore, the genetic code evolved as pathways for synthesis of new amino acids became available. One hypothesis proposes that early in the evolution of the genetic code four amino acids-valine, alanine, aspartic acid, and glycine-were coded by GNC codons (N = any base) with the remaining codons being nonsense codons. The other sixteen amino acids were subsequently added to the genetic code by changing nonsense codons into sense codons for these amino acids. Improvement in protein function is presumed to be the driving force behind the evolution of the code, but how improved function was achieved by adding amino acids has not been examined. Based on an analysis of amino acid function in proteins, an evolutionary mechanism for expansion of the genetic code is described in which individual coded amino acids were replaced by new amino acids that used nonsense codons differing by one base change from the sense codons previously used. The improved or altered protein function afforded by the changes in amino acid function provided the selective advantage underlying the expansion of the genetic code. Analysis of amino acid properties and functions explains why amino acids are found in their respective positions in the genetic code.

  19. Ligand specificity and conformational stability of human fatty acid-binding proteins.

    Science.gov (United States)

    Zimmerman, A W; van Moerkerk, H T; Veerkamp, J H

    2001-09-01

    Fatty acid binding proteins (FABPs) are small cytosolic proteins with virtually identical backbone structures that facilitate the solubility and intracellular transport of fatty acids. At least eight different types of FABP occur, each with a specific tissue distribution and possibly with a distinct function. To define the functional characteristics of all eight human FABPs, viz. heart (H), brain (B), myelin (M), adipocyte (A), epidermal (E), intestinal (I), liver (L) and ileal lipid-binding protein (I-LBP), we studied their ligand specificity, their conformational stability and their immunological crossreactivity. Additionally, binding of bile acids to I-LBP was studied. The FABP types showed differences in fatty acid binding affinity. Generally, the affinity for palmitic acid was lower than for oleic and arachidonic acid. All FABP types, except E-FABP, I-FABP and I-LBP interacted with 1-anilinonaphtalene-8-sulphonic acid (ANS). Only L-FABP, I-FABP and M-FABP showed binding of 11-((5-dimethylaminonaphtalene-1-sulfonyl)amino)undecanoic acid (DAUDA). I-LBP showed increasing binding of bile acids in the order taurine-conjugated>glycine-conjugated>unconjugated bile acids. A hydroxylgroup of bile acids at position 7 decreased and at position 12 increased the binding affinity to I-LBP. The fatty acid-binding affinity and the conformation of FABP types were differentially affected in the presence of urea. Our results demonstrate significant differences in ligand binding, conformational stability and surface properties between different FABP types which may point to a specific function in certain cells and tissues. The preference of I-LBP (but not L-FABP) for conjugated bile acids is in accordance with a specific role in bile acid reabsorption in the ileum.

  20. Long-chain n-3 fatty acids - New anabolic compounds improving protein metabolism

    Science.gov (United States)

    Previous animal studies demonstrated that chronic feeding of long-chain n-3 polyunsaturated fatty acids (LCn-3PUFA) that modifies muscle membrane fatty acid composition promotes protein anabolism by blunting the age-associated deterioration in insulin sensitivity. The current study assessed, as a pr...

  1. COMPARATIVE PATHOGENESIS OF HALOACETIC ACID AND PROTEIN KINASE INHIBITOR EMBRYOTOXICITY IN MOUSE WHOLE EMBRYO CULTURE

    Science.gov (United States)

    Comparative pathogenesis of haloacetic acid and protein kinase inhibitor embryotoxicity in mouse whole embryo culture.Ward KW, Rogers EH, Hunter ES 3rd.Curriculum in Toxicology, University of North Carolina at Chapel Hill, 27599-7270, USA.Haloacetic acids ...

  2. Amino acid composition and crude protein values of some Cyanobacteria from Çanakkale (Turkey).

    Science.gov (United States)

    Akgül, Rıza; Kızılkaya, Bayram; Akgül, Füsun; Erduğan, Hüseyin

    2015-09-01

    Cyanobacteria (blue-green algae) form an important component of integrated nutrient managements in agriculture and are exploited in commercial biotechnological ventures. In this study, Rivularia bullata (Poir) Berkeley ex Bornet & Flahault, Nostocs pongiaeforme C. Agardh ex Bornet & Flahault were researched for their amino acid composition and crude protein values. R. bullata was collected from coastal zones of the Gulf of Saros and N. spongiaeforme from the Ayazma Stream. The levels of amino acids were measured in algae samples using EZ: fast kits (EZ: fast GC/FID Protein Hydrolysate Amino Acid Kit) by gas chromatography. The crude proteins of samples were determined by the Kjeldahl method and were calculated using a nitrogen conversion factor of 6.25. Thirty-two amino acids were investigated, for N. spongiaeforme eight free essential amino acids (EAA), eight free non-essential amino acids (NEAA) and eleven other amino acids (OAA); for R. bullata eight EAA, eight NEAA and eight OAA were detected. Aspartic acid is the major constituent for both species. The total protein percents were determined for N. spongiaeforme as % 19.83 and for R. bullata as % 6.15. When considering the increasing world population and reducing natural products; Cyanobacteria will benew feed sources for all living.

  3. Metabolically inert perfluorinated fatty acids directly activate uncoupling protein 1 in brown-fat mitochondria

    OpenAIRE

    Shabalina, Irina G.; Kalinovich, Anastasia V.; Cannon, Barbara; Nedergaard, Jan

    2015-01-01

    The metabolically inert perfluorinated fatty acids perfluorooctane sulfonate (PFOS) and perfluorooctanoate (PFOA) can display fatty acid-like activity in biological systems. The uncoupling protein 1 (UCP1) in brown adipose tissue is physiologically (re)activated by fatty acids, including octanoate. This leads to bioenergetically uncoupled energy dissipation (heat production, thermogenesis). We have examined here the possibility that PFOA/PFOS can directly (re)activate UCP1 in isolated mouse b...

  4. Protein Quantification by Derivatization-Free High-Performance Liquid Chromatography of Aromatic Amino Acids

    Science.gov (United States)

    Hesse, Almut

    2016-01-01

    Amino acid analysis is considered to be the gold standard for quantitative peptide and protein analysis. Here, we would like to propose a simple HPLC/UV method based on a reversed-phase separation of the aromatic amino acids tyrosine (Tyr), phenylalanine (Phe), and optionally tryptophan (Trp) without any derivatization. The hydrolysis of the proteins and peptides was performed by an accelerated microwave technique, which needs only 30 minutes. Two internal standard compounds, homotyrosine (HTyr) and 4-fluorophenylalanine (FPhe) were used for calibration. The limit of detection (LOD) was estimated to be 0.05 µM (~10 µg/L) for tyrosine and phenylalanine at 215 nm. The LOD for a protein determination was calculated to be below 16 mg/L (~300 ng BSA absolute). Aromatic amino acid analysis (AAAA) offers excellent accuracy and a precision of about 5% relative standard deviation, including the hydrolysis step. The method was validated with certified reference materials (CRM) of amino acids and of a pure protein (bovine serum albumin, BSA). AAAA can be used for the quantification of aromatic amino acids, isolated peptides or proteins, complex peptide or protein samples, such as serum or milk powder, and peptides or proteins immobilized on solid supports. PMID:27559481

  5. Total nitrogen vs. amino-acid profile as indicator of protein content of beef.

    Science.gov (United States)

    Hall, Nicolette G; Schönfeldt, Hettie C

    2013-10-01

    In most cited food composition studies and tables, the proximate system measures protein as total nitrogen (N) (determined by Kjeldahl or Dumas method) multiplied by a specific factor. A factor of 6.25 is used for determining total protein from total N (Jones, Munsey, & Walker, 1942). Although more expensive, it is considered more accurate to base protein content of foods on amino acid data (Greenfield & Southgate, 2003). A study on the nutrient composition of beef analysed the full amino-acid profile of fifteen retail cuts from three age groups and six fat codes, as well as determined total nitrogen content to determine proximate protein composition. For all cuts, the correlation coefficient of total amino acids to protein (N×6.25) was 0.635. This indicates a poor correlation for predicting actual protein content (as determined by total amino acid count), based on the nitrogen factor of 6.25. On average, the sum of amino acids per cut amounted to 91% of total determined protein (N×6.25) for the same cut.

  6. Amino acid composition analysis of human secondary transport proteins and implications for reliable membrane topology prediction.

    Science.gov (United States)

    Saidijam, Massoud; Azizpour, Sonia; Patching, Simon G

    2016-07-08

    Secondary transporters in humans are a large group of proteins that transport a wide range of ions, metals, organic and inorganic solutes involved in energy transduction, control of membrane potential and osmotic balance, metabolic processes and in the absorption or efflux of drugs and xenobiotics. They are also emerging as important targets for development of new drugs and as target sites for drug delivery to specific organs or tissues. We have performed amino acid composition (AAC) and phylogenetic analyses and membrane topology predictions for 336 human secondary transport proteins and used the results to confirm protein classification and to look for trends and correlations with structural domains and specific substrates and/or function. Some proteins showed statistically high contents of individual amino acids or of groups of amino acids with similar physicochemical properties. One recurring trend was a correlation between high contents of charged and/or polar residues with misleading results in predictions of membrane topology, which was especially prevalent in Mitochondrial Carrier family proteins. We demonstrate how charged or polar residues located in the middle of transmembrane helices can interfere with their identification by membrane topology tools resulting in missed helices in the prediction. Comparison of AAC in the human proteins with that in 235 secondary transport proteins from Escherichia coli revealed similar overall trends along with differences in average contents for some individual amino acids and groups of similar amino acids that are presumed to result from a greater number of functions and complexity in the higher organism.

  7. Hidden thermodynamic information in protein amino acid mutation tables

    Science.gov (United States)

    Phillips, J. C.

    2017-03-01

    We combine the standard 1992 20 × 20 substitution matrix based on block alignment, BLOSUM62, with the standard 1982 amino acid hydropathicity scale KD as well as the modern 2007 hydropathicity scale MZ, and compare the results. The 20-parameter KD and MZ hydropathicity scales have different thermodynamic character, corresponding to first- and second-order transitions. The KD and MZ comparisons show that the mutation rates reflect quantitative iteration of qualitative amino acid-phobic and -philic binary 2 × 10 properties that define quaternary 4 × 5 subgroups (but not quinary 5 × 4 subgroups), with the modern MZ bioinformatic scale giving much better results. The quaternary 5-mer MZ 4 × 5 subgroups are called mutons (Mu5). Among all hydropathicity scales, the MZ scale uniquely exhibits a smooth, deep mutational minimum at its center associated with alanine, glycine, the smallest amino acid, and histidine.

  8. Reasons for the occurrence of the twenty coded protein amino acids

    Science.gov (United States)

    Weber, A. L.; Miller, S. L.

    1981-01-01

    Factors involved in the selection of the 20 protein L-alpha-amino acids during chemical evolution and the early stages of Darwinian evolution are discussed. The selection is considered on the basis of the availability in the primitive ocean, function in proteins, the stability of the amino acid and its peptides, stability to racemization, and stability on the transfer RNA. It is concluded that aspartic acid, glutamic acid, arginine, lysine, serine and possibly threonine are the best choices for acidic, basic and hydroxy amino acids. The hydrophobic amino acids are reasonable choices, except for the puzzling absences of alpha-amino-n-butyric acid, norvaline and norleucine. The choices of the sulfur and aromatic amino acids seem reasonable, but are not compelling. Asparagine and glutamine are apparently not primitive. If life were to arise on another planet, it would be expected that the catalysts would be poly-alpha-amino acids and that about 75% of the amino acids would be the same as on the earth.

  9. Heart-type fatty-acid-binding protein (FABP3 is a lysophosphatidic acid-binding protein in human coronary artery endothelial cells

    Directory of Open Access Journals (Sweden)

    Ryoko Tsukahara

    2014-01-01

    Full Text Available Fatty-acid-binding protein 3, muscle and heart (FABP3, also known as heart-type FABP, is a member of the family of intracellular lipid-binding proteins. It is a small cytoplasmic protein with a molecular mass of about 15 kDa. FABPs are known to be carrier proteins for transporting fatty acids and other lipophilic substances from the cytoplasm to the nucleus, where these lipids are released to a group of nuclear receptors such as peroxisome proliferator-activated receptors (PPARs. In this study, using lysophosphatidic acid (LPA-coated agarose beads, we have identified FABP3 as an LPA carrier protein in human coronary artery endothelial cells (HCAECs. Administration of LPA to HCAECs resulted in a dose-dependent increase in PPARγ activation. Furthermore, the LPA-induced PPARγ activation was abolished when the FABP3 expression was reduced using small interfering RNA (siRNA. We further show that the nuclear fraction of control HCAECs contained a significant amount of exogenously added LPA, whereas FABP3 siRNA-transfected HCAECs had a decreased level of LPA in the nucleus. Taken together, these results suggest that FABP3 governs the transcriptional activities of LPA by targeting them to cognate PPARγ in the nucleus.

  10. Leucine-Enriched Essential Amino Acids Augment Mixed Protein Synthesis, But Not Collagen Protein Synthesis, in Rat Skeletal Muscle after Downhill Running

    OpenAIRE

    2016-01-01

    Mixed and collagen protein synthesis is elevated for as many as 3 days following exercise. Immediately after exercise, enhanced amino acid availability increases synthesis of mixed muscle protein, but not muscle collagen protein. However, the potential for synergic effects of amino acid ingestion with exercise on both mixed and collagen protein synthesis remains unclear. We investigated muscle collagen protein synthesis in rats following post-exercise ingestion of leucine-enriched essential a...

  11. Analysis of Protein Amino Acids in Tobacco Using Microwave Digestion of Plant Material

    Directory of Open Access Journals (Sweden)

    Moldoveanu SC

    2014-12-01

    Full Text Available This paper describes a technique using microwave digestion and gas chromatography-mass spectrometry (GC-MS, which makes possible the analysis of protein amino acids in tobacco. The technique involves first the measurement of free amino acids, a hydrolysis using microwave digestion, and a measurement of total resulting amino acids. The content of protein amino acids is determined from the difference of total and free amino acids. The digestion is performed with aqueous 6 N HCl (with 1% phenol for two hours in a microwave at 120°C in sealed vials. The GC-MS analysis is performed after the amino acids are derivatized with N-methyl-N-(t-butyldimethylsilyltrifluoroacetamide (MTBSTFA. The technique provides reliable results with less than 10% relative standard deviation (RSD for most amino acids. Only the determination of very low level amino acids is affected by larger errors. The method provides results for free amino acids that are in very good agreement with those obtained by high performance liquid chromatography (HPLC, and also results for protein levels in tobacco in agreement with data previously reported in the literature. Results are given for several single grade tobaccos and for tobacco blends from four Kentucky reference cigarettes.

  12. Protein homeostasis disorders of key enzymes of amino acids metabolism: mutation-induced protein kinetic destabilization and new therapeutic strategies.

    Science.gov (United States)

    Pey, Angel L

    2013-12-01

    Many inborn errors of amino acids metabolism are caused by single point mutations affecting the ability of proteins to fold properly (i.e., protein homeostasis), thus leading to enzyme loss-of-function. Mutations may affect protein homeostasis by altering intrinsic physical properties of the polypeptide (folding thermodynamics, and rates of folding/unfolding/misfolding) as well as the interaction of partially folded states with elements of the protein homeostasis network (such as molecular chaperones and proteolytic machineries). Understanding these mutational effects on protein homeostasis is required to develop new therapeutic strategies aimed to target specific features of the mutant polypeptide. Here, I review recent work in three different diseases of protein homeostasis associated to inborn errors of amino acids metabolism: phenylketonuria, inherited homocystinuria and primary hyperoxaluria type I. These three different genetic disorders involve proteins operating in different cell organelles and displaying different structural complexities. Mutations often decrease protein kinetic stability of the native state (i.e., its half-life for irreversible denaturation), which can be studied using simple kinetic models amenable to biophysical and biochemical characterization. Natural ligands and pharmacological chaperones are shown to stabilize mutant enzymes, thus supporting their therapeutic application to overcome protein kinetic destabilization. The role of molecular chaperones in protein folding and misfolding is also discussed as well as their potential pharmacological modulation as promising new therapeutic approaches. Since current available treatments for these diseases are either burdening or only successful in a fraction of patients, alternative treatments must be considered covering studies from protein structure and biophysics to studies in animal models and patients.

  13. Isoelectric Point, Electric Charge, and Nomenclature of the Acid-Base Residues of Proteins

    Science.gov (United States)

    Maldonado, Andres A.; Ribeiro, Joao M.; Sillero, Antonio

    2010-01-01

    The main object of this work is to present the pedagogical usefulness of the theoretical methods, developed in this laboratory, for the determination of the isoelectric point (pI) and the net electric charge of proteins together with some comments on the naming of the acid-base residues of proteins. (Contains 8 figures and 4 tables.)

  14. 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...

  15. The complete amino acid sequence of the basic nuclear protein of bull spermatozoa

    NARCIS (Netherlands)

    Coelingh, J.P.; Monfoort, Cornelis H.; Rozijn, Thomas H.; Gevers Leuven, Jan A.; Schiphof, R.; Steyn-Parvé, Elizabeth P.; Braunitzer, Gerhard; Schrank, Barbara; Ruhfus, Annette

    1972-01-01

    The complete amino acid sequence of the basic nuclear protein of bull spermatozoa has been established. The sequence was partially deduced by characterization of peptides isolated from thermolysine and chymotryptic digests of the reduced and S-aminoethylated protein. The complete sequence of the fir

  16. Effect of microfluidized and stearic acid modified soy protein in natural rubber

    Science.gov (United States)

    Microfluidized and stearic acid modified soy protein aggregates were used to reinforced natural rubber. The size of soy protein particles was reduced with a microfluidizing and ball milling process. Filler size reduction with longer ball milling time tends to increase tensile strength of the rubber ...

  17. Umami taste amino acids produced by hydrolyzing extracted protein from tomato seed meal

    Science.gov (United States)

    Enzymatic hydrolysis was performed for extracting protein to prepare umami taste amino acids from defatted tomato seed meal (DTSM) which is a by-product of tomato processing. Papain was used as an enzyme for the hydrolysis of DTSM. The particle size distribution of DTSM, protein concentration and fr...

  18. Urinary excretion of fatty acid-binding proteins in idiopathic membranous nephropathy.

    NARCIS (Netherlands)

    Hofstra, J.M.; Deegens, J.K.J.; Steenbergen, E.J.; Wetzels, J.F.M.

    2008-01-01

    BACKGROUND: It is suggested that proteinuria contributes to progressive renal failure by inducing tubular cell injury. The site of injury is unknown. Most studies have used markers of proximal tubular cell damage. Fatty acid-binding proteins (FABPs) are intracellular carrier proteins with different

  19. Isoelectric Point, Electric Charge, and Nomenclature of the Acid-Base Residues of Proteins

    Science.gov (United States)

    Maldonado, Andres A.; Ribeiro, Joao M.; Sillero, Antonio

    2010-01-01

    The main object of this work is to present the pedagogical usefulness of the theoretical methods, developed in this laboratory, for the determination of the isoelectric point (pI) and the net electric charge of proteins together with some comments on the naming of the acid-base residues of proteins. (Contains 8 figures and 4 tables.)

  20. Isolation and characterization of undenatured chlorogenic acid free sunflower (Helianthus annuus) Proteins

    NARCIS (Netherlands)

    Gonzales-Perez, S.; Merck, K.B.; Vereijken, J.M.; Koningsveld, van G.A.; Gruppen, H.; Voragen, A.G.J.

    2002-01-01

    A method for obtaining sunflower protein (SFP) isolate, nondenatured and free of chlorogenic acid (CGA), has been developed. During the isolating procedure, the extent of CGA removal and protein denaturation was monitored. The defatted flour contained 2.5 percent CGA as the main phenolic compound. P

  1. Immunotoxicity of nucleic acid reduced BioProtein - a bacterial derived single cell protein - in Wistar rats

    DEFF Research Database (Denmark)

    Mølck, Anne-marie; Poulsen, Morten; Christensen, Hanne Risager;

    2002-01-01

    BioProtein is a single cell protein produced by a mixed methanotrophic and heterotrophic bacteria culture using natural gas as energy source, which has been approved for animal feed. BioProtein contains a large amount of nucleic acids making the product less suitable for human consumption...... the same tendency, although, not statistically significant (P = 0.09). The subsets of cells identified as neutrophils and eosinophils were increased and lymphocytes decreased. The histopathological examination revealed histiocytosis and accumulation of foamy macrophages in the mesenteric lymph nodes...

  2. Amino acid composition and biological effects of supplementing broad bean and corn proteins with Nigella sativa (black cumin) cake protein.

    Science.gov (United States)

    al-Gaby, A M

    1998-10-01

    The biological effects of supplementing broad bean (Vicia faba) or corn (Zea maize) meal protein with black cumin (Nigella sativa) cake protein as well as their amino acid composition were investigated. The percentage of total protein content of Nigella cake was 22.7%. Lysine is existent in abundant amounts in faba meal protein, while leucine is the most abundant in corn meal protein (chemical score = 156) and valine is higher in Nagella cake protein. compared with rats fed sole corn or faba meal protein, substitution of 25% of corn or faba meal protein with Nigella cake protein in the diet remarkably raised the growth rate of rats and resulted in significant higher levels of rat total serum lipids and triglycerides. Also, the supplemented diet caused significant increases in serum total protein and its two fractions albumin and globulin and insignificantly increase the activity of serum phosphatases and transaminases within normal ranges. The supplementation did not have any adverse nutritional effects in the levels of lipid fractions in the serum.

  3. Specific protein regions influence substrate specificity and product length in polyunsaturated fatty acid condensing enzymes.

    Science.gov (United States)

    Vrinten, Patricia L; Hoffman, Travis; Bauer, Jörg; Qiu, Xiao

    2010-05-11

    We describe a condensing enzyme from Pythium irregulare (PirELO) that shows highest activity on the 18-carbon, Delta-6 desaturated fatty acids, stearidonic acid and gamma-linolenic acid. However, this enzyme is also capable of elongating a number of other fatty acids including the 20-carbon, Delta-5 desaturated fatty acid eicosapentaenoic acid. Surprisingly, a Phytophthora infestans condensing enzyme (PinELO) with very high homology to PirELO did not show activity with 20-carbon fatty acids. A series of chimeric proteins for these two enzymes were constructed to investigate the influence of different regions on substrate and product length. The substitution of a region from near the center of PirELO into PinELO resulted in an enzyme having EPA-elongating activity similar to that of PirELO. Only eight amino acids differed between the two proteins in this region; however, substitution of the same region from PinELO into PirELO produced a protein which was almost inactive. The addition of a small region from near the N-terminus of PinELO was sufficient to restore activity with GLA, indicating that amino acids from these two regions interact to determine protein structure or function. Predicted topology models for PirELO and PinELO placed the two regions described here near the luminal-proximal ends of the first and fourth/fifth transmembrane helixes, at the opposite end of the condensing enzyme from four conserved regions thought to form a catalytic ring. Thus, protein characteristics determined by specific luminal-proximal regions of fatty acid condensing enzymes have a major influence on substrate specificity and final product length.

  4. Protein Content and Amino Acid Composition in Grains of Wheat-Related Species

    Institute of Scientific and Technical Information of China (English)

    JIANG Xiao-ling; TIAN Ji-chun; HAO Zhi; ZHANG Wei-dong

    2008-01-01

    The protein content and amino acid composition for 17 wheat-related species(WRS)and three common wheats(control) were determined and analyzed,and the essential amino acids(EAAs)in WRS were evaluated according to FAO/WHO amino acid recommendations.The results showed that the mean protein content for WRS was 16.67%,which was 23.21% higher than that for the control.The mean contents(g 100 g-1 protein)of most amino acids for WRS were lysine 2.74%,threonine 2.83%,phenylalanine 4.17%,isoleucine 3.42%,valine 3.90%,histidine 2.81%,glutamic acid 29.96%,proline 9.12%,glycine 3.59%,alanine 3.37%,and cysteine 1.57%,which were higher than those for the control.The contents of the other 6 amino acids for WRS were lower than those for the control.The materials(Triticum monococcum L.,Triticum carthlicum Nevski,and Triticum turgidum L.)contained relatively high concentration of the most deficient EAAs(lysine, threonine,and methionine).Comparing with FAO/WHO amino acid recommendations,the amino acid scores(AAS)of lysine(49.8%),threonine(70.7%),and sulfur-containing amino acids(74.8%)were the lowest,which were considered as the main limiting amino acids in WRS.It was observed that the materials with Triticum urartu Tum.(AA)and Aegilops speltoides Tausch.(SS)genomes had relatively high contents of protein and EAA.The contents of protein(16.91%), phenylalanine(4.78%),isoleucine(3.53%),leucine(6.16%),and valine(4.09%)for the diploid materials were higher than those for the other materials.These results will provide some information for selecting parents in breeding about nutrient quality and utilization of fine gene in wheat.

  5. Controlled overproduction of proteins by lactic acid bacteria

    NARCIS (Netherlands)

    Kuipers, Oscar P.; Ruyter, Pascalle G.G.A. de; Kleerebezem, Michiel; Vos, Willem M. de

    1997-01-01

    Lactic acid bacteria are widely used in industrial food fermentations, contributing to flavour, texture and preservation of the fermented products. Here we describe recent advances in the development of controlled gene expression systems, which allow the regulated overproduction of any desirable

  6. Mosaic protein and nucleic acid vaccines against hepatitis C virus

    Science.gov (United States)

    Yusim, Karina; Korber, Bette T. M.; Kuiken, Carla L.; Fischer, William M.

    2013-06-11

    The invention relates to immunogenic compositions useful as HCV vaccines. Provided are HCV mosaic polypeptide and nucleic acid compositions which provide higher levels of T-cell epitope coverage while minimizing the occurrence of unnatural and rare epitopes compared to natural HCV polypeptides and consensus HCV sequences.

  7. Glycation inhibits trichloroacetic acid (TCA)-induced whey protein precipitation

    Science.gov (United States)

    Four different WPI saccharide conjugates were successfully prepared to test whether glycation could inhibit WPI precipitation induced by trichloroacetic acid (TCA). Conjugates molecular weights after glycation were analyzed with SDS-PAGE. No significant secondary structure change due to glycation wa...

  8. Mitogen-activated protein kinase and abscisic acid signal transduction

    NARCIS (Netherlands)

    Heimovaara-Dijkstra, S.; Testerink, C.; Wang, M.

    1998-01-01

    The phytohormone abscisic acid (ABA) is a classical plant hormone, responsible for regulation of abscission, diverse aspects of plant and seed development, stress responses and germination. It was found that ABA signal transduction in plants can involve the activity of type 2C-phosphatases (PP2C), c

  9. Tetrahymena thermophila acidic ribosomal protein L37 contains an archaebacterial type of C-terminus

    DEFF Research Database (Denmark)

    Hansen, T S; Andreasen, P H; Dreisig, H;

    1991-01-01

    We have cloned and characterized a Tetrahymena thermophila macronuclear gene (L37) encoding the acidic ribosomal protein (A-protein) L37. The gene contains a single intron located in the 3'-part of the coding region. Two major and three minor transcription start points (tsp) were mapped 39 to 63...... nucleotides upstream from the translational start codon. The uppermost tsp mapped to the first T in a putative T. thermophila RNA polymerase II initiator element, TATAA. The coding region of L37 predicts a protein of 109 amino acid (aa) residues. A substantial part of the deduced aa sequence was verified...... by protein sequencing. The T. thermophila L37 clearly belongs to the P1-type family of eukaryotic A-proteins, but the C-terminal region has the hallmarks of archaebacterial A-proteins....

  10. Shark myelin basic protein: amino acid sequence, secondary structure, and self-association.

    Science.gov (United States)

    Milne, T J; Atkins, A R; Warren, J A; Auton, W P; Smith, R

    1990-09-01

    Myelin basic protein (MBP) from the Whaler shark (Carcharhinus obscurus) has been purified from acid extracts of a chloroform/methanol pellet from whole brains. The amino acid sequence of the majority of the protein has been determined and compared with the sequences of other MBPs. The shark protein has only 44% homology with the bovine protein, but, in common with other MBPs, it has basic residues distributed throughout the sequence and no extensive segments that are predicted to have an ordered secondary structure in solution. Shark MBP lacks the triproline sequence previously postulated to form a hairpin bend in the molecule. The region containing the putative consensus sequence for encephalitogenicity in the guinea pig contains several substitutions, thus accounting for the lack of activity of the shark protein. Studies of the secondary structure and self-association have shown that shark MBP possesses solution properties similar to those of the bovine protein, despite the extensive differences in primary structure.

  11. Isotopomer distributions in amino acids from a highly expressed protein as a proxy for those from total protein

    Energy Technology Data Exchange (ETDEWEB)

    Shaikh, Afshan; Shaikh, Afshan S.; Tang, Yinjie; Mukhopadhyay, Aindrila; Keasling, Jay D.

    2008-06-27

    {sup 13}C-based metabolic flux analysis provides valuable information about bacterial physiology. Though many biological processes rely on the synergistic functions of microbial communities, study of individual organisms in a mixed culture using existing flux analysis methods is difficult. Isotopomer-based flux analysis typically relies on hydrolyzed amino acids from a homogeneous biomass. Thus metabolic flux analysis of a given organism in a mixed culture requires its separation from the mixed culture. Swift and efficient cell separation is difficult and a major hurdle for isotopomer-based flux analysis of mixed cultures. Here we demonstrate the use of a single highly-expressed protein to analyze the isotopomer distribution of amino acids from one organism. Using the model organism E. coli expressing a plasmid-borne, his-tagged Green Fluorescent Protein (GFP), we show that induction of GFP does not affect E. coli growth kinetics or the isotopomer distribution in nine key metabolites. Further, the isotopomer labeling patterns of amino acids derived from purified GFP and total cell protein are indistinguishable, indicating that amino acids from a purified protein can be used to infer metabolic fluxes of targeted organisms in a mixed culture. This study provides the foundation to extend isotopomer-based flux analysis to study metabolism of individual strains in microbial communities.

  12. Protein and lipid deposition rates in male broiler chickens : separate responses to amino acids and protein-free energy

    NARCIS (Netherlands)

    Eits, R.M.; Kwakkel, R.P.; Verstegen, M.W.A.; Stoutjesdijk, P.; Greef, de K.H.

    2002-01-01

    Two experiments of similar design were conducted with male broiler chickens over two body weight ranges, 200 to 800 g in Experiment 1 and 800 to 1,600 g in Experiment 2. The data were used to test the hypothesis that protein deposition rate increases (linearly) with increasing amino acid intake, unt

  13. An information-theoretic classification of amino acids for the assessment of interfaces in protein-protein docking.

    Science.gov (United States)

    Jardin, Christophe; Stefani, Arno G; Eberhardt, Martin; Huber, Johannes B; Sticht, Heinrich

    2013-09-01

    Docking represents a versatile and powerful method to predict the geometry of protein-protein complexes. However, despite significant methodical advances, the identification of good docking solutions among a large number of false solutions still remains a difficult task. We have previously demonstrated that the formalism of mutual information (MI) from information theory can be adapted to protein docking, and we have now extended this approach to enhance its robustness and applicability. A large dataset consisting of 22,934 docking decoys derived from 203 different protein-protein complexes was used for an MI-based optimization of reduced amino acid alphabets representing the protein-protein interfaces. This optimization relied on a clustering analysis that allows one to estimate the mutual information of whole amino acid alphabets by considering all structural features simultaneously, rather than by treating them individually. This clustering approach is fast and can be applied in a similar fashion to the generation of reduced alphabets for other biological problems like fold recognition, sequence data mining, or secondary structure prediction. The reduced alphabets derived from the present work were converted into a scoring function for the evaluation of docking solutions, which is available for public use via the web service score-MI: http://score-MI.biochem.uni-erlangen.de.

  14. Identification of Acidic pH-dependent Ligands of Pentameric C-reactive Protein*

    OpenAIRE

    Hammond, David J.; Sanjay K. Singh; Thompson, James A.; Beeler, Bradley W.; Rusiñol, Antonio E.; Pangburn, Michael K.; Potempa, Lawrence A.; Agrawal, Alok

    2010-01-01

    C-reactive protein (CRP) is a phylogenetically conserved protein; in humans, it is present in the plasma and at sites of inflammation. At physiological pH, native pentameric CRP exhibits calcium-dependent binding specificity for phosphocholine. In this study, we determined the binding specificities of CRP at acidic pH, a characteristic of inflammatory sites. We investigated the binding of fluid-phase CRP to six immobilized proteins: complement factor H, oxidized low-density lipoprotein, compl...

  15. Regulation of polyisoprenylated methylated protein methyl esterase by polyunsaturated fatty acids and prostaglandins

    OpenAIRE

    Amissah, Felix; Taylor, Shalina; Duverna, Randolph; Ayuk-Takem, Lambert T.; Lamango, Nazarius S

    2011-01-01

    Polyisoprenylation is a set of secondary modifications involving proteins whose aberrant activities are implicated in cancers and degenerative disorders. The last step of the pathway involves an ester-forming polyisoprenylated protein methyl transferase- and hydrolytic polyisoprenylated methylated protein methyl esterase (PMPMEase)-catalyzed reactions. Omega-3 and omega-6 polyunsaturated fatty acids (PUFAs) have been linked with antitumorigeneis and tumorigenesis, respectively. PUFAs are stru...

  16. Production of hydrophobic amino acids from biobased resources: wheat gluten and rubber seed proteins.

    Science.gov (United States)

    Widyarani; Sari, Yessie W; Ratnaningsih, Enny; Sanders, Johan P M; Bruins, Marieke E

    2016-09-01

    Protein hydrolysis enables production of peptides and free amino acids that are suitable for usage in food and feed or can be used as precursors for bulk chemicals. Several essential amino acids for food and feed have hydrophobic side chains; this property may also be exploited for subsequent separation. Here, we present methods for selective production of hydrophobic amino acids from proteins. Selectivity can be achieved by selection of starting material, selection of hydrolysis conditions, and separation of achieved hydrolysate. Several protease combinations were applied for hydrolysis of rubber seed protein concentrate, wheat gluten, and bovine serum albumin (BSA). High degree of hydrolysis (>50 %) could be achieved. Hydrophobic selectivity was influenced by the combination of proteases and by the extent of hydrolysis. Combination of Pronase and Peptidase R showed the highest selectivity towards hydrophobic amino acids, roughly doubling the content of hydrophobic amino acids in the products compared to the original substrates. Hydrophobic selectivity of 0.6 mol-hydrophobic/mol-total free amino acids was observed after 6 h hydrolysis of wheat gluten and 24 h hydrolysis of rubber seed proteins and BSA. The results of experiments with rubber seed proteins and wheat gluten suggest that this process can be applied to agro-industrial residues.

  17. Biopolymers: protein and nucleic acids. Annual report, 15 September 1987-14 September 1988

    Energy Technology Data Exchange (ETDEWEB)

    Richards, J.H.; Abelson, J.N.; Dervan, P.B.; Hood, L.H.; Simon, M.I.

    1988-09-15

    The work focuses on learning the principles that govern interactions between proteins and nucleic acids, both DNA and RNA (specifically tRNA). With these principles as guides peptides (of about 50 amino acids) that bind to specific regions of DNA are being synthesized. Various reactive functionalities are being attached to the synthetic peptides to generate reagents that cleave DNA specifically at the site to which the peptide binds. The work also involves biophysical studies of the protein/nucleic acid complexes in order to expand our understanding of the principles of protein binding to nucleic acids. Development of improved procedures for the chemical synthesis of peptides forms another important aspect of the program.

  18. Accelerated protein digestion and amino acid absorption after Roux-en-Y gastric bypass

    DEFF Research Database (Denmark)

    Bojsen-Møller, Kirstine N; Jacobsen, Siv H; Dirksen, Carsten;

    2015-01-01

    . CONCLUSIONS: RYGB accelerates caseinate digestion and amino acid absorption, resulting in faster and higher but more transient postprandial elevation of plasma amino acids. Changes are likely mediated by accelerated intestinal nutrient entry and clearly demonstrate that protein digestion is not impaired after......BACKGROUND: Roux-en-Y gastric bypass (RYGB) involves exclusion of major parts of the stomach and changes in admixture of gastro-pancreatic enzymes, which could have a major impact on protein digestion and amino acid absorption. OBJECTIVE: We investigated the effect of RYGB on amino acid appearance...... in the systemic circulation from orally ingested protein and from endogenous release. DESIGN: Nine obese glucose-tolerant subjects, with a mean body mass index (in kg/m(2)) of 39.2 (95% CI: 35.2, 43.3) and mean glycated hemoglobin of 5.3% (95% CI: 4.9%, 5.6%), were studied before and 3 mo after RYGB. Leucine...

  19. Prediction algorithm for amino acid types with their secondary structure in proteins (PLATON) using chemical shifts.

    Science.gov (United States)

    Labudde, D; Leitner, D; Krüger, M; Oschkinat, H

    2003-01-01

    The algorithm PLATON is able to assign sets of chemical shifts derived from a single residue to amino acid types with its secondary structure (amino acid species). A subsequent ranking procedure using optionally two different penalty functions yields predictions for possible amino acid species for the given set of chemical shifts. This was demonstrated in the case of the alpha-spectrin SH3 domain and applied to 9 further protein data sets taken from the BioMagRes database. A database consisting of reference chemical shift patterns (reference CSPs) was generated from assigned chemical shifts of proteins with known 3D-structure. This reference CSP database is used in our approach for extracting distributions of amino acid types with their most likely secondary structure elements (namely alpha-helix, beta-sheet, and coil) for single amino acids by comparison with query CSPs. Results obtained for the 10 investigated proteins indicates that the percentage of correct amino acid species in the first three positions in the ranking list, ranges from 71.4% to 93.2% for the more favorable penalty function. Where only the top result of the ranking list for these 10 proteins is considered, 36.5% to 83.1% of the amino acid species are correctly predicted. The main advantage of our approach, over other methods that rely on average chemical shift values is the ability to increase database content by incorporating newly derived CSPs, and therefore to improve PLATON's performance over time.

  20. Fatty acid transport protein expression in human brain and potential role in fatty acid transport across human brain microvessel endothelial cells.

    Science.gov (United States)

    Mitchell, Ryan W; On, Ngoc H; Del Bigio, Marc R; Miller, Donald W; Hatch, Grant M

    2011-05-01

    The blood-brain barrier (BBB), formed by the brain capillary endothelial cells, provides a protective barrier between the systemic blood and the extracellular environment of the CNS. Passage of fatty acids from the blood to the brain may occur either by diffusion or by proteins that facilitate their transport. Currently several protein families have been implicated in fatty acid transport. The focus of the present study was to identify the fatty acid transport proteins (FATPs) expressed in the brain microvessel endothelial cells and characterize their involvement in fatty acid transport across an in vitro BBB model. The major fatty acid transport proteins expressed in human brain microvessel endothelial cells (HBMEC), mouse capillaries and human grey matter were FATP-1, -4 and fatty acid binding protein 5 and fatty acid translocase/CD36. The passage of various radiolabeled fatty acids across confluent HBMEC monolayers was examined over a 30-min period in the presence of fatty acid free albumin in a 1 : 1 molar ratio. The apical to basolateral permeability of radiolabeled fatty acids was dependent upon both saturation and chain length of the fatty acid. Knockdown of various fatty acid transport proteins using siRNA significantly decreased radiolabeled fatty acid transport across the HBMEC monolayer. Our findings indicate that FATP-1 and FATP-4 are the predominant fatty acid transport proteins expressed in the BBB based on human and mouse expression studies. While transport studies in HBMEC monolayers support their involvement in fatty acid permeability, fatty acid translocase/CD36 also appears to play a prominent role in transport of fatty acids across HBMEC.

  1. Protein and Essential Amino Acids to Protect Musculoskeletal Health during Spaceflight: Evidence of a Paradox?

    Directory of Open Access Journals (Sweden)

    Kyle J. Hackney

    2014-07-01

    Full Text Available Long-duration spaceflight results in muscle atrophy and a loss of bone mineral density. In skeletal muscle tissue, acute exercise and protein (e.g., essential amino acids stimulate anabolic pathways (e.g., muscle protein synthesis both independently and synergistically to maintain neutral or positive net muscle protein balance. Protein intake in space is recommended to be 12%–15% of total energy intake (≤1.4 g∙kg−1∙day−1 and spaceflight is associated with reduced energy intake (~20%, which enhances muscle catabolism. Increasing protein intake to 1.5–2.0 g∙kg−1∙day−1 may be beneficial for skeletal muscle tissue and could be accomplished with essential amino acid supplementation. However, increased consumption of sulfur-containing amino acids is associated with increased bone resorption, which creates a dilemma for musculoskeletal countermeasures, whereby optimizing skeletal muscle parameters via essential amino acid supplementation may worsen bone outcomes. To protect both muscle and bone health, future unloading studies should evaluate increased protein intake via non-sulfur containing essential amino acids or leucine in combination with exercise countermeasures and the concomitant influence of reduced energy intake.

  2. Distribution and Variation of Ribonucleic Acid (RNA) and Protein and Its Hydrolysis Products in Lake Sediments

    Institute of Scientific and Technical Information of China (English)

    梁小兵; 万国江; 黄荣贵

    2002-01-01

    Protein and RNA in lake sediments tend to be decomposed progressively with time and sedimentation depth. Their concentrations tend to decrease starting from the sedimentation depth of 17 cm and that of 19 cm, respectively. However, the products of their decomposition-amino acids and nucleotides show different rules of variation. At the depth from 27 cm to 30 cm the amino acids are most abundant in the pore waters of lake sediments. Such variation tendency seems to be related to the extent to which microbes utilize amino acids and nucleotides. Due to polymerization in the geological processes and the adsorption of protein on minerals and organic polymers, below the sedimentation depth of 17 cm there is still a certain amount of protein in the sediments. With the time passing by, protein has been well preserved in various sediment layers, indicating that its decomposition is relatively limited. The peak values of protein content in the sediments of the two lakes are produced in the surface layers at the depth of 10 cm, implicating that the surface sediments are favorable to the release of protein.The contents of amino acids in the pore waters of lake sediments are closely related to the activities of microbes. Below the depth of 27 cm, the amino acids are significantly accumulated in Lake Aha sediments, probably indicating the weakening of microbial activities.

  3. Lipid binding protein response to a bile acid library: a combined NMR and statistical approach.

    Science.gov (United States)

    Tomaselli, Simona; Pagano, Katiuscia; Boulton, Stephen; Zanzoni, Serena; Melacini, Giuseppe; Molinari, Henriette; Ragona, Laura

    2015-11-01

    Primary bile acids, differing in hydroxylation pattern, are synthesized from cholesterol in the liver and, once formed, can undergo extensive enzyme-catalysed glycine/taurine conjugation, giving rise to a complex mixture, the bile acid pool. Composition and concentration of the bile acid pool may be altered in diseases, posing a general question on the response of the carrier (bile acid binding protein) to the binding of ligands with different hydrophobic and steric profiles. A collection of NMR experiments (H/D exchange, HET-SOFAST, ePHOGSY NOESY/ROESY and (15) N relaxation measurements) was thus performed on apo and five different holo proteins, to monitor the binding pocket accessibility and dynamics. The ensemble of obtained data could be rationalized by a statistical approach, based on chemical shift covariance analysis, in terms of residue-specific correlations and collective protein response to ligand binding. The results indicate that the same residues are influenced by diverse chemical stresses: ligand binding always induces silencing of motions at the protein portal with a concomitant conformational rearrangement of a network of residues, located at the protein anti-portal region. This network of amino acids, which do not belong to the binding site, forms a contiguous surface, sensing the presence of the bound lipids, with a signalling role in switching protein-membrane interactions on and off.

  4. A biotin enrichment strategy identifies novel carbonylated amino acids in proteins from human plasma.

    Science.gov (United States)

    Havelund, Jesper F; Wojdyla, Katarzyna; Davies, Michael J; Jensen, Ole N; Møller, Ian Max; Rogowska-Wrzesinska, Adelina

    2017-03-06

    Protein carbonylation is an irreversible protein oxidation correlated with oxidative stress, various diseases and ageing. Here we describe a peptide-centric approach for identification and characterisation of up to 14 different types of carbonylated amino acids in proteins. The modified residues are derivatised with biotin-hydrazide, enriched and characterised by tandem mass spectrometry. The strength of the method lies in an improved elution of biotinylated peptides from monomeric avidin resin using hot water (95°C) and increased sensitivity achieved by reduction of analyte losses during sample preparation and chromatography. For the first time MS/MS data analysis utilising diagnostic biotin fragment ions is used to pinpoint sites of biotin labelling and improve the confidence of carbonyl peptide assignments. We identified a total of 125 carbonylated residues in bovine serum albumin after extensive in vitro metal ion-catalysed oxidation. Furthermore, we assigned 133 carbonylated sites in 36 proteins in native human plasma protein samples. The optimised workflow enabled detection of 10 hitherto undetected types of carbonylated amino acids in proteins: aldehyde and ketone modifications of leucine, valine, alanine, isoleucine, glutamine, lysine and glutamic acid (+14Da), an oxidised form of methionine - aspartate semialdehyde (-32Da) - and decarboxylated glutamic acid and aspartic acid (-30Da).

  5. Interference from alpha-amino acid and protein on determination of formaldehyde in food

    Science.gov (United States)

    Lu, Xiumin; Zhang, Xiaofeng; Fu, Yujie; Xiang, Jinxin

    2005-12-01

    The disturbance of alpha-amino acids and proteins on the analysis of formaldehyde content in food was investigated by electrochemical assay. Results show that the pH decreases gradually from 9.91 to 4.36 with increasing aspartic acid concentration. The recovery rate changes from 8% to 100% after different amounts of formaldehyde were added into protein solutions. For edible bamboo shoots, the recovery rate of formaldehyde is 80% to 100%. For shrimp kernel, however, the recovery rate of formaldehyde is 8% to 60%. These results indicate that the consumed quantity of formaldehyde is correlative with the protein concentration in foods. Therefore, the determinate formaldehyde content in food is actually not the totally applied amount, but just the residue after its reaction with the alpha-amino acids or free amino groups on the protein surface.

  6. Incorporation of noncanonical amino acids into Rosetta and use in computational protein-peptide interface design.

    Directory of Open Access Journals (Sweden)

    P Douglas Renfrew

    Full Text Available Noncanonical amino acids (NCAAs can be used in a variety of protein design contexts. For example, they can be used in place of the canonical amino acids (CAAs to improve the biophysical properties of peptides that target protein interfaces. We describe the incorporation of 114 NCAAs into the protein-modeling suite Rosetta. We describe our methods for building backbone dependent rotamer libraries and the parameterization and construction of a scoring function that can be used to score NCAA containing peptides and proteins. We validate these additions to Rosetta and our NCAA-rotamer libraries by showing that we can improve the binding of a calpastatin derived peptides to calpain-1 by substituting NCAAs for native amino acids using Rosetta. Rosetta (executables and source, auxiliary scripts and code, and documentation can be found at (http://www.rosettacommons.org/.

  7. G-protein-coupled receptors for free fatty acids

    DEFF Research Database (Denmark)

    Milligan, Graeme; Ulven, Trond; Murdoch, Hannah;

    2014-01-01

    It is becoming evident that nutrients and metabolic intermediates derived from such nutrients regulate cellular function by activating a number of cell-surface G-protein coupled receptors (GPCRs). Until now, members of the GPCR family have largely been considered as the molecular targets that com...

  8. Leucine-Enriched Essential Amino Acids Augment Mixed Protein Synthesis, But Not Collagen Protein Synthesis, in Rat Skeletal Muscle after Downhill Running

    Directory of Open Access Journals (Sweden)

    Hiroyuki Kato

    2016-06-01

    Full Text Available Mixed and collagen protein synthesis is elevated for as many as 3 days following exercise. Immediately after exercise, enhanced amino acid availability increases synthesis of mixed muscle protein, but not muscle collagen protein. However, the potential for synergic effects of amino acid ingestion with exercise on both mixed and collagen protein synthesis remains unclear. We investigated muscle collagen protein synthesis in rats following post-exercise ingestion of leucine-enriched essential amino acids. We determined fractional protein synthesis rates (FSR at different time points following exercise. Mixed protein and collagen protein FSRs in skeletal muscle were determined by measuring protein-bound enrichments of hydroxyproline and proline, and by measuring the intracellular enrichment of proline, using injections of flooding d3-proline doses. A leucine-enriched mixture of essential amino acids (or distilled water as a control was administrated 30 min or 1 day post-exercise. The collagen protein synthesis in the vastus lateralis was elevated for 2 days after exercise. Although amino acid administration did not increase muscle collagen protein synthesis, it did lead to augmented mixed muscle protein synthesis 1 day following exercise. Thus, contrary to the regulation of mixed muscle protein synthesis, muscle collagen protein synthesis is not affected by amino acid availability after damage-inducing exercise.

  9. Leucine-Enriched Essential Amino Acids Augment Mixed Protein Synthesis, But Not Collagen Protein Synthesis, in Rat Skeletal Muscle after Downhill Running.

    Science.gov (United States)

    Kato, Hiroyuki; Suzuki, Hiromi; Inoue, Yoshiko; Suzuki, Katsuya; Kobayashi, Hisamine

    2016-06-28

    Mixed and collagen protein synthesis is elevated for as many as 3 days following exercise. Immediately after exercise, enhanced amino acid availability increases synthesis of mixed muscle protein, but not muscle collagen protein. However, the potential for synergic effects of amino acid ingestion with exercise on both mixed and collagen protein synthesis remains unclear. We investigated muscle collagen protein synthesis in rats following post-exercise ingestion of leucine-enriched essential amino acids. We determined fractional protein synthesis rates (FSR) at different time points following exercise. Mixed protein and collagen protein FSRs in skeletal muscle were determined by measuring protein-bound enrichments of hydroxyproline and proline, and by measuring the intracellular enrichment of proline, using injections of flooding d₃-proline doses. A leucine-enriched mixture of essential amino acids (or distilled water as a control) was administrated 30 min or 1 day post-exercise. The collagen protein synthesis in the vastus lateralis was elevated for 2 days after exercise. Although amino acid administration did not increase muscle collagen protein synthesis, it did lead to augmented mixed muscle protein synthesis 1 day following exercise. Thus, contrary to the regulation of mixed muscle protein synthesis, muscle collagen protein synthesis is not affected by amino acid availability after damage-inducing exercise.

  10. Cinnamic Acid and Its Derivatives Inhibit Fructose-Mediated Protein Glycation

    Directory of Open Access Journals (Sweden)

    Sirintorn Yibchok-anun

    2012-02-01

    Full Text Available Cinnamic acid and its derivatives have shown a variety of pharmacologic properties. However, little is known about the antiglycation properties of cinnamic acid and its derivatives. The present study sought to characterize the protein glycation inhibitory activity of cinnamic acid and its derivatives in a bovine serum albumin (BSA/fructose system. The results demonstrated that cinnamic acid and its derivatives significantly inhibited the formation of advanced glycation end products (AGEs by approximately 11.96–63.36% at a concentration of 1 mM. The strongest inhibitory activity against the formation of AGEs was shown by cinnamic acid. Furthermore, cinnamic acid and its derivatives reduced the level of fructosamine, the formation of Nε-(carboxymethyl lysine (CML, and the level of amyloid cross β-structure. Cinnamic acid and its derivatives also prevented oxidative protein damages, including effects on protein carbonyl formation and thiol oxidation of BSA. Our findings may lead to the possibility of using cinnamic acid and its derivatives for preventing AGE-mediated diabetic complications.

  11. Revisiting amino acid substitution matrices for identifying distantly related proteins.

    Science.gov (United States)

    Yamada, Kazunori; Tomii, Kentaro

    2014-02-01

    Although many amino acid substitution matrices have been developed, it has not been well understood which is the best for similarity searches, especially for remote homology detection. Therefore, we collected information related to existing matrices, condensed it and derived a novel matrix that can detect more remote homology than ever. Using principal component analysis with existing matrices and benchmarks, we developed a novel matrix, which we designate as MIQS. The detection performance of MIQS is validated and compared with that of existing general purpose matrices using SSEARCH with optimized gap penalties for each matrix. Results show that MIQS is able to detect more remote homology than the existing matrices on an independent dataset. In addition, the performance of our developed matrix was superior to that of CS-BLAST, which was a novel similarity search method with no amino acid matrix. We also evaluated the alignment quality of matrices and methods, which revealed that MIQS shows higher alignment sensitivity than that with the existing matrix series and CS-BLAST. Fundamentally, these results are expected to constitute good proof of the availability and/or importance of amino acid matrices in sequence analysis. Moreover, with our developed matrix, sophisticated similarity search methods such as sequence-profile and profile-profile comparison methods can be improved further. Newly developed matrices and datasets used for this study are available at http://csas.cbrc.jp/Ssearch/.

  12. Role of Protein and Amino Acids in Infant and Young Child Nutrition: Protein and Amino Acid Needs and Relationship with Child Growth.

    Science.gov (United States)

    Uauy, Ricardo; Kurpad, Anura; Tano-Debrah, Kwaku; Otoo, Gloria E; Aaron, Grant A; Toride, Yasuhiko; Ghosh, Shibani

    2015-01-01

    Over a third of all deaths of children under the age of five are linked to undernutrition. At a 90% coverage level, a core group of ten interventions inclusive of infant and young child nutrition could save one million lives of children under 5 y of age (15% of all deaths) (Lancet 2013). The infant and young child nutrition package alone could save over 220,000 lives in children under 5 y of age. High quality proteins (e.g. milk) in complementary, supplementary and rehabilitation food products have been found to be effective for good growth. Individual amino acids such as lysine and arginine have been found to be factors linked to growth hormone release in young children via the somatotropic axis and high intakes are inversely associated with fat mass index in pre-pubertal lean girls. Protein intake in early life is positively associated with height and weight at 10 y of age. This paper will focus on examining the role of protein and amino acids in infant and young child nutrition by examining protein and amino acid needs in early life and the subsequent relationship with stunting.

  13. Maternal Low Quality Protein Diet Alters Plasma Amino Acid Concentrations of Weaning Rats

    Directory of Open Access Journals (Sweden)

    Arzu Kabasakal Cetin

    2015-12-01

    Full Text Available Several studies have indicated the influence of a maternal low protein diet on the fetus. However, the effect of a maternal low quality protein diet on fetal growth and development is largely unknown. Wistar rats (11 weeks old were mated and maintained on either a chow diet with 20% casein (n = 6 as the control group (C, or a low quality protein diet with 20% wheat gluten (n = 7 as the experimental group (WG through gestation and lactation. Maternal body weights were similar in both groups throughout the study. Birth weights were not influenced by maternal diet and offspring body weights during lactation were similar between the groups. Offspring’s plasma amino acid profiles showed that plasma methionine, glutamine and lysine were significantly lower and aspartic acid, ornithine and glycine-proline were significantly higher in the WG. Plant based protein comprises an important part of protein intake in developing countries. It is well-known that these diets can be inadequate in terms of essential amino acids. The current study shows differential effects of a maternal low quality protein diet on the offspring’s plasma amino acids. Future studies will examine further aspects of the influence of maternal low quality protein diets on fetal growth and development.

  14. The folding type of a protein is relevant to the amino acid composition.

    Science.gov (United States)

    Nakashima, H; Nishikawa, K; Ooi, T

    1986-01-01

    The folding types of 135 proteins, the three-dimensional structures of which are known, were analyzed in terms of the amino acid composition. The amino acid composition of a protein was expressed as a point in a multidimensional space spanned with 20 axes, on which the corresponding contents of 20 amino acids in the protein were represented. The distribution pattern of proteins in this composition space was examined in relation to five folding types, alpha, beta, alpha/beta, alpha + beta, and irregular type. The results show that amino acid compositions of the alpha, beta, and alpha/beta types are located in different regions in the composition space, thus allowing distinct separation of proteins depending on the folding types. The points representing proteins of the alpha + beta and irregular types, however, are widely scattered in the space, and the existing regions overlap with those of the other folding types. A simple method of utilizing the "distance" in the space was found to be convenient for classification of proteins into the five folding types. The assignment of the folding type with this method gave an accuracy of 70% in the coincidence with the experimental data.

  15. Role of gamma-aminobutyricacidB(GABA(B)) receptors in the regulation of kainic acid-induced cell death in mouse hippocampus.

    Science.gov (United States)

    Lee, Han Kyu; Seo, Young Jun; Choi, Seong Soo; Kwon, Min Soo; Shim, Eon Jeong; Lee, Jin Young; Suh, Hong Won

    2005-12-31

    Kainic acid (KA) is well-known as an excitatory, neurotoxic substance. In mice, KA administered intracerebroventricularly (i.c.v.) lead to morphological damage of hippocampus expecially concentrated on the CA3 pyramidal neurons. In the present study, the possible role of gamma-aminobutyric acid B (GABA(B)) receptors in hippocampal cell death induced by KA (0.1 microg) administered i.c.v. was examined. 5-Aminovaleric acid (5-AV; GABA(B) receptors antagonist, 20 mug) reduced KA-induced CA3 pyramidal cell death. KA increased the phosphorylated extracellular signal-regulated kinase (p-ERK) and Ca(2+)/calmodulin-dependent protein kinase II (p-CaMK II) immunoreactivities (IRs) 30 min after KA treatment, and c-Fos, c-Jun IR 2 h, and glial fibrillary acidic protein (GFAP), complement receptor type 3 (OX-42) IR 1 day in hippocampal area in KA-injected mice. 5-AV attenuated KA-induced p-CaMK II, GFAP and OX-42 IR in the hippocampal CA3 region. These results suggest that p-CaMK II may play as an important regulator on hippocampal cell death induced by KA administered i.c.v. in mice. Activated astrocytes, which was presented by GFAP IR, and activated microglia, which was presented by the OX-42 IR, may be a good indicator for measuring the cell death in hippocampal regions by KA excitotoxicity. Furthermore, it showed that GABA(B) receptors appear to be involved in hippocampal CA3 pyramidal cell death induced by KA administered i.c.v. in mice.

  16. Sequential activation of microglia and astrocyte cytokine expression precedes increased Iba-1 or GFAP immunoreactivity following systemic immune challenge.

    Science.gov (United States)

    Norden, Diana M; Trojanowski, Paige J; Villanueva, Emmanuel; Navarro, Elisa; Godbout, Jonathan P

    2016-02-01

    Activation of the peripheral immune system elicits a coordinated response from the central nervous system. Key to this immune to brain communication is that glia, microglia, and astrocytes, interpret and propagate inflammatory signals in the brain that influence physiological and behavioral responses. One issue in glial biology is that morphological analysis alone is used to report on glial activation state. Therefore, our objective was to compare behavioral responses after in vivo immune (lipopolysaccharide, LPS) challenge to glial specific mRNA and morphological profiles. Here, LPS challenge induced an immediate but transient sickness response with decreased locomotion and social interaction. Corresponding with active sickness behavior (2-12 h), inflammatory cytokine mRNA expression was elevated in enriched microglia and astrocytes. Although proinflammatory cytokine expression in microglia peaked 2-4 h after LPS, astrocyte cytokine, and chemokine induction was delayed and peaked at 12 h. Morphological alterations in microglia (Iba-1(+)) and astrocytes (GFAP(+)), however, were undetected during this 2-12 h timeframe. Increased Iba-1 immunoreactivity and de-ramified microglia were evident 24 and 48 h after LPS but corresponded to the resolution phase of activation. Morphological alterations in astrocytes were undetected after LPS. Additionally, glial cytokine expression did not correlate with morphology after four repeated LPS injections. In fact, repeated LPS challenge was associated with immune and behavioral tolerance and a less inflammatory microglial profile compared with acute LPS challenge. Overall, induction of glial cytokine expression was sequential, aligned with active sickness behavior, and preceded increased Iba-1 or GFAP immunoreactivity after LPS challenge.

  17. Defining meal requirements for protein to optimize metabolic roles of amino acids.

    Science.gov (United States)

    Layman, Donald K; Anthony, Tracy G; Rasmussen, Blake B; Adams, Sean H; Lynch, Christopher J; Brinkworth, Grant D; Davis, Teresa A

    2015-04-29

    Dietary protein provides essential amino acids (EAAs) for the synthesis of new proteins plus an array of other metabolic functions; many of these functions are sensitive to postprandial plasma and intracellular amino acid concentrations. Recent research has focused on amino acids as metabolic signals that influence the rate of protein synthesis, inflammation responses, mitochondrial activity, and satiety, exerting their influence through signaling systems including mammalian/mechanistic target of rapamycin complex 1 (mTORC1), general control nonrepressed 2 (GCN2), glucagon-like peptide 1 (GLP-1), peptide YY (PYY), serotonin, and insulin. These signals represent meal-based responses to dietary protein. The best characterized of these signals is the leucine-induced activation of mTORC1, which leads to the stimulation of skeletal muscle protein synthesis after ingestion of a meal that contains protein. The response of this metabolic pathway to dietary protein (i.e., meal threshold) declines with advancing age or reduced physical activity. Current dietary recommendations for protein are focused on total daily intake of 0.8 g/kg body weight, but new research suggests daily needs for older adults of ≥1.0 g/kg and identifies anabolic and metabolic benefits to consuming at least 20-30 g protein at a given meal. Resistance exercise appears to increase the efficiency of EAA use for muscle anabolism and to lower the meal threshold for stimulation of protein synthesis. Applying this information to a typical 3-meal-a-day dietary plan results in protein intakes that are well within the guidelines of the Dietary Reference Intakes for acceptable macronutrient intakes. The meal threshold concept for dietary protein emphasizes a need for redistribution of dietary protein for optimum metabolic health.

  18. Cellular nucleic acid binding protein binds G-rich single-stranded nucleic acids and may function as a nucleic acid chaperone.

    Science.gov (United States)

    Armas, Pablo; Nasif, Sofía; Calcaterra, Nora B

    2008-02-15

    Cellular nucleic acid binding protein (CNBP) is a small single-stranded nucleic acid binding protein made of seven Zn knuckles and an Arg-Gly rich box. CNBP is strikingly conserved among vertebrates and was reported to play broad-spectrum functions in eukaryotic cells biology. Neither its biological function nor its mechanisms of action were elucidated yet. The main goal of this work was to gain further insights into the CNBP biochemical and molecular features. We studied Bufo arenarum CNBP (bCNBP) binding to single-stranded nucleic acid probes representing the main reported CNBP putative targets. We report that, although bCNBP is able to bind RNA and single-stranded DNA (ssDNA) probes in vitro, it binds RNA as a preformed dimer whereas both monomer and dimer are able to bind to ssDNA. A systematic analysis of variant probes shows that the preferred bCNBP targets contain unpaired guanosine-rich stretches. These data expand the knowledge about CNBP binding stoichiometry and begins to dissect the main features of CNBP nucleic acid targets. Besides, we show that bCNBP presents a highly disordered predicted structure and promotes the annealing and melting of nucleic acids in vitro. These features are typical of proteins that function as nucleic acid chaperones. Based on these data, we propose that CNBP may function as a nucleic acid chaperone through binding, remodeling, and stabilizing nucleic acids secondary structures. This novel CNBP biochemical activity broadens the field of study about its biological function and may be the basis to understand the diverse ways in which CNBP controls gene expression.

  19. Photo-CIDNP studies of amino acids and proteins

    CERN Document Server

    Lopez, J J

    2001-01-01

    half. Chapter 6: CIDNP in Micellar Solutions The presence of detergent, below and above the critical micelle concentration, is shown to affect CIDNP intensities, due to electrostatic interactions between charged dye and detergent molecules. In the last part of this chapter, photo-CIDNP experiments with the membrane protein gramicidin A, incorporated in detergent and lipid micelles, are described. Chapter 7: CIDNP Study of the Tryptophan Radical CIDNP spectra are characteristic of the transient radical intermediates which are generated during the flash photolysis. Here, the tryptophan radicals g-factor is estimated with the help of the CIDNP dependence on the magnetic field in which the flash-photolysis takes place. The ultimate aim of the research described in this thesis is the development of methods with which ope may study the structure and function of proteins on a molecular level. This is done with the help of a combination of NMR (Nuclear Magnetic Resonance) and flash photolysis, in which light initiate...

  20. Highly Charged Protein Ions: The Strongest Organic Acids to Date.

    Science.gov (United States)

    Zenaidee, Muhammad A; Leeming, Michael G; Zhang, Fangtong; Funston, Toby T; Donald, William A

    2017-07-10

    The basicity of highly protonated cytochrome c (cyt c) and myoglobin (myo) ions were investigated using tandem mass spectrometry, ion-molecule reactions (IMRs), and theoretical calculations as a function of charge state. Surprisingly, highly charged protein ions (HCPI) can readily protonate non-polar molecules and inert gases, including Ar, O2 , and N2 in thermal IMRs. The most HCPIs that can be observed are over 130 kJ mol(-1) less basic than the least basic neutral organic molecules known (tetrafluoromethane and methane). Based on theoretical calculations, it is predicted that protonated cyt c and myo ions should spontaneously lose a proton to vacuum for charge states in which every third residue is protonated. In this study, HCPIs are formed where every fourth residue on average is protonated. These results indicate that protein ions in higher charge states can be formed using a low-pressure ion source to reduce proton-transfer reactions between protein ions and gases from the atmosphere. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Heterologous protein display on the cell surface of lactic acid bacteria mediated by the s-layer protein

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    Han Lanlan

    2011-10-01

    Full Text Available Abstract Background Previous studies have revealed that the C-terminal region of the S-layer protein from Lactobacillus is responsible for the cell wall anchoring, which provide an approach for targeting heterologous proteins to the cell wall of lactic acid bacteria (LAB. In this study, we developed a new surface display system in lactic acid bacteria with the C-terminal region of S-layer protein SlpB of Lactobacillus crispatus K2-4-3 isolated from chicken intestine. Results Multiple sequence alignment revealed that the C-terminal region (LcsB of Lb. crispatus K2-4-3 SlpB had a high similarity with the cell wall binding domains SA and CbsA of Lactobacillus acidophilus and Lb. crispatus. To evaluate the potential application as an anchoring protein, the green fluorescent protein (GFP or beta-galactosidase (Gal was fused to the N-terminus of the LcsB region, and the fused proteins were successfully produced in Escherichia coli, respectively. After mixing them with the non-genetically modified lactic acid bacteria cells, the fused GFP-LcsB and Gal-LcsB were functionally associated with the cell surface of various lactic acid bacteria tested. In addition, the binding capacity could be improved by SDS pretreatment. Moreover, both of the fused proteins could simultaneously bind to the surface of a single cell. Furthermore, when the fused DNA fragment of gfp:lcsB was inserted into the Lactococcus lactis expression vector pSec:Leiss:Nuc, the GFP could not be secreted into the medium under the control of the nisA promoter. Western blot, in-gel fluorescence assay, immunofluorescence microscopy and SDS sensitivity analysis confirmed that the GFP was successfully expressed onto the cell surface of L. lactis with the aid of the LcsB anchor. Conclusion The LcsB region can be used as a functional scaffold to target the heterologous proteins to the cell surfaces of lactic acid bacteria in vitro and in vivo, and has also the potential for biotechnological

  2. The "Jekyll and Hyde" Actions of Nucleic Acids on the Prion-like Aggregation of Proteins.

    Science.gov (United States)

    Silva, Jerson L; Cordeiro, Yraima

    2016-07-22

    Protein misfolding results in devastating degenerative diseases and cancer. Among the culprits involved in these illnesses are prions and prion-like proteins, which can propagate by converting normal proteins to the wrong conformation. For spongiform encephalopathies, a real prion can be transmitted among individuals. In other disorders, the bona fide prion characteristics are still under investigation. Besides inducing misfolding of native proteins, prions bind nucleic acids and other polyanions. Here, we discuss how nucleic acid binding might influence protein misfolding for both disease-related and benign, functional prions and why the line between bad and good amyloids might be more subtle than previously thought. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  3. Tracers to investigate protein and amino acid metabolism in human subjects.

    Science.gov (United States)

    Wagenmakers, A J

    1999-11-01

    Three tracer methods have been used to measure protein synthesis, protein breakdown and protein oxidation at whole-body level. The method using L-[1-(13)C]leucine is considered the method of reference. These methods have contributed greatly to the existing knowledge on whole-body protein turnover and its regulation by feeding, fasting, hormones and disease. How exercise and ingestion of mixed protein-containing meals affect whole-body protein metabolism is still open to debate, as there are discrepancies in results obtained with different tracers. The contribution of whole-body methods to the future gain of knowledge is expected to be limited due to the fact that most physiological disturbances have been investigated extensively, and due to the lack of information on the relative contribution of various tissues and proteins to whole-body changes. Tracer amino acid-incorporation methods are most suited to investigate these latter aspects of protein metabolism. These methods have shown that some tissues (liver and gut) have much higher turnover rates and deposit much more protein than others (muscle). Massive differences also exist between the fractional synthesis rates of individual proteins. The incorporation methods have been properly validated, although minor disagreements remain on the identity of the true precursor pool (the enrichment of which should be used in the calculations). Arterio-venous organ balance studies have shown that little protein is deposited in skeletal muscle following a protein-containing meal, while much more protein is deposited in liver and gut. The amount deposited in the feeding period in each of these tissues is released again during overnight fasting. The addition of tracers to organ balance studies allows the simultaneous estimation of protein synthesis and protein breakdown, and provides information on whether changes in net protein balance are caused primarily by a change in protein synthesis or in protein breakdown. In the case

  4. Seasonal variations in the amino acid profile and protein nutritional value of Saccharina latissima cultivated in a commercial IMTA system

    DEFF Research Database (Denmark)

    Silva Marinho, Goncalo; Holdt, Susan Løvstad; Angelidaki, Irini

    2015-01-01

    Seaweeds have potential for the provision of biomass for food and feed supplements. The demand is increasing especially for proteins as ingredients; however, the amino acid profile is essential for evaluation of the nutritional value of proteins. The year-round protein concentration and amino acid....... Aspartic and glutamic acids dominated the amino acid profile, accounting for up to 49 % of the total. Greatest seasonal differences in amino acid composition occurred in July, with leucine contributing most (22.7–26.7 %) of the observed differences. A maximal essential amino acid (EAA) score of 68...

  5. Models of protein and amino acid requirements for cattle

    Directory of Open Access Journals (Sweden)

    Luis Orlindo Tedeschi

    2015-03-01

    Full Text Available Protein supply and requirements by ruminants have been studied for more than a century. These studies led to the accumulation of lots of scientific information about digestion and metabolism of protein by ruminants as well as the characterization of the dietary protein in order to maximize animal performance. During the 1980s and 1990s, when computers became more accessible and powerful, scientists began to conceptualize and develop mathematical nutrition models, and to program them into computers to assist with ration balancing and formulation for domesticated ruminants, specifically dairy and beef cattle. The most commonly known nutrition models developed during this period were the National Research Council (NRC in the United States, Agricultural Research Council (ARC in the United Kingdom, Institut National de la Recherche Agronomique (INRA in France, and the Commonwealth Scientific and Industrial Research Organization (CSIRO in Australia. Others were derivative works from these models with different degrees of modifications in the supply or requirement calculations, and the modeling nature (e.g., static or dynamic, mechanistic, or deterministic. Circa 1990s, most models adopted the metabolizable protein (MP system over the crude protein (CP and digestible CP systems to estimate supply of MP and the factorial system to calculate MP required by the animal. The MP system included two portions of protein (i.e., the rumen-undegraded dietary CP - RUP - and the contributions of microbial CP - MCP as the main sources of MP for the animal. Some models would explicitly account for the impact of dry matter intake (DMI on the MP required for maintenance (MPm; e.g., Cornell Net Carbohydrate and Protein System - CNCPS, the Dutch system - DVE/OEB, while others would simply account for scurf, urinary, metabolic fecal, and endogenous contributions independently of DMI. All models included milk yield and its components in estimating MP required for lactation

  6. A Complex Prime Numerical Representation of Amino Acids for Protein Function Comparison.

    Science.gov (United States)

    Chen, Duo; Wang, Jiasong; Yan, Ming; Bao, Forrest Sheng

    2016-08-01

    Computationally assessing the functional similarity between proteins is an important task of bioinformatics research. It can help molecular biologists transfer knowledge on certain proteins to others and hence reduce the amount of tedious and costly benchwork. Representation of amino acids, the building blocks of proteins, plays an important role in achieving this goal. Compared with symbolic representation, representing amino acids numerically can expand our ability to analyze proteins, including comparing the functional similarity of them. Among the state-of-the-art methods, electro-ion interaction pseudopotential (EIIP) is widely adopted for the numerical representation of amino acids. However, it could suffer from degeneracy that two different amino acid sequences have the same numerical representation, due to the design of EIIP. In light of this challenge, we propose a complex prime numerical representation (CPNR) of amino acids, inspired by the similarity between a pattern among prime numbers and the number of codons of amino acids. To empirically assess the effectiveness of the proposed method, we compare CPNR against EIIP. Experimental results demonstrate that the proposed method CPNR always achieves better performance than EIIP. We also develop a framework to combine the advantages of CPNR and EIIP, which enables us to improve the performance and study the unique characteristics of different representations.

  7. Search for conserved amino acid residues of the α-crystallin proteins of vertebrates.

    Science.gov (United States)

    Shiliaev, Nikita G; Selivanova, Olga M; Galzitskaya, Oxana V

    2016-04-01

    [Formula: see text]-crystallin is the major eye lens protein and a member of the small heat-shock protein (sHsp) family. [Formula: see text]-crystallins have been shown to support lens clarity by preventing the aggregation of lens proteins. We performed the bioinformatics analysis of [Formula: see text]-crystallin sequences from vertebrates to find conserved amino acid residues as the three-dimensional (3D) structure of [Formula: see text]-crystallin is not identified yet. We are the first who demonstrated that the N-terminal region is conservative along with the central domain for vertebrate organisms. We have found that there is correlation between the conserved and structured regions. Moreover, amyloidogenic regions also correspond to the structured regions. We analyzed the amino acid composition of [Formula: see text]-crystallin A and B chains. Analyzing the occurrence of each individual amino acid residue, we have found that such amino acid residues as leucine, serine, lysine, proline, phenylalanine, histidine, isoleucine, glutamic acid, and valine change their content simultaneously in A and B chains in different classes of vertebrates. Aromatic amino acids occur more often in [Formula: see text]-crystallins from vertebrates than on the average in proteins among 17 animal proteomes. We obtained that the identity between A and B chains in the mammalian group is 0.35, which is lower than the published 0.60.

  8. Lactic acid induces aberrant amyloid precursor protein processing by promoting its interaction with endoplasmic reticulum chaperone proteins.

    Directory of Open Access Journals (Sweden)

    Yiwen Xiang

    Full Text Available BACKGROUND: Lactic acid, a natural by-product of glycolysis, is produced at excess levels in response to impaired mitochondrial function, high-energy demand, and low oxygen availability. The enzyme involved in the production of β-amyloid peptide (Aβ of Alzheimer's disease, BACE1, functions optimally at lower pH, which led us to investigate a potential role of lactic acid in the processing of amyloid precursor protein (APP. METHODOLOGY/PRINCIPAL FINDINGS: Lactic acid increased levels of Aβ40 and 42, as measured by ELISA, in culture medium of human neuroblastoma cells (SH-SY5Y, whereas it decreased APP metabolites, such as sAPPα. In cell lysates, APP levels were increased and APP was found to interact with ER-chaperones in a perinuclear region, as determined by co-immunoprecipitation and fluorescence microscopy studies. Lactic acid had only a very modest effect on cellular pH, did increase the levels of ER chaperones Grp78 and Grp94 and led to APP aggregate formation reminiscent of aggresomes. CONCLUSIONS/SIGNIFICANCE: These findings suggest that sustained elevations in lactic acid levels could be a risk factor in amyloidogenesis related to Alzheimer's disease through enhanced APP interaction with ER chaperone proteins and aberrant APP processing leading to increased generation of amyloid peptides and APP aggregates.

  9. Effect of maternal micronutrients (folic acid, vitamin B12) and omega 3 fatty acids on liver fatty acid desaturases and transport proteins in Wistar rats.

    Science.gov (United States)

    Wadhwani, Nisha S; Manglekar, Rupali R; Dangat, Kamini D; Kulkarni, Asmita V; Joshi, Sadhana R

    2012-01-01

    A disturbed fatty acid metabolism increases the risk of adult non-communicable diseases. This study examines the effect of maternal micronutrients on the fatty acid composition, desaturase activity, mRNA levels of fatty acid desaturases and transport proteins in the liver. Pregnant female rats were divided into 6 groups at 2 levels of folic acid both in the presence and absence of vitamin B(12). The vitamin B(12) deficient groups were supplemented with omega 3 fatty acid. An imbalance of maternal micronutrients reduces liver docosahexaenoic acid, increases Δ5 desaturase activity but decreases mRNA levels, decreases Δ6 desaturase activity but not mRNA levels as compared to control. mRNA level of Δ5 desaturase reverts back to the levels of the control group as a result of omega 3 fatty acid supplementation. Our data for the first time indicates that maternal micronutrients differentially alter the activity and expression of fatty acid desaturases in the liver.

  10. Dihedral angle preferences of DNA and RNA binding amino acid residues in proteins.

    Science.gov (United States)

    Ponnuraj, Karthe; Saravanan, Konda Mani

    2017-04-01

    A protein can interact with DNA or RNA molecules to perform various cellular processes. Identifying or analyzing DNA/RNA binding site amino acid residues is important to understand molecular recognition process. It is quite possible to accurately model DNA/RNA binding amino acid residues in experimental protein-DNA/RNA complex by using the electron density map whereas, locating/modeling the binding site amino acid residues in the predicted three dimensional structures of DNA/RNA binding proteins is still a difficult task. Considering the above facts, in the present work, we have carried out a comprehensive analysis of dihedral angle preferences of DNA and RNA binding site amino acid residues by using a classical Ramachandran map. We have computed backbone dihedral angles of non-DNA/RNA binding residues and used as control dataset to make a comparative study. The dihedral angle preference of DNA and RNA binding site residues of twenty amino acid type is presented. Our analysis clearly revealed that the dihedral angles (φ, ψ) of DNA/RNA binding amino acid residues prefer to occupy (-89° to -60°, -59° to -30°) bins. The results presented in this paper will help to model/locate DNA/RNA binding amino acid residues with better accuracy. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Amino acid starvation has opposite effects on mitochondrial and cytosolic protein synthesis.

    Directory of Open Access Journals (Sweden)

    Mark A Johnson

    Full Text Available Amino acids are essential for cell growth and proliferation for they can serve as precursors of protein synthesis, be remodelled for nucleotide and fat biosynthesis, or be burnt as fuel. Mitochondria are energy producing organelles that additionally play a central role in amino acid homeostasis. One might expect mitochondrial metabolism to be geared towards the production and preservation of amino acids when cells are deprived of an exogenous supply. On the contrary, we find that human cells respond to amino acid starvation by upregulating the amino acid-consuming processes of respiration, protein synthesis, and amino acid catabolism in the mitochondria. The increased utilization of these nutrients in the organelle is not driven primarily by energy demand, as it occurs when glucose is plentiful. Instead it is proposed that the changes in the mitochondrial metabolism complement the repression of cytosolic protein synthesis to restrict cell growth and proliferation when amino acids are limiting. Therefore, stimulating mitochondrial function might offer a means of inhibiting nutrient-demanding anabolism that drives cellular proliferation.

  12. Amino acid composition, score and in vitro protein digestibility of foods commonly consumed in Norhwest Mexico

    Directory of Open Access Journals (Sweden)

    Graciela Caire-Juvera

    2013-04-01

    Full Text Available A better knowledge of the amino acid composition of foods commonly consumed in different regions is essential to calculate their scores and, therefore, to predict their protein quality. This paper presents the amino acid composition, amino acid score and in vitro protein digestibility of fifteen foods that are commonly consumed in Northwest Mexico. The foods were prepared by the traditional methods and were analyzed by reverse-phase HPLC. The chemical score for each food was determined using the recommendations for children of 1-2 years of age, and the digestibility was evaluated using a multienzyme technique. Lysine was the limiting amino acid in cereal-based products (scores 15 to 54, and methionine and cysteine were limiting in legume products (scores 41 to 47, boiled beef (score = 75 and hamburger (score = 82. The method of preparation had an effect on the content of certain amino acids, some of them increased and others decreased their content. Meat products and regional cheese provided a high amino acid score (scores 67 to 91 and digestibility (80.7 to 87.8%. Bologna, a processed meat product, had a lower digestibility (75.4%. Data on the amino acid composition of foods commonly consumed in Mexico can be used to provide valuable information on food analysis and protein quality, and to contribute to nutrition and health research and health programs.

  13. Nucleic acid-mediated intracellular protein delivery by lipid-like nanoparticles.

    Science.gov (United States)

    Eltoukhy, Ahmed A; Chen, Delai; Veiseh, Omid; Pelet, Jeisa M; Yin, Hao; Dong, Yizhou; Anderson, Daniel G

    2014-08-01

    Intracellular protein delivery has potential biotechnological and therapeutic application, but remains technically challenging. In contrast, a plethora of nucleic acid carriers have been developed, with lipid-based nanoparticles (LNPs) among the most clinically advanced reagents for oligonucleotide delivery. Here, we validate the hypothesis that oligonucleotides can serve as packaging materials to facilitate protein entrapment within and intracellular delivery by LNPs. Using two distinct model proteins, horseradish peroxidase and NeutrAvidin, we demonstrate that LNPs can yield efficient intracellular protein delivery in vitro when one or more oligonucleotides have been conjugated to the protein cargo. Moreover, in experiments with NeutrAvidin in vivo, we show that oligonucleotide conjugation significantly enhances LNP-mediated protein uptake within various spleen cell populations, suggesting that this approach may be particularly suitable for improved delivery of protein-based vaccines to antigen-presenting cells. Copyright © 2014 Elsevier Ltd. All rights reserved.

  14. Heat-stable proteins and abscisic acid action in barley aleurone cells

    Energy Technology Data Exchange (ETDEWEB)

    Jacobsen, J.V. (CSIRO, Canberra (Australia)); Shaw, D.C. (Australian National Univ., Canberra (Australia))

    1989-12-01

    ({sup 35}S)Methionine labeling experiments showed that abscisic acid (ABA) induced the synthesis of at least 25 polypeptides in mature barley (Hordeum vulgare) aleurone cells. The polypeptides were not secreted. Whereas most of the proteins extracted from aleurone cells were coagulated by heating to 100{degree}C for 10 minutes, most of the ABA-induced polypeptides remained in solution (heat-stable). ABA had little effect on the spectrum of polypeptides that were synthesized and secreted by aleurone cells, and most of these secreted polypeptides were also heat-stable. Coomassie blue staining of sodium dodecyl sulfate polyacrylamide gels indicated that ABA-induced polypeptides already occurred in high amounts in mature aleurone layers having accumulated during grain development. About 60% of the total protein extracted from mature aleurone was heat stable. Amino acid analyses of total preparations of heat-stable and heat-labile proteins showed that, compared to heat-labile proteins, heat-stable intracellular proteins were characterized by higher glutamic acid/glutamine (Glx) and glycine levels and lower levels of neutral amino acids. Secreted heat-stable proteins were rich in Glx and proline. The possibilities that the accumulation of the heat-stable polypeptides during grain development is controlled by ABA and that the function of these polypeptides is related to their abundance and extraordinary heat stability are considered.

  15. Effect of protein/essential amino acids and resistance training on skeletal muscle hypertrophy: A case for whey protein

    Directory of Open Access Journals (Sweden)

    Stout Jeffrey R

    2010-06-01

    Full Text Available Abstract Regardless of age or gender, resistance training or provision of adequate amounts of dietary protein (PRO or essential amino acids (EAA can increase muscle protein synthesis (MPS in healthy adults. Combined PRO or EAA ingestion proximal to resistance training, however, can augment the post-exercise MPS response and has been shown to elicit a greater anabolic effect than exercise plus carbohydrate. Unfortunately, chronic/adaptive response data comparing the effects of different protein sources is limited. A growing body of evidence does, however, suggest that dairy PRO, and whey in particular may: 1 stimulate the greatest rise in MPS, 2 result in greater muscle cross-sectional area when combined with chronic resistance training, and 3 at least in younger individuals, enhance exercise recovery. Therefore, this review will focus on whey protein supplementation and its effects on skeletal muscle mass when combined with heavy resistance training.

  16. The 73 kilodalton heat shock cognate protein purified from rat brain contains nonesterified palmitic and stearic acids.

    Science.gov (United States)

    Guidon, P T; Hightower, L E

    1986-08-01

    A protein related to the 71 kilodalton inducible rat heat shock protein was purified to electrophoretic homogeneity in milligram amounts from brain tissue of nonheat-stressed rats. The protein has been designated as a stress cognate protein based on previous studies and data presented herein that this protein cross-reacted with a monoclonal antibody originally raised against the Drosophila 70 kilodalton heat shock protein. The purified protein had an apparent molecular mass of 73 kilodaltons when analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis and an apparent mass of 150 kilodaltons as determined by nondissociative gel chromatography, suggesting that the purified protein is a homodimer. The purified protein had isoelectric points of 5.0 under nondissociative conditions and 5.6 when exposed to protein denaturants, suggesting loss of bound anionic molecules and/or net exposure of basic residues upon denaturation. Chloroform/methanol extraction of the purified protein and subsequent analyses by thin layer and gas-liquid chromatography resulted in the identification of palmitic and stearic acids noncovalently bound to the protein. Approximately four molecules of fatty acids were bound per dimer with palmitic and stearic acids present in a one-to-one ratio. The purified protein did not bind exogenously added radioactive palmitate, indicating that the fatty acid-binding sites of the cognate protein were fully occupied and that the associated fatty acids were too tightly bound to exchange readily. The possible significance of the fatty acids associated with the 73 kilodalton stress cognate protein is discussed.

  17. Augmentation of protein-derived acetic acid production by heat-alkaline-induced changes in protein structure and conformation.

    Science.gov (United States)

    Wang, Xu; Li, Yanbo; Liu, Junxin; Ren, Nan-Qi; Qu, Jiuhui

    2016-01-01

    Waste-derived acetic acid (HAc) is an attractive feedstock for microbe-mediated biofuel production. However, fermentative conversion of HAc from waste-activated sludge (WAS) has low yield because of the high concentration of proteins not readily utilizable by microorganisms without prior hydrolysis. We investigated a combined technology for HAc augmentation during sludge protein fermentation. The maximal HAc yield increased over two-fold, reaching 0.502 ± 0.021 g/g protein (0.36 ± 0.01 g COD/g COD, ∼52% of the total volatile fatty acids) when synthetic sludge protein was heated at 120 °C for 30 min, treated at pH 12 for 24 h, and fermented at pH 9 for 72 h. Comprehensive analysis illustrated that the heat-alkaline pretreatment significantly induced protein fragmentation, simultaneously increasing the efficiency of protein biohydrolysis (from 35.5% to 85.9%) by inducing conformational changes indicative of protein unfolding. Consequently, the native α-helix content was decreased from 67.3% to 32.5% by conversion to an unordered shape, whose content increased from 27.5% to 45.5%; disulfide bonds were cleaved, whereas the main S-S stretching pattern was altered from gauche-gauche-gauche to gauche-gauche-trans, consequently causing increased protein susceptibility to proteolytic hydrolysis (76.3% vs. 47.0%). Economic analysis indicated that anaerobic fermentation with appropriate heat-alkaline pretreatment is a cost-effective approach for waste conversion to energy sources such as HAc.

  18. Effect of Aacupuncture at Yanglingquan(GB 34) on the Expression of Tyrosine Hydroxylase and Glialfibfillary Acidic Protein in Mice with Parkinson's Disease%针刺筋会穴阳陵泉对帕金森模型小鼠黑质TH和GFAP表达的影响

    Institute of Scientific and Technical Information of China (English)

    陈伶利; 李杰; 李新华; 陈北阳; 莫莉

    2011-01-01

    目的 观察比较1-甲基-4-苯基-1,2,3,6四氢吡啶(MPTP) 诱导的帕金森小鼠模型及其接受针刺治疗后黑质酪氨酸羟化酶(TH)和星形胶质细胞的胶质原纤维性蛋白(GFAP)的表达变化.方法 以腹腔注射(30 mg/kg) MPTP诱导7 d形成帕金森小鼠模型,针刺双侧筋会穴"阳陵泉"及"舞蹈震颤区",每日1次共治疗21 d.针刺结束后,采用免疫荧光组化检测小鼠脑黑质TH的和GFAP的表达变化.结果 在7 d造模后,模型组和针刺组TH阳性细胞显著丢失;治疗结束后,与模型组比较,针刺组TH 阳性细胞数量增加,正常组和针刺组GFAP的阳性细胞数量减少.结论 MPTP可促进帕金森模型小鼠表达;针刺筋会穴阳陵泉可对MPTP 诱导小鼠黑质多巴胺能神经有保护作用,能明显地减弱MPTP伤害性刺激引起的行为反应.%Objective To observe the effect of acupuncture at Yanglingquan (GB 34) on the expression of nigral neurons tyrosine hydroxylase(TH) and glialfibfillary acidic protein (GFAP) in mice model with Parkinson's disease (PD). Methods The mouse model with PD was induced by peritoneal injection of 1- Methyl- 4- phenyl- 1,2,3,6- tetrahydropyridine(MPTP,30 mg/kg) for 7 successive days and divided into acupuncture group acupunctured at Yanglingquan (GB 34) and choreatic section once per day for 12 days and control group. After acupuncture,the expression of mice nigral nurons TH and GFAP were measured through immunofluorescence histochemistry. Results The TH positive neurons in substantial nigra of mice in control group and acupuncture group were lessened distinctly after the 7 day. The expression of TH in the acupuncture group was higher than that in the control group (P<0. 05).The expressions of GFAP in the acupuncture group and the control group were shorter than that in the model group after the 21 day (P<0. 05). Conclusion The acupuncture at Yanglingquan (GB 34) showed a protective effect on dopaminergic neurons of mouse with Parkitson

  19. Stability and structure of protein-lipoamino acid colloidal particles: toward nasal delivery of pharmaceutically active proteins.

    Science.gov (United States)

    Bijani, Christian; Arnarez, Clément; Brasselet, Sabrina; Degert, Corinne; Broussaud, Olivier; Elezgaray, Juan; Dufourc, Erick J

    2012-04-03

    To circumvent the painful intravenous injection of proteins in the treatment of children with growth deficiency, anemia, and calcium insufficiency, we investigated the stability and structure of protein-lipoamino acid complexes that could be nasally sprayed. Preparations that ensure a colloidal and structural stability of recombinant human growth hormone (rhGH), recombinant human erythropoietin (rhEPO), and salmon calcitonin (sCT) mixed with lauroyl proline (LP) were established. Protein structure was controlled by circular dichroism, and very small sizes of ca. 5 nm were determined by dynamic light scattering. The colloidal preparations could be sprayed with a droplet size of 20-30 μm. The molecular structure of aggregates was investigated by all-atom molecular dynamics. Whereas a lauroyl proline capping of globular proteins rhGH and rhEPO with preservation of their active structure was observed, a mixed micelle of sCT and lipoamino acids was formed. In the latter, aggregated LP constitutes the inner core and the surface is covered with calcitonins that acquire a marked α-helix character. Hydrophobic/philic interaction balance between proteins and LP drives the particles' stability. Passage through nasal cells grown at confluence was markedly increased by the colloidal preparations and could reach a 20 times increase in the case of EPO. Biological implications of such colloidal preparations are discussed in terms of furtiveness.

  20. Effect of increased protein intake on renal acid load and renal hemodynamic responses.

    Science.gov (United States)

    Teunissen-Beekman, Karianna F M; Dopheide, Janneke; Geleijnse, Johanna M; Bakker, Stephan J L; Brink, Elizabeth J; de Leeuw, Peter W; van Baak, Marleen A

    2016-03-01

    Increased protein intake versus maltodextrin intake for 4 weeks lowers blood pressure. Concerns exist that high-protein diets reduce renal function. Effects of acute and 4-week protein intake versus maltodextrin intake on renal acid load, glomerular filtration rate and related parameters were compared in this study. Seventy-nine overweight individuals with untreated elevated blood pressure and normal kidney function were randomized to consume a mix of protein isolates (60 g/day) or maltodextrin (60 g/day) for 4 weeks in energy balance. Twenty-four-hour urinary potential renal acid load (uPRAL) was compared between groups. A subgroup (maltodextrin N = 27, protein mix N = 25) participated in extra test days investigating fasting levels and postprandial effects of meals supplemented with a moderate protein- or maltodextrin-load on glomerular filtration rate, effective renal plasma flow, plasma renin, aldosterone, pH, and bicarbonate. uPRAL was significantly higher in the protein group after 4 weeks (P ≤ 0.001). Postprandial filtration fraction decreased further after the protein-supplemented breakfast than after the maltodextrin-supplemented breakfast after 4 weeks of supplementation (P ≤ 0.001). Fasting and postprandial levels of glomerular filtration rate, effective renal plasma flow, renin, aldosterone, angiotensin-converting enzyme, pH and bicarbonate did not differ between groups. In conclusion, 4 weeks on an increased protein diet (25% of energy intake) increased renal acid load, but did not affect renal function. Postprandial changes, except for filtration fraction, also did not differ between groups. These data suggest that a moderate increase in protein intake by consumption of a protein mix for 4 weeks causes no (undesirable) effects on kidney function in overweight and obese individuals with normal kidney function.

  1. Effects of plant proteins on postprandial, free plasma amino acid concentrations in rainbow trout (Oncorhynchus mykiss)

    DEFF Research Database (Denmark)

    Larsen, Bodil Katrine; Dalsgaard, Anne Johanne Tang; Pedersen, Per Bovbjerg

    2012-01-01

    proteins from wheat, peas, field beans, sunflower and soybean. Blood samples were obtained from the caudal vein of 7 fish in each dietary treatment group prior to feeding, as well as: 2, 4, 6, 8, 12, 24, 48 and 72 h after feeding (sampling 7 new fish at each time point), and plasma amino acid......Postprandial patterns in plasma free amino acid concentrations were investigated in juvenile rainbow trout (Oncorhynchus mykiss) fed either a fish meal based diet (FM) or a diet (VEG) where 59% of fish meal protein (corresponding to 46% of total dietary protein) was replaced by a matrix of plant...... concentrations were subsequently measured by HPLC. Nutrient digestibility and ammonia excretion of the two experimental diets were measured in a parallel experiment using a modified Guelph setup. Results showed that the appearance of most amino acids (essential and non-essential) in the plasma was delayed...

  2. Murine protein H is comprised of 20 repeating units, 61 amino acids in length

    DEFF Research Database (Denmark)

    Kristensen, Torsten; Tack, B F

    1986-01-01

    A cDNA library constructed from size-selected (greater than 28 S) poly(A)+ RNA isolated from the livers of C57B10. WR mice was screened by using a 249-base-pair (bp) cDNA fragment encoding 83 amino acid residues of human protein H as a probe. Of 120,000 transformants screened, 30 hybridized......, 448 bp of 3'-untranslated sequence, and a polyadenylylated tail of undetermined length. Murine pre-protein H was deduced to consist of an 18-amino acid signal peptide and 1216 residues of H-protein sequence. Murine H was composed of 20 repetitive units, each about 61 amino acid residues in length...

  3. Computational simulations of protein folding to engineer amino acid sequences to encourage desired supersecondary structure formation.

    Science.gov (United States)

    Gerstman, Bernard S; Chapagain, Prem P

    2013-01-01

    The dynamics of protein folding are complicated because of the various types of amino acid interactions that create secondary, supersecondary, and tertiary interactions. Computational modeling can be used to simulate the biophysical and biochemical interactions that determine protein folding. Effective folding to a desired protein configuration requires a compromise between speed, stability, and specificity. If the primary sequence of amino acids emphasizes one of these characteristics, the others might suffer and the folding process may not be optimized. We provide an example of a model peptide whose primary sequence produces a highly stable supersecondary two-helix bundle structure, but at the expense of lower speed and specificity of the folding process. We show how computational simulations can be used to discover the configuration of the kinetic trap that causes the degradation in the speed and specificity of folding. We also show how amino acid sequences can be engineered by specific substitutions to optimize the folding to the desired supersecondary structure.

  4. Primary structures of three highly acidic ribosomal proteins S6, S12 and S15 from the archaebacterium Halobacterium marismortui.

    Science.gov (United States)

    Kimura, J; Arndt, E; Kimura, M

    1987-11-16

    The amino acid sequences of three extremely acidic ribosomal proteins, S6, S12, and S15, from Halobacterium marismortui have been determined. The sequences were obtained by the sequence analysis of peptides derived by enzymatic digestion with trypsin. Stapylococcus aureus protease and chymotrypsin, as well as by cleavage with dilute HCl. The proteins, S6, S12 and S15, consist of 116, 147 and 102 amino acid residues, and have molecular masses of 12,251, 16,440 and 11,747 Da, respectively. Comparison of the amino acid sequences of these proteins with ribosomal protein sequences of other organisms revealed that halobacterial protein S12 has homology with the eukaryotic protein S16A from Saccharomyces cerevisiae, while S15 is significantly related to the Xenopus laevis S19 protein. No homology was found between these halobacterial proteins and any eubacterial ribosomal proteins.

  5. Binding of acylated peptides and fatty acids to phospholipid vesicles: pertinence to myristoylated proteins.

    Science.gov (United States)

    Peitzsch, R M; McLaughlin, S

    1993-10-01

    We studied the binding of fatty acids and acylated peptides to phospholipid vesicles by making electrophoretic mobility and equilibrium dialysis measurements. The binding energies of the anionic form of the fatty acids and the corresponding acylated glycines were identical; the energies increased by 0.8 kcal/mol per number of carbons in the acyl chain (Ncarbon = 10, 12, 14, 16), a value identical to that for the classical entropy-driven hydrophobic effect discussed by Tanford [The Hydrophobic Effect (1980) Wiley, New York]. The unitary Gibbs free binding energy, delta Gou, of myristoylated glycine, 8 kcal/mol, is independent of the nature of the electrically neutral lipids used to form the vesicles. Similar binding energies were obtained with other myristoylated peptides (e.g., Gly-Ala, Gly-Ala-Ala). The 8 kcal/mol, which corresponds to an effective dissociation constant of 10(-4) M for myristoylated peptides with lipids, provides barely enough energy to attach a myristoylated protein in the cytoplasm to the plasma membrane. Thus, other factors that reduce (e.g., hydrophobic interaction of myristate with the covalently attached protein) or enhance (e.g., electrostatic interactions of basic residues with acidic lipids; protein-protein interactions with intrinsic receptor proteins) the interaction of myristoylated proteins with membranes are likely to be important and may cause reversible translocation of these proteins to the membrane.(ABSTRACT TRUNCATED AT 250 WORDS)

  6. Microbial production of amino acid-modified spider dragline silk protein with intensively improved mechanical properties.

    Science.gov (United States)

    Zhang, Haibo; Zhou, Fengli; Jiang, Xinglin; Cao, Mingle; Wang, Shilu; Zou, Huibin; Cao, Yujin; Xian, Mo; Liu, Huizhou

    2016-08-17

    Spider dragline silk is a remarkably strong fiber with impressive mechanical properties, which were thought to result from the specific structures of the underlying proteins and their molecular size. In this study, silk protein 11R26 from the dragline silk protein of Nephila clavipes was used to analyze the potential effects of the special amino acids on the function of 11R26. Three protein derivatives, ZF4, ZF5, and ZF6, were obtained by site-directed mutagenesis, based on the sequence of 11R26, and among these derivatives, serine was replaced with cysteine, isoleucine, and arginine, respectively. After these were expressed and purified, the mechanical performance of the fibers derived from the four proteins was tested. Both hardness and average elastic modulus of ZF4 fiber increased 2.2 times compared with those of 11R26. The number of disulfide bonds in ZF4 protein was 4.67 times that of 11R26, which implied that disulfide bonds outside the poly-Ala region affect the mechanical properties of spider silk more efficiently. The results indicated that the mechanical performances of spider silk proteins with small molecular size can be enhanced by modification of the amino acids residues. Our research not only has shown the feasibility of large-scale production of spider silk proteins but also provides valuable information for protein rational design.

  7. Improved proteomic analysis following trichloroacetic acid extraction of Bacillus anthracis spore proteins.

    Science.gov (United States)

    Deatherage Kaiser, Brooke L; Wunschel, David S; Sydor, Michael A; Warner, Marvin G; Wahl, Karen L; Hutchison, Janine R

    2015-11-01

    Proteomic analysis of bacterial samples provides valuable information about cellular responses and functions under different environmental pressures. Analysis of cellular proteins is dependent upon efficient extraction from bacterial samples, which can be challenging with increasing complexity and refractory characteristics. While no single method can recover 100% of the bacterial proteins, selected protocols can improve overall protein isolation, peptide recovery, or enrichment for certain classes of proteins. The method presented here is technically simple, does not require specialized equipment such as a mechanical disrupter, and is effective for protein extraction of the particularly challenging sample type of Bacillus anthracis Sterne spores. The ability of Trichloroacetic acid (TCA) extraction to isolate proteins from spores and enrich for spore-specific proteins was compared to the traditional mechanical disruption method of bead beating. TCA extraction improved the total average number of proteins identified within a sample as compared to bead beating (547 vs 495, respectively). Further, TCA extraction enriched for 270 spore proteins, including those typically identified by first isolating the spore coat and exosporium layers. Bead beating enriched for 156 spore proteins more typically identified from whole spore proteome analyses. The total average number of proteins identified was equal using TCA or bead beating for easily lysed samples, such as B. anthracis vegetative cells. As with all assays, supplemental methods such as implementation of an alternative preparation method may simplify sample preparation and provide additional insight to the protein biology of the organism being studied.

  8. Analysis of soybean root proteins affected by gibberellic acid treatment under flooding stress.

    Science.gov (United States)

    Oh, Myeong Won; Nanjo, Yohei; Komatsu, Setsuko

    2014-01-01

    Flooding is a serious abiotic stress for soybean because it restricts growth and reduces grain yields. To investigate the effect of gibberellic acid (GA) on soybean under flooding stress, root proteins were analyzed using a gel-free proteomic technique. Proteins were extracted from the roots of 4-days-old soybean seedlings exposed to flooding stress in the presence and absence of exogenous GA3 for 2 days. A total of 307, 324, and 250 proteins were identified from untreated, and flooding-treated soybean seedlings without or with GA3, respectively. Secondary metabolism- and cell-related proteins, and proteins involved in protein degradation/synthesis were decreased by flooding stress; however, the levels of these proteins were restored by GA3 supplementation under flooding. Fermentation- and cell wall-related proteins were not affected by GA3 supplementation. Furthermore, putative GA-responsive proteins, which were identified by the presence of a GA-responsive element in the promoter region, were less abundant by flooding stress; however, these proteins were more abundant by GA3 supplementation under flooding. Taken together, these results suggest that GA3 affects the abundance of proteins involved in secondary metabolism, cell cycle, and protein degradation/synthesis in soybeans under flooding stress.

  9. Nitrogen effects on proteins, chlorophylls and fatty acids during the growth of Arthrospira platensis.

    Science.gov (United States)

    Ayachi, Samah; El Abed, Amor; Dhifi, Wissal; Marzouk, Brahim

    2007-06-01

    Spirulina platensis (=Arthrospira platensis) is a tunisian strain which has been isolated for the first time in Oued Essed (Sousse, Sidi Bou Ali). Biomass evolution, proteins, chlorophylls and fatty acids composition of this alga were monitored by varying nitrogen concentrations in the culture medium. Nitrogen stress was provoked by adding sodium nitrate (NaNO3) in the culture medium with concentrations varying from 0 to 5 g/l. Results obtained showed that nitrogen depletion increased total proteins and total chlorophylls. The addition of NaNO3 (5g/l) led to an increase of total fatty acids amounts and modify fatty acids composition. Optimal quantities of palmitic, gamma -linolenic and oleic acids were obtained with NaNO3 free-cultures. Thus, the tunisian strain has valuable biological substances, worthy to determine the optimal conditions for its propagation.

  10. Phenylboronic acid-salicylhydroxamic acid bioconjugates. 2. Polyvalent immobilization of protein ligands for affinity chromatography.

    Science.gov (United States)

    Wiley, J P; Hughes, K A; Kaiser, R J; Kesicki, E A; Lund, K P; Stolowitz, M L

    2001-01-01

    Phenylboronic acid bioconjugates prepared from alkaline phosphatase by reaction with either 2,5-dioxopyrrolidinyl 3-[N-[3-(1,3,2-dioxaboran-2-yl)phenyl]carbamoyl]propanoate (PBA-XX-NHS) or 2,5-dioxopyrrolidinyl 6-[[3,5-di-(1,3,2-dioxaboran-2-yl)phenyl]carbonylamino]hexanoate (PDBA-X-NHS) were compared with respect to the efficiency with which they were immobilized on salicylhydroxamic acid-modified Sepharose (SHA-X-Sepharose) by boronic acid complex formation. When immobilized on moderate capacity SHA-X-Sepharose (5.4 micromol of SHA/mL of gel), PDBA-alkaline phosphatase conjugates were shown to be stable with respect to both the alkaline (pH 11.0) and acidic (pH 2.5) buffers utilized to recover anti-alkaline phosphatase during affinity chromatography. Boronic acid complex formation was compared to covalent immobilization of alkaline phosphatase on Affi-Gel 10 and Affi-Gel 15. PDBA-AP.SHA-X-Sepharose was shown to afford superior performance to both Affi-Gel 10 and Affi-Gel 15 with respect to immobilization of alkaline phosphatase, retention of anti-alkaline phosphatase and recovery of anti-alkaline phosphatase under alkaline conditions. High capacity SHA-X-Sepharose (> or = 7 micromol of SHA/mL of gel) was shown to afford superior performance to moderate capacity SHA-X-Sepharose (4.5 micromol of SHA/mL of gel) with respect to stability at pH 11.0 and pH 2.5 when a PDBA-alphaHuman IgG conjugate with a low incorporation ratio of only 1.5:1 was immobilized on SHA-X-Sepharose and subsequently utilized for affinity chromatography of Human IgG. The results are interpreted in terms of either a bivalent or trivalent interaction involving boronic acid complex formation.

  11. Fatty acid binding protein 7 and n-3 poly unsaturated fatty acid supply in early rat brain development.

    Science.gov (United States)

    Maximin, Elise; Langelier, Bénédicte; Aïoun, Josiane; Al-Gubory, Kaïs H; Bordat, Christian; Lavialle, Monique; Heberden, Christine

    2016-03-01

    Fatty acid binding protein 7 (FABP7), abundant in the embryonic brain, binds with the highest affinity to docosahexaenoic acid (DHA) and is expressed in the early stages of embryogenesis. Here, we have examined the consequences of the exposure to different DHA levels and of the in utero depletion of FABP7 on early rat brain development. Neurodevelopment was evaluated through the contents of two proteins, connexin 43 (Cx43) and cyclin-dependent kinase 5 (CDK5), both involved in neuroblast proliferation, differentiation, and migration. The dams were fed with diets presenting different DHA contents, from deficiency to supplementation. DHA brain embryos contents already differed at embryonic day 11.5 and the differences kept increasing with time. Cx43 and CDK5 contents were positively associated with the brain DHA levels. When FABP7 was depleted in vivo by injections of siRNA in the telencephalon, the enhancement of the contents of both proteins was lost in supplemented animals, but FABP7 depletion did not modify phospholipid compositions regardless of the diets. Thus, FABP7 is a necessary mediator of the effect of DHA on these proteins synthesis, but its role in DHA uptake is not critical, although FABP7 is localized in phospholipid-rich areas. Our study shows that high contents of DHA associated with FABP7 are necessary to promote early brain development, which prompted us to recommend DHA supplementation early in pregnancy.

  12. Tolerance to acetic acid is improved by mutations of the TATA-binding protein gene.

    Science.gov (United States)

    An, Jieun; Kwon, Hyeji; Kim, Eunjung; Lee, Young Mi; Ko, Hyeok Jin; Park, Hongjae; Choi, In-Geol; Kim, Sooah; Kim, Kyoung Heon; Kim, Wankee; Choi, Wonja

    2015-03-01

    Screening a library of overexpressing mutant alleles of the TATA-binding gene SPT15 yielded two Saccharomyces cerevisiae strains (MRRC 3252 and 3253) with enhanced tolerance to acetic acid. They were also tolerant to propionic acid and hydrogen peroxide. Transcriptome profile analysis identified 58 upregulated genes and 106 downregulated genes in MRRC 3252. Stress- and protein synthesis-related transcription factors were predominantly enriched in the upregulated and downregulated genes respectively. Eight deletion mutants for some of the highly downregulated genes were acetic acid-tolerant. The level of intracellular reactive oxygen species was considerably lessened in MRRC 3252 and 3253 upon exposure to acetic acid. Metabolome profile analysis revealed that intracellular concentrations of 5 and 102 metabolites were increased and decreased, respectively, in MRRC 3252, featuring a large increase of urea and a significant decrease of amino acids. The dur1/2Δmutant, in which the urea degradation gene DUR1/2 is deleted, displayed enhanced tolerance to acetic acid. Enhanced tolerance to acetic acid was also observed on the medium containing a low concentration of amino acids. Taken together, this study identified two SPT15 alleles, nine gene deletions and low concentration of amino acids in the medium that confer enhanced tolerance to acetic acid.

  13. Radiolytic Modification of Sulfur Containing Acidic Amino Residues in Model Peptides: Fundamental Studies for Protein Footprinting

    Energy Technology Data Exchange (ETDEWEB)

    Xu,G.; Chance, M.

    2005-01-01

    Protein footprinting based on hydroxyl radical-mediated modification and quantitative mass spectroscopic analysis is a proven technique for examining protein structure, protein-ligand interactions, and structural allostery upon protein complex formation. The reactive and solvent-accessible amino acid side chains function as structural probes; however, correct structural analysis depends on the identification and quantification of all the relevant oxidative modifications within the protein sequence. Sulfur-containing amino acids are oxidized readily and the mechanisms of oxidation are particularly complex, although they have been extensively investigated by EPR and other spectroscopic methods. Here we have undertaken a detailed mass spectrometry study (using electrospray ionization mass spectrometry and tandem mass spectrometry) of model peptides containing cysteine (Cys-SH), cystine (disulfide bonded Cys), and methionine after oxidation using {gamma}-rays or synchrotron X-rays and have compared these results to those expected from oxidation mechanisms proposed in the literature. Radiolysis of cysteine leads to cysteine sulfonic acid (+48 Da mass shift) and cystine as the major products; other minor products including cysteine sulfinic acid (+32 Da mass shift) and serine (-16 Da mass shift) are observed. Radiolysis of cystine results in the oxidative opening of the disulfide bond and generation of cysteine sulfonic acid and sulfinic acid; however, the rate of oxidation is significantly less than that for cysteine. Radiolysis of methionine gives rise primarily to methionine sulfoxide (+16 Da mass shift); this can be further oxidized to methionine sulfone (+32 Da mass shift) or another product with a -32 Da mass shift likely due to aldehyde formation at the {gamma}-carbon. Due to the high reactivity of sulfur-containing amino acids, the extent of oxidation is easily influenced by secondary oxidation events or the presence of redox reagents used in standard proteolytic

  14. Recognizing protein–protein interfaces with empirical potentials and reduced amino acid alphabets

    Directory of Open Access Journals (Sweden)

    Wodak Shoshana

    2007-07-01

    Full Text Available Abstract Background In structural genomics, an important goal is the detection and classification of protein–protein interactions, given the structures of the interacting partners. We have developed empirical energy functions to identify native structures of protein–protein complexes among sets of decoy structures. To understand the role of amino acid diversity, we parameterized a series of functions, using a hierarchy of amino acid alphabets of increasing complexity, with 2, 3, 4, 6, and 20 amino acid groups. Compared to previous work, we used the simplest possible functional form, with residue–residue interactions and a stepwise distance-dependence. We used increased computational ressources, however, constructing 290,000 decoys for 219 protein–protein complexes, with a realistic docking protocol where the protein partners are flexible and interact through a molecular mechanics energy function. The energy parameters were optimized to correctly assign as many native complexes as possible. To resolve the multiple minimum problem in parameter space, over 64000 starting parameter guesses were tried for each energy function. The optimized functions were tested by cross validation on subsets of our native and decoy structures, by blind tests on series of native and decoy structures available on the Web, and on models for 13 complexes submitted to the CAPRI structure prediction experiment. Results Performance is similar to several other statistical potentials of the same complexity. For example, the CAPRI target structure is correctly ranked ahead of 90% of its decoys in 6 cases out of 13. The hierarchy of amino acid alphabets leads to a coherent hierarchy of energy functions, with qualitatively similar parameters for similar amino acid types at all levels. Most remarkably, the performance with six amino acid classes is equivalent to that of the most detailed, 20-class energy function. Conclusion This suggests that six carefully chosen amino

  15. Biochemical Roles for Conserved Residues in the Bacterial Fatty Acid-binding Protein Family.

    Science.gov (United States)

    Broussard, Tyler C; Miller, Darcie J; Jackson, Pamela; Nourse, Amanda; White, Stephen W; Rock, Charles O

    2016-03-18

    Fatty acid kinase (Fak) is a ubiquitous Gram-positive bacterial enzyme consisting of an ATP-binding protein (FakA) that phosphorylates the fatty acid bound to FakB. In Staphylococcus aureus, Fak is a global regulator of virulence factor transcription and is essential for the activation of exogenous fatty acids for incorporation into phospholipids. The 1.2-Å x-ray structure of S. aureus FakB2, activity assays, solution studies, site-directed mutagenesis, and in vivo complementation were used to define the functions of the five conserved residues that define the FakB protein family (Pfam02645). The fatty acid tail is buried within the protein, and the exposed carboxyl group is bound by a Ser-93-fatty acid carboxyl-Thr-61-His-266 hydrogen bond network. The guanidinium of the invariant Arg-170 is positioned to potentially interact with a bound acylphosphate. The reduced thermal denaturation temperatures of the T61A, S93A, and H266A FakB2 mutants illustrate the importance of the hydrogen bond network in protein stability. The FakB2 T61A, S93A, and H266A mutants are 1000-fold less active in the Fak assay, and the R170A mutant is completely inactive. All FakB2 mutants form FakA(FakB2)2 complexes except FakB2(R202A), which is deficient in FakA binding. Allelic replacement shows that strains expressing FakB2 mutants are defective in fatty acid incorporation into phospholipids and virulence gene transcription. These conserved residues are likely to perform the same critical functions in all bacterial fatty acid-binding proteins.

  16. Evaluation of salivary sialic acid, total protein, and total sugar in oral cancer: A preliminary report

    Directory of Open Access Journals (Sweden)

    Sanjay P

    2008-01-01

    Full Text Available Aim: Detection of cancer at the early stage is of utmost importance to decrease the morbidity and mortality of the disease. Apart from the conventional biopsy, noninvasive methods like analysis of saliva may provide a cost-effective approach for screening a large population. Thus, this study aimed to estimate salivary levels of sialic acid, total protein, and total sugar in the oral cancer patients and in healthy control group to evaluate their role in diagnosis and prognosis of oral cancer. Study Design: Unstimulated whole saliva samples were collected from 30 healthy controls (Group I and 30 squamous cell carcinoma patients (group II. Estimations of salivary levels of sialic acid, total protein, and total sugar were performed. This was correlated histopathologically with the grades of carcinoma. Statistical Analysis and Results: The Student′s ′ t ′ test and multivariate regression analysis were performed. The results showed that salivary levels of total protein, total sugar, protein-bound sialic acid, and free sialic acid were significantly higher in oral cancer patients compared to those of normal healthy controls ( P values in all the results were less than 0.001. The salivary free sialic acid levels were found to be significantly higher in well-differentiated squamous cell carcinoma than in moderately differentiated carcinoma ( P < 0.001. However, protein-bound sialic acid, total proteins, and total sugars did not show any statistical significance between well and moderately differentiated carcinomas. Conclusion: Biochemical analysis of saliva can be used in early detection of cancer and is best correlated with histopathological degree of squamous cell carcinoma.

  17. Release of free amino acids upon oxidation of peptides and proteins by hydroxyl radicals.

    Science.gov (United States)

    Liu, Fobang; Lai, Senchao; Tong, Haijie; Lakey, Pascale S J; Shiraiwa, Manabu; Weller, Michael G; Pöschl, Ulrich; Kampf, Christopher J

    2017-03-01

    Hydroxyl radical-induced oxidation of proteins and peptides can lead to the cleavage of the peptide, leading to a release of fragments. Here, we used high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) and pre-column online ortho-phthalaldehyde (OPA) derivatization-based amino acid analysis by HPLC with diode array detection and fluorescence detection to identify and quantify free amino acids released upon oxidation of proteins and peptides by hydroxyl radicals. Bovine serum albumin (BSA), ovalbumin (OVA) as model proteins, and synthetic tripeptides (comprised of varying compositions of the amino acids Gly, Ala, Ser, and Met) were used for reactions with hydroxyl radicals, which were generated by the Fenton reaction of iron ions and hydrogen peroxide. The molar yields of free glycine, aspartic acid, asparagine, and alanine per peptide or protein varied between 4 and 55%. For protein oxidation reactions, the molar yields of Gly (∼32-55% for BSA, ∼10-21% for OVA) were substantially higher than those for the other identified amino acids (∼5-12% for BSA, ∼4-6% for OVA). Upon oxidation of tripeptides with Gly in C-terminal, mid-chain, or N-terminal positions, Gly was preferentially released when it was located at the C-terminal site. Overall, we observe evidence for a site-selective formation of free amino acids in the OH radical-induced oxidation of peptides and proteins, which may be due to a reaction pathway involving nitrogen-centered radicals.

  18. Antibacterial drug treatment increases intestinal bile acid absorption via elevated levels of ileal apical sodium-dependent bile acid transporter but not organic solute transporter α protein.

    Science.gov (United States)

    Miyata, Masaaki; Hayashi, Kenjiro; Yamakawa, Hiroki; Yamazoe, Yasushi; Yoshinari, Kouichi

    2015-01-01

    Antibacterial drug treatment increases the bile acid pool size and hepatic bile acid concentration through the elevation of hepatic bile acid synthesis. However, the involvement of intestinal bile acid absorption in the increased bile acid pool size remains unclear. To determine whether intestinal bile acid absorption contributes to the increased bile acid pool in mice treated with antibacterial drugs, we evaluated the levels of bile acid transporter proteins and the capacity of intestinal bile acid absorption. Ileal apical sodium-dependent bile acid transporter (ASBT) mRNA and protein levels were significantly increased in ampicillin (ABPC)-treated mice, whereas organic solute transporter α (OSTα) mRNA levels, but not protein levels, significantly decreased in mice. Similar alterations in the expression levels of bile acid transporters were observed in mice treated with bacitracin/neomycin/streptomycin. The capacity for intestinal bile acid absorption was evaluated by an in situ loop method. Increased ileal absorption of taurochenodeoxycholic acid was observed in mice treated with ABPC. These results suggest that intestinal bile acid absorption is elevated in an ASBT-dependent manner in mice treated with antibacterial drugs.

  19. Relationship between serum adiopocyte fatty acid binding protein and atherosclerosis in chronic kidney disease

    Institute of Scientific and Technical Information of China (English)

    吴晶

    2014-01-01

    Objective To investigate the expression of serum adiopocyte fatty acid binding protein(A-FABP)in chronic kidney disease(CKD)and the role that A-FABP plays in CKD with atherosclerosis.Methods A total of 138 patients with CKD and 20 health control volunteers(HC)were involved in this study.The levels of serum AFABP,free fatty acid(FFA),interleukin-6(IL-6),

  20. Hepatic phenotype of liver fatty acid binding protein gene-ablated mice

    OpenAIRE

    Martin, Gregory G.; Atshaves, Barbara P.; Huang, Huan; McIntosh, Avery L.; Williams, Brad J.; Pai, Pei-Jing; Russell, David H.; Kier, Ann B.; Schroeder, Friedhelm

    2009-01-01

    Although the function of liver fatty acid binding protein in hepatic fatty acid metabolism has been extensively studied, its potential role in hepatic cholesterol homeostasis is less clear. Although hepatic cholesterol accumulation was initially reported in L-FABP-null female mice, that study was performed with early N2 backcross generation mice. To resolve whether the hepatic cholesterol phenotype in these L-FABP−/− mice was attributable to genetic inhomogeneity, these L-FABP−/− mice were fu...

  1. Fish protein hydrolysate elevates plasma bile acids and reduces visceral adipose tissue mass in rats

    DEFF Research Database (Denmark)

    Liaset, Bjørn; Madsen, Lise; Hao, Qin

    2009-01-01

    Conjugation of bile acids (BAs) to the amino acids taurine or glycine increases their solubility and promotes liver BA secretion. Supplementing diets with taurine or glycine modulates BA metabolism and enhances fecal BA excretion in rats. However, it is still unclear whether dietary proteins...... varying in taurine and glycine contents alter BA metabolism, and thereby modulate the recently discovered systemic effects of BAs. Here we show that rats fed a diet containing saithe fish protein hydrolysate (saithe FPH), rich in taurine and glycine, for 26 days had markedly elevated fasting plasma BA...

  2. In vivo synthesized 34S enriched amino acid standards for species specific isotope dilution of proteins

    DEFF Research Database (Denmark)

    Hermann, Gerrit; Moller, Laura Hyrup; Gammelgaard, Bente

    2016-01-01

    A generic quantification approach was introduced addressing the characterization of protein standards while fulfilling the principles of metrology. Traceable absolute quantification was achieved combining a proven biochemical method, i.e. protein hydrolysis followed by amino acid quantification...... in yeast fermentations provided species specific isotopically enriched standards for IDA quantification of cysteine and methionine in the oxidized forms, methionine sulfone and cysteic acid. Reverse isotope dilution mass spectrometry (IDMS) characterization by inductively coupled plasma mass spectrometry...... and methionine sulfone, respectively, was assessed. The established IDA method was validated for the absolute quantification of commercially available lysozyme and ceruloplasmin standards including the calculation of a total combined uncertainty budget....

  3. Incorporation of Amino Acids with Long-Chain Terminal Olefins into Proteins

    OpenAIRE

    Exner, Matthias P.; Sebastian Köhling; Julie Rivollier; Sandrine Gosling; Puneet Srivastava; Palyancheva, Zheni I; Piet Herdewijn; Marie-Pierre Heck; Jörg Rademann; Nediljko Budisa

    2016-01-01

    The increasing need for site-specific protein decorations that mimic natural posttranslational modifications requires access to a variety of noncanonical amino acids with moieties enabling bioorthogonal conjugation chemistry. Here we present the incorporation of long-chain olefinic amino acids into model proteins with rational variants of pyrrolysyl-tRNA synthetase (PylRS). Nε-heptenoyl lysine was incorporated for the first time using the known promiscuous variant PylRS(Y306A/Y384F), and Nε-p...

  4. A Panel of Autoantibodies Against Neural Proteins as Peripheral Biomarker for Pesticide-Induced Neurotoxicity.

    Science.gov (United States)

    El Rahman, Heba Allah Abd; Salama, Mohamed; Gad El-Hak, Seham A; El-Harouny, Mona A; ElKafrawy, Passent; Abou-Donia, Mohamed B

    2017-09-05

    In the present study, we screened the sera of subjects chronically exposed to mixtures of pesticides (composed mainly of organophosphorus compounds (OPs) and others) and developed neurological symptoms for the presence of autoantibodies against cytoskeletal neural proteins. OPs have a well-characterized clinical profile resulting from acute cholinergic crisis. However, some of these compounds cause neuronal degeneration and demyelination known as organophosphorus compound-induced delayed neurotoxicity (OPIDN) and/or organophosphorus compound-induced chronic neurotoxicity (OPICN). Studies from our group have demonstrated the presence of autoantibodies to essential neuronal and glial proteins against cytoskeletal neural proteins in patients with chemical-induced brain injury. In this study, we screened the serum of 50 pesticide-exposed subjects and 25 non-exposed controls, using Western blot analysis against the following proteins: neurofilament triplet proteins (NFPs), tubulin, microtubule-associated tau proteins (Tau), microtubule-associated protein-2 (MAP-2), myelin basic protein (MBP), myelin-associated glycoprotein (MAG), glial fibrillary acidic protein (GFAP), calcium-calmodulin kinase II (CaMKII), glial S100-B protein, and alpha-synuclein (SNCA). Serum reactivity was measured as arbitrary chemiluminescence units. As a group, exposed subjects had significantly higher levels of autoantibody reactivity in all cases examined. The folds of increase in of autoantibodies against neural proteins of the subjects compared to healthy humans in descending order were as follows: MBP, 7.67, MAG 5.89, CaMKII 5.50, GFAP 5.1, TAU 4.96, MAP2 4.83, SNCA 4.55, NFP 4.55, S-100B 2.43, and tubulin 1.78. This study has demonstrated the presence of serum autoantibodies to central nervous system-specific proteins in a group of farmers chronically exposed to pesticides who developed neurological signs and symptoms of neural injury. These autoantibodies can be used as future diagnostic

  5. 玻璃体腔移植人脐带间充质干细胞对糖尿病大鼠视网膜形态及胶质细胞原纤维酸性蛋白和视紫红质表达的影响%The influence of human umbilical cord mesenchymal stem cells transplantation into vitreous cavity of diabetic rats on retinal morphology and the expression level of glial fibrillary acidic protein and rhodopsin

    Institute of Scientific and Technical Information of China (English)

    邹媛媛; 汪建涛; 李筱荣

    2016-01-01

    Objective To observe the influence of human umbilical cord mesenchymal stem cells (hUCMSC) transplantation into vitreous cavity of diabetic rats on the retinal morphology,and the expression of glial fibrillary acidic protein (GFAP) and rhodopsin (RHO).Methods 78 male SpragueDawley rats were used.70 rats were injected with streptozotocin by tail vein injection at a dose of 40 mg/kg to establish the diabetes mellitus model,and another 8 rats were injected with 0.1 mol/L pH 4.0 citric acid buffer at the same dose as the normal control group.After 5 weeks of modeling,10 rats were taken as the control group of diabetic model.hUCMSC suspension was injected into the right eye vitreous cavity of the remaining 60 rats,and the same volume of Dulbecco's modified Eagle/F12 medium was injected into the left vitreous cavity as control eyes.1,2 and 4 weeks after transplantation,follow-up experiments were performed.The experimental eyes were labeled as U1,U2,and U4 groups,while the control eyes were recorded as D1,D2,D4,and each group consisted of 20 eyes.After paraffin section and hematoxylin-eosin staining,the structure of the retina was observed by optical microscopy and the thickness of the outer nuclear layer and the inner nuclear layer (INL) were measured.The distribution and migration of hUCMSC in rat retina were observed by frozen section-tissue immunofluorescence assay.The mRNA and protein expression of GFAP and RHO in the retina were detected by real-time quantitative polymerase chain reaction (PCR) and Western blot assays.Results The results of optical microscope observation showed the normal structure of retina in normal control group.The retinal nerve fiber layer (NFL) was thinned and the number of retinal ganglion cells (RGC) in the control group of diabetic rats was decreased.The decreased number and disorder arrangement of RGC were observed as well in U1,D1 rats.The RGC number of U2,U4,D2,D4 rats was gradually decreased.Compared with D4 group,the thickness of INL

  6. The human fatty acid-binding protein family: Evolutionary divergences and functions

    Directory of Open Access Journals (Sweden)

    Smathers Rebecca L

    2011-03-01

    Full Text Available Abstract Fatty acid-binding proteins (FABPs are members of the intracellular lipid-binding protein (iLBP family and are involved in reversibly binding intracellular hydrophobic ligands and trafficking them throughout cellular compartments, including the peroxisomes, mitochondria, endoplasmic reticulum and nucleus. FABPs are small, structurally conserved cytosolic proteins consisting of a water-filled, interior-binding pocket surrounded by ten anti-parallel beta sheets, forming a beta barrel. At the superior surface, two alpha-helices cap the pocket and are thought to regulate binding. FABPs have broad specificity, including the ability to bind long-chain (C16-C20 fatty acids, eicosanoids, bile salts and peroxisome proliferators. FABPs demonstrate strong evolutionary conservation and are present in a spectrum of species including Drosophila melanogaster, Caenorhabditis elegans, mouse and human. The human genome consists of nine putatively functional protein-coding FABP genes. The most recently identified family member, FABP12, has been less studied.

  7. The Prevalence of STIV c92-Like Proteins in Acidic Thermal Environments

    Directory of Open Access Journals (Sweden)

    Jamie C. Snyder

    2011-01-01

    Full Text Available A new type of viral-induced lysis system has recently been discovered for two unrelated archaeal viruses, STIV and SIRV2. Prior to the lysis of the infected host cell, unique pyramid-like lysis structures are formed on the cell surface by the protrusion of the underlying cell membrane through the overlying external S-layer. It is through these pyramid structures that assembled virions are released during lysis. The STIV viral protein c92 is responsible for the formation of these lysis structures. We searched for c92-like proteins in viral sequences present in multiple viral and cellular metagenomic libraries from Yellowstone National Park acidic hot spring environments. Phylogenetic analysis of these proteins demonstrates that, although c92-like proteins are detected in these environments, some are quite divergent and may represent new viral families. We hypothesize that this new viral lysis system is common within diverse archaeal viral populations found within acidic hot springs.

  8. Protein location prediction using atomic composition and global features of the amino acid sequence

    Energy Technology Data Exchange (ETDEWEB)

    Cherian, Betsy Sheena, E-mail: betsy.skb@gmail.com [Centre for Bioinformatics, University of Kerala, Kariyavattom Campus, Thiruvananthapuram, Kerala (India); Nair, Achuthsankar S. [Centre for Bioinformatics, University of Kerala, Kariyavattom Campus, Thiruvananthapuram, Kerala (India)

    2010-01-22

    Subcellular location of protein is constructive information in determining its function, screening for drug candidates, vaccine design, annotation of gene products and in selecting relevant proteins for further studies. Computational prediction of subcellular localization deals with predicting the location of a protein from its amino acid sequence. For a computational localization prediction method to be more accurate, it should exploit all possible relevant biological features that contribute to the subcellular localization. In this work, we extracted the biological features from the full length protein sequence to incorporate more biological information. A new biological feature, distribution of atomic composition is effectively used with, multiple physiochemical properties, amino acid composition, three part amino acid composition, and sequence similarity for predicting the subcellular location of the protein. Support Vector Machines are designed for four modules and prediction is made by a weighted voting system. Our system makes prediction with an accuracy of 100, 82.47, 88.81 for self-consistency test, jackknife test and independent data test respectively. Our results provide evidence that the prediction based on the biological features derived from the full length amino acid sequence gives better accuracy than those derived from N-terminal alone. Considering the features as a distribution within the entire sequence will bring out underlying property distribution to a greater detail to enhance the prediction accuracy.

  9. Structure of the ordered hydration of amino acids in proteins: analysis of crystal structures

    Energy Technology Data Exchange (ETDEWEB)

    Biedermannová, Lada, E-mail: lada.biedermannova@ibt.cas.cz; Schneider, Bohdan [Institute of Biotechnology CAS, Videnska 1083, 142 20 Prague (Czech Republic)

    2015-10-27

    The hydration of protein crystal structures was studied at the level of individual amino acids. The dependence of the number of water molecules and their preferred spatial localization on various parameters, such as solvent accessibility, secondary structure and side-chain conformation, was determined. Crystallography provides unique information about the arrangement of water molecules near protein surfaces. Using a nonredundant set of 2818 protein crystal structures with a resolution of better than 1.8 Å, the extent and structure of the hydration shell of all 20 standard amino-acid residues were analyzed as function of the residue conformation, secondary structure and solvent accessibility. The results show how hydration depends on the amino-acid conformation and the environment in which it occurs. After conformational clustering of individual residues, the density distribution of water molecules was compiled and the preferred hydration sites were determined as maxima in the pseudo-electron-density representation of water distributions. Many hydration sites interact with both main-chain and side-chain amino-acid atoms, and several occurrences of hydration sites with less canonical contacts, such as carbon–donor hydrogen bonds, OH–π interactions and off-plane interactions with aromatic heteroatoms, are also reported. Information about the location and relative importance of the empirically determined preferred hydration sites in proteins has applications in improving the current methods of hydration-site prediction in molecular replacement, ab initio protein structure prediction and the set-up of molecular-dynamics simulations.

  10. Additive effects of lupin protein and phytic acid on aortic calcification in ApoE deficient mice

    Directory of Open Access Journals (Sweden)

    Alexandra Schutkowski

    2015-03-01

    A two-factorial study with ApoE knockout mice was conducted in which mice received lupin protein isolate or casein with or without phytase. Phytic acid was added to the casein diets to a final concentration identical to the lupin protein diets. Here we show that the serum concentrations of cholesterol, lathosterol and desmosterol were lower and the faecal bile acid excretion was higher in the groups fed lupin proteins than in the groups fed casein (p < 0.05. Mice that received the lupin protein diet containing phytic acid were characterized by a lower aortic calcification than mice of the other three groups (p < 0.05. In conclusion, our results show that the cholesterol lowering properties of lupin protein isolate were not caused by phytic acid. However, the hypocalcific action of lupin proteins appears to depend on the combination of lupin proteins and phytic acid.

  11. Sialic Acid Binding Properties of Soluble Coronavirus Spike (S1 Proteins: Differences between Infectious Bronchitis Virus and Transmissible Gastroenteritis Virus

    Directory of Open Access Journals (Sweden)

    Christine Winter

    2013-07-01

    Full Text Available The spike proteins of a number of coronaviruses are able to bind to sialic acids present on the cell surface. The importance of this sialic acid binding ability during infection is, however, quite different. We compared the spike protein of transmissible gastroenteritis virus (TGEV and the spike protein of infectious bronchitis virus (IBV. Whereas sialic acid is the only receptor determinant known so far for IBV, TGEV requires interaction with its receptor aminopeptidase N to initiate infection of cells. Binding tests with soluble spike proteins carrying an IgG Fc-tag revealed pronounced differences between these two viral proteins. Binding of the IBV spike protein to host cells was in all experiments sialic acid dependent, whereas the soluble TGEV spike showed binding to APN but had no detectable sialic acid binding activity. Our results underline the different ways in which binding to sialoglycoconjugates is mediated by coronavirus spike proteins.

  12. DNA binding protein identification by combining pseudo amino acid composition and profile-based protein representation

    Science.gov (United States)

    Liu, Bin; Wang, Shanyi; Wang, Xiaolong

    2015-10-01

    DNA-binding proteins play an important role in most cellular processes. Therefore, it is necessary to develop an efficient predictor for identifying DNA-binding proteins only based on the sequence information of proteins. The bottleneck for constructing a useful predictor is to find suitable features capturing the characteristics of DNA binding proteins. We applied PseAAC to DNA binding protein identification, and PseAAC was further improved by incorporating the evolutionary information by using profile-based protein representation. Finally, Combined with Support Vector Machines (SVMs), a predictor called iDNAPro-PseAAC was proposed. Experimental results on an updated benchmark dataset showed that iDNAPro-PseAAC outperformed some state-of-the-art approaches, and it can achieve stable performance on an independent dataset. By using an ensemble learning approach to incorporate more negative samples (non-DNA binding proteins) in the training process, the performance of iDNAPro-PseAAC was further improved. The web server of iDNAPro-PseAAC is available at http://bioinformatics.hitsz.edu.cn/iDNAPro-PseAAC/.

  13. Proteomic Analysis of Responsive Proteins Induced in Japanese Birch Plantlet Treated with Salicylic Acid

    Directory of Open Access Journals (Sweden)

    Hiromu Suzuki

    2014-07-01

    Full Text Available The present study was performed to unravel the mechanisms of systemic acquired resistance (SAR establishment and resistance signaling pathways against the canker-rot fungus (Inonotus obliquus strain IO-U1 infection in Japanese birch plantlet No.8. Modulation of protein-profile induced by salicylic acid (SA-administration was analyzed, and SA-responsive proteins were identified. In total, 5 specifically expressed, 3 significantly increased, and 3 significantly decreased protein spots were identified using liquid chromatography/tandem mass spectrometry (LC/MS/MS and the sequence tag method. These proteins were malate dehydrogenase, succinate dehydrogenase, phosphoglycerate kinase, diaminopimalate decarboxylase, arginase, chorismate mutase, cyclophilin, aminopeptidase, and unknown function proteins. These proteins are considered to be involved in SAR-establishment mechanisms in the Japanese birch plantlet No 8.

  14. Probing the structural dynamics of proteins and nucleic acids with optical tweezers.

    Science.gov (United States)

    Ritchie, Dustin B; Woodside, Michael T

    2015-10-01

    Conformational changes are an essential feature of most molecular processes in biology. Optical tweezers have emerged as a powerful tool for probing conformational dynamics at the single-molecule level because of their high resolution and sensitivity, opening new windows on phenomena ranging from folding and ligand binding to enzyme function, molecular machines, and protein aggregation. By measuring conformational changes induced in a molecule by forces applied by optical tweezers, new insight has been gained into the relationship between dynamics and function. We discuss recent advances from studies of how structure forms in proteins and RNA, including non-native structures, fluctuations in disordered proteins, and interactions with chaperones assisting native folding. We also review the development of assays probing the dynamics of complex protein-nucleic acid and protein-protein assemblies that reveal the dynamic interactions between biomolecular machines and their substrates.

  15. Proteomic Analysis of Responsive Proteins Induced in Japanese Birch Plantlet Treated with Salicylic Acid

    Science.gov (United States)

    Suzuki, Hiromu; Takashima, Yuya; Ishiguri, Futoshi; Yoshizawa, Nobuo; Yokota, Shinso

    2014-01-01

    The present study was performed to unravel the mechanisms of systemic acquired resistance (SAR) establishment and resistance signaling pathways against the canker-rot fungus (Inonotus obliquus strain IO-U1) infection in Japanese birch plantlet No.8. Modulation of protein-profile induced by salicylic acid (SA)-administration was analyzed, and SA-responsive proteins were identified. In total, 5 specifically expressed, 3 significantly increased, and 3 significantly decreased protein spots were identified using liquid chromatography/tandem mass spectrometry (LC/MS/MS) and the sequence tag method. These proteins were malate dehydrogenase, succinate dehydrogenase, phosphoglycerate kinase, diaminopimalate decarboxylase, arginase, chorismate mutase, cyclophilin, aminopeptidase, and unknown function proteins. These proteins are considered to be involved in SAR-establishment mechanisms in the Japanese birch plantlet No 8.

  16. Fatty acids induce leukotriene C4 synthesis in macrophages in a fatty acid binding protein-dependent manner.

    Science.gov (United States)

    Long, Eric K a; Hellberg, Kristina; Foncea, Rocio; Hertzel, Ann V; Suttles, Jill; Bernlohr, David A

    2013-07-01

    Obesity results in increased macrophage recruitment to adipose tissue that promotes a chronic low-grade inflammatory state linked to increased fatty acid efflux from adipocytes. Activated macrophages produce a variety of pro-inflammatory lipids such as leukotriene C4 (LTC4) and 5-, 12-, and 15-hydroxyeicosatetraenoic acid (HETE) suggesting the hypothesis that fatty acids may stimulate eicosanoid synthesis. To assess if eicosanoid production increases with obesity, adipose tissue of leptin deficient ob/ob mice was analyzed. In ob/ob mice, LTC4 and 12-HETE levels increased in the visceral (but not subcutaneous) adipose depot while the 5-HETE levels decreased and 15-HETE abundance was unchanged. Since macrophages produce the majority of inflammatory molecules in adipose tissue, treatment of RAW264.7 or primary peritoneal macrophages with free fatty acids led to increased secretion of LTC4 and 5-HETE, but not 12- or 15-HETE. Fatty acid binding proteins (FABPs) facilitate the intracellular trafficking of fatty acids and other hydrophobic ligands and in vitro stabilize the LTC4 precursor leukotriene A4 (LTA4) from non-enzymatic hydrolysis. Consistent with a role for FABPs in LTC4 synthesis, treatment of macrophages with HTS01037, a specific FABP inhibitor, resulted in a marked decrease in both basal and fatty acid-stimulated LTC4 secretion but no change in 5-HETE production or 5-lipoxygenase expression. These results indicate that the products of adipocyte lipolysis may stimulate the 5-lipoxygenase pathway leading to FABP-dependent production of LTC4 and contribute to the insulin resistant state.

  17. A single whey acidic protein domain containing protein (SWD) inhibits bacteria invasion and dissemination in shrimp Marsupenaeus japonicus.

    Science.gov (United States)

    Jiang, Hai-Shan; Sun, Chen; Wang, Tong; Zhao, Xiao-Fan; Wang, Jin-Xing

    2013-08-01

    The single whey acidic protein (WAP) domain containing proteins (SWDs) in crustacean belong to type III crustins and have antiprotease activities and/or antimicrobial activities. Their functions in vivo in crustacean immunity need to be clarify. In this study, a new single WAP domain containing protein (SWD) was obtained from Marsupenaeus japonicus, designated as MjSWD. The full-length cDNA of MjSWD was 522 bp.The open reading frame of MjSWD encoded a protein of 79 amino acids, with a 24 amino acid signal peptide and a WAP domain. Tissue distribution analysis revealed that MjSWD transcripts were generally expressed in all the tested tissues, including hemocytes, heart, hepatopancreas, gill, stomach and intestine. The time course expression of MjSWD was analyzed by quantitative real time PCR, and the results exhibited that MjSWD was upregulated after bacteria (Vibrio anguillarum, Staphylococcus aureus) and white spot syndrome virus (WSSV) challenge in gills and stomach of the shrimp. The purified recombinant protein of MjSWD could bind to several Gram-negative and Gram-positive bacteria though binding to microbial polysaccharides (peptidoglycan). MjSWD could inhibit the activity of Subtilisin A and Proteinase K and bacteria-secreted proteases. The results of natural infection with MjSWD incubated bacteria showed that the inhibition of MjSWD against bacterial secreted proteases was contributed to inhibiting bacteria invasion and dissemination in the shrimp. The MjSWD is, thus, involved in the shrimp antibacterial innate immunity.

  18. Meal composition and plasma amino acid ratios: Effect of various proteins or carbohydrates, and of various protein concentrations

    Science.gov (United States)

    Yokogoshi, Hidehiko; Wurtman, Richard J.

    1986-01-01

    The effects of meals containing various proteins and carbohydrates, and of those containing various proportions of protein (0 percent to 20 percent of a meal, by weight) or of carbohydrate (0 percent to 75 percent), on plasma levels of certain large neutral amino acids (LNAA) in rats previously fasted for 19 hours were examined. Also the plasma tryptophan ratios (the ratio of the plasma trytophan concentration to the summed concentrations of the other large neutral amino acids) and other plasma amino acid ratios were calculated. (The plasma tryptophan ratio has been shown to determine brain tryptophan levels and, thereby, to affect the synthesis and release of the neurotransmitter serotonin). A meal containing 70 percent to 75 percent of an insulin-secreting carbohydrate (dextrose or dextrin) increased plasma insulin levels and the tryptophan ratio; those containing 0 percent or 25 percent carbohydrate failed to do so. Addition of as little as 5 percent casein to a 70 percent carbohydrate meal fully blocked the increase in the plasma tryptophan ratio without affecting the secretion of insulin - probably by contributing much larger quantities of the other LNAA than of tryptophan to the blood. Dietary proteins differed in their ability to suppress the carbohydrate-induced rise in the plasma tryptophan ratio. Addition of 10 percent casein, peanut meal, or gelatin fully blocked this increase, but lactalbumin failed to do so, and egg white did so only partially. (Consumption of the 10 percent gelatin meal also produced a major reduction in the plasma tyrosine ratio, and may thereby have affected brain tyrosine levels and catecholamine synthesis.) These observations suggest that serotonin-releasing neurons in brains of fasted rats are capable of distinguishing (by their metabolic effects) between meals poor in protein but rich in carbohydrates that elicit insulin secretion, and all other meals. The changes in brain serotonin caused by carbohydrate-rich, protein

  19. Meal composition and plasma amino acid ratios: Effect of various proteins or carbohydrates, and of various protein concentrations

    Science.gov (United States)

    Yokogoshi, Hidehiko; Wurtman, Richard J.

    1986-01-01

    The effects of meals containing various proteins and carbohydrates, and of those containing various proportions of protein (0 percent to 20 percent of a meal, by weight) or of carbohydrate (0 percent to 75 percent), on plasma levels of certain large neutral amino acids (LNAA) in rats previously fasted for 19 hours were examined. Also the plasma tryptophan ratios (the ratio of the plasma trytophan concentration to the summed concentrations of the other large neutral amino acids) and other plasma amino acid ratios were calculated. (The plasma tryptophan ratio has been shown to determine brain tryptophan levels and, thereby, to affect the synthesis and release of the neurotransmitter serotonin). A meal containing 70 percent to 75 percent of an insulin-secreting carbohydrate (dextrose or dextrin) increased plasma insulin levels and the tryptophan ratio; those containing 0 percent or 25 percent carbohydrate failed to do so. Addition of as little as 5 percent casein to a 70 percent carbohydrate meal fully blocked the increase in the plasma tryptophan ratio without affecting the secretion of insulin - probably by contributing much larger quantities of the other LNAA than of tryptophan to the blood. Dietary proteins differed in their ability to suppress the carbohydrate-induced rise in the plasma tryptophan ratio. Addition of 10 percent casein, peanut meal, or gelatin fully blocked this increase, but lactalbumin failed to do so, and egg white did so only partially. (Consumption of the 10 percent gelatin meal also produced a major reduction in the plasma tyrosine ratio, and may thereby have affected brain tyrosine levels and catecholamine synthesis.) These observations suggest that serotonin-releasing neurons in brains of fasted rats are capable of distinguishing (by their metabolic effects) between meals poor in protein but rich in carbohydrates that elicit insulin secretion, and all other meals. The changes in brain serotonin caused by carbohydrate-rich, protein

  20. A possible mode of the specifi-crecognition of nucleic acids by proteins

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Seven sets of protein target sites, which occur in several gene promoters, have been analyzed. The results suggest that there is a possible mode of specific recognition of double-helical nucleic acids by proteins. This recognition mode is related to a special topological property of double-helical DNA, which is termed base spatial pattern (BSP) of DNA segment. BSP is the spatial topological property determined only by the spatial arrangement of the bases on double-helical DNA segment.

  1. Effects of Salvianolic Acid B on Protein Expression in Human Umbilical Vein Endothelial Cells

    OpenAIRE

    Tsong-Min Chang; Guey-Yueh Shi; Hua-Lin Wu; Chieh-Hsi Wu; Yan-Di Su; Hui-Lin Wang; Hsin-Yun Wen; Huey-Chun Huang

    2011-01-01

    Salvianolic acid B (Sal B), a pure water-soluble compound extracted from Radix Salviae miltiorrhizae, has been reported to possess potential cardioprotective efficacy. To identify proteins or pathways by which Sal B might exert its protective activities on the cardiovascular system, two-dimensional gel electrophoresis-based comparative proteomics was performed, and proteins altered in their expression level after Sal B treatment were identified by MALDI-TOF MS/MS. Human umbilical vein endothe...

  2. Protein and Amino Acid Restriction, Aging and Disease: from yeast to humans

    OpenAIRE

    Mirzaei, Hamed; Suarez, Jorge A.; Valter D Longo

    2014-01-01

    Many of the effects of dietary restriction (DR) on longevity and health span in model organisms have been linked to reduced protein and amino acid (AA) intake and the stimulation of specific nutrient signaling pathways. Studies in yeast have shown that addition of serine, threonine, and valine in media promotes cellular sensitization and aging by activating different but connected pathways. Protein or essential AA restriction extends both lifespan and healthspan in rodent models. In humans, p...

  3. Diverse roles of the nucleic acid binding protein KHSRP in cell differentiation and disease

    Science.gov (United States)

    Briata, Paola; Bordo, Domenico; Puppo, Margherita; Gorlero, Franco; Rossi, Martina; Bizzozzero, Nora Perrone; Gherzi, Roberto

    2015-01-01

    The single-stranded nucleic acid binding protein KHSRP (KH-Type Splicing Regulatory Protein) modulates RNA life and gene expression at various levels. KHSRP controls important cellular functions as different as proliferation, differentiation, metabolism and response to infectious agents. We summarize and discuss experimental evidence providing a potential link between changes in KHSRP expression/function and human diseases including neuromuscular disorders, obesity, type II diabetes, and cancer. PMID:26708421

  4. Prediction of endoplasmic reticulum resident proteins using fragmented amino acid composition and support vector machine

    Directory of Open Access Journals (Sweden)

    Ravindra Kumar

    2017-09-01

    Full Text Available Background The endoplasmic reticulum plays an important role in many cellular processes, which includes protein synthesis, folding and post-translational processing of newly synthesized proteins. It is also the site for quality control of misfolded proteins and entry point of extracellular proteins to the secretory pathway. Hence at any given point of time, endoplasmic reticulum contains two different cohorts of proteins, (i proteins involved in endoplasmic reticulum-specific function, which reside in the lumen of the endoplasmic reticulum, called as endoplasmic reticulum resident proteins and (ii proteins which are in process of moving to the extracellular space. Thus, endoplasmic reticulum resident proteins must somehow be distinguished from newly synthesized secretory proteins, which pass through the endoplasmic reticulum on their way out of the cell. Approximately only 50% of the proteins used in this study as training data had endoplasmic reticulum retention signal, which shows that these signals are not essentially present in all endoplasmic reticulum resident proteins. This also strongly indicates the role of additional factors in retention of endoplasmic reticulum-specific proteins inside the endoplasmic reticulum. Methods This is a support vector machine based method, where we had used different forms of protein features as inputs for support vector machine to develop the prediction models. During training leave-one-out approach of cross-validation was used. Maximum performance was obtained with a combination of amino acid compositions of different part of proteins. Results In this study, we have reported a novel support vector machine based method for predicting endoplasmic reticulum resident proteins, named as ERPred. During training we achieved a maximum accuracy of 81.42% with leave-one-out approach of cross-validation. When evaluated on independent dataset, ERPred did prediction with sensitivity of 72.31% and specificity of 83

  5. Allspice and Clove As Source of Triterpene Acids Activating the G Protein-Coupled Bile Acid Receptor TGR5

    Directory of Open Access Journals (Sweden)

    Angela Ladurner

    2017-07-01

    Full Text Available Worldwide, metabolic diseases such as obesity and type 2 diabetes have reached epidemic proportions. A major regulator of metabolic processes that gained interest in recent years is the bile acid receptor TGR5 (Takeda G protein-coupled receptor 5. This G protein-coupled membrane receptor can be found predominantly in the intestine, where it is mainly responsible for the secretion of the incretins glucagon-like peptide 1 (GLP-1 and peptide YY (PYY. The aim of this study was (i to identify plant extracts with TGR5-activating potential, (ii to narrow down their activity to the responsible constituents, and (iii to assess whether the intestinal microbiota produces transformed metabolites with a different activity profile. Chenodeoxycholic acid (CDCA served as positive control for both, the applied cell-based luciferase reporter gene assay for TGR5 activity and the biotransformation assay using mouse fecal slurry. The suitability of the workflow was demonstrated by the biotransformation of CDCA to lithocholic acid resulting in a distinct increase in TGR5 activity. Based on a traditional Tibetan formula, 19 plant extracts were selected and investigated for TGR5 activation. Extracts from the commonly used spices Syzygium aromaticum (SaroE, clove, Pimenta dioica (PdioE, allspice, and Kaempferia galanga (KgalE, aromatic ginger significantly increased TGR5 activity. After biotransformation, only KgalE showed significant differences in its metabolite profile, which, however, did not alter its TGR5 activity compared to non-transformed KgalE. UHPLC-HRMS (high-resolution mass spectrometry analysis revealed triterpene acids (TTAs as the main constituents of the extracts SaroE and PdioE. Identification and quantification of TTAs in these two extracts as well as comparison of their TGR5 activity with reconstituted TTA mixtures allowed the attribution of the TGR5 activity to TTAs. EC50s were determined for the main TTAs, i.e., oleanolic acid (2.2 ± 1.6

  6. Allspice and Clove As Source of Triterpene Acids Activating the G Protein-Coupled Bile Acid Receptor TGR5.

    Science.gov (United States)

    Ladurner, Angela; Zehl, Martin; Grienke, Ulrike; Hofstadler, Christoph; Faur, Nadina; Pereira, Fátima C; Berry, David; Dirsch, Verena M; Rollinger, Judith M

    2017-01-01

    Worldwide, metabolic diseases such as obesity and type 2 diabetes have reached epidemic proportions. A major regulator of metabolic processes that gained interest in recent years is the bile acid receptor TGR5 (Takeda G protein-coupled receptor 5). This G protein-coupled membrane receptor can be found predominantly in the intestine, where it is mainly responsible for the secretion of the incretins glucagon-like peptide 1 (GLP-1) and peptide YY (PYY). The aim of this study was (i) to identify plant extracts with TGR5-activating potential, (ii) to narrow down their activity to the responsible constituents, and (iii) to assess whether the intestinal microbiota produces transformed metabolites with a different activity profile. Chenodeoxycholic acid (CDCA) served as positive control for both, the applied cell-based luciferase reporter gene assay for TGR5 activity and the biotransformation assay using mouse fecal slurry. The suitability of the workflow was demonstrated by the biotransformation of CDCA to lithocholic acid resulting in a distinct increase in TGR5 activity. Based on a traditional Tibetan formula, 19 plant extracts were selected and investigated for TGR5 activation. Extracts from the commonly used spices Syzygium aromaticum (SaroE, clove), Pimenta dioica (PdioE, allspice), and Kaempferia galanga (KgalE, aromatic ginger) significantly increased TGR5 activity. After biotransformation, only KgalE showed significant differences in its metabolite profile, which, however, did not alter its TGR5 activity compared to non-transformed KgalE. UHPLC-HRMS (high-resolution mass spectrometry) analysis revealed triterpene acids (TTAs) as the main constituents of the extracts SaroE and PdioE. Identification and quantification of TTAs in these two extracts as well as comparison of their TGR5 activity with reconstituted TTA mixtures allowed the attribution of the TGR5 activity to TTAs. EC50s were determined for the main TTAs, i.e., oleanolic acid (2.2 ± 1.6 μM), ursolic

  7. Molecular basis for amino acid sensing by family C G-protein-coupled receptors

    DEFF Research Database (Denmark)

    Wellendorph, Petrine; Bräuner-Osborne, Hans

    2009-01-01

    -alpha;-amino acid receptor G-protein-coupled receptor family C, group 6, subtype A (GPRC6A) and seven orphan receptors. Aside from the orphan receptors, the family C GPCRs are dimeric receptors characterized by a large extracellular Venus flytrap domain which bind the endogenous agonists. Except from the GABA(B1......-2) and T1R2-3 receptor, all receptors are either activated or positively modulated by amino acids. In this review, we outline mutational, biophysical and structural studies which have elucidated the interaction of the amino acids with the Venus flytrap domains, molecular mechanisms of receptor selectivity...

  8. Preliminary research on amino acid composition and nutritional value of clover proteins

    Directory of Open Access Journals (Sweden)

    L. Kłyszejko-Stefanowicz

    2015-06-01

    Full Text Available The amino acid composition and nutritional value of 5 clover varieties including 3 Polish ones ('Gloria', 'Hruszowska', 'Skrzeszowicka' and 2 of foreign origin ('Rotra' and 'Violetta' were investigated. No significant differences in the total protein content (19.2–20.0% of dry matter as well as in qualitative amino acid composition were found among the clover varieties under examination. EAA index (Essential amino acid index calculated according to Oser for 'Gloria' and 'Hruszowska' showed the highest nutritional value was – 40. The lowest value of EAA index was found for 'Violetta' cvar. – 32, intermediate values however for Rotra and Skrzeszowicka was 37 and 36.

  9. Effects of different fatty acid chain lengths on fatty acid oxidation-related protein expression levels in rat skeletal muscles.

    Science.gov (United States)

    Ishizawa, Rie; Masuda, Kazumi; Sakata, Susumu; Nakatani, Akira

    2015-01-01

    Skeletal muscles can adapt to dietary interventions that affect energy metabolism. Dietary intake of medium-chain fatty acids (MCFAs) enhances mitochondrial oxidation of fatty acids (FAO) in type IIa skeletal muscle fibers. However, the effect of MCFAs diet on mitochondrial or cytoplasmic FAO-related protein expression levels in different types of muscle fibers remains unclear. This study aims to examine the effects of a high-fat diet, containing MCFAs, on mitochondrial enzyme activities and heart-type fatty acid-binding protein (H-FABP) levels in different types of skeletal muscle fibers. Five-week-old male Wistar rats were assigned to one of the following three dietary conditions: standard chow (SC, 12% of calories from fat), high-fat MCFA, or high-fat long-chain fatty acids (LCFAs) diet (60% of calories from fat for both). The animals were provided food and water ad libitum for 4 weeks, following which citrate synthase (CS) activity and H-FABP concentration were analyzed. The epididymal fat pads (EFP) were significantly smaller in the MCFA group than in the LCFA group (p increase in CS activity compared with that observed in SC-fed controls in all types of skeletal muscle fibers (triceps, surface portion of gastrocnemius (gasS), deep portion of gastrocnemius (gasD), and soleus; p increase the expression of the mitochondrial enzyme CS, but not that of H-FABP, in both fast- and slow-twitch muscle fibers, suggesting that H-FABP expression is dependent on the chain length of fatty acids in the cytoplasm of skeletal muscles cells.

  10. Immunohistochemical localization of D-β-aspartic acid-containing proteins in pterygium.

    Science.gov (United States)

    Kaji, Yuichi; Oshika, Tetsuro; Nejima, Ryouhei; Mori, Saiyo; Miyata, Kazunori; Fujii, Noriko

    2015-12-10

    Biologically uncommon D-β-aspartic acid (D-β-Asp) residues have been reported to accumulate in organs affected by age-related disorders. In the present study, we investigated the localization of D-β-Asp-containing proteins in cases of pterygium, one of the most prominent age-related ocular conditions. Immunohistochemical localization of D-β-Asp-containing proteins was investigated in surgical specimens of pterygium from 20 patients and control specimens from 10 patients. Strong immunoreactivity to D-β-Asp-containing proteins was observed in subepithelial elastotic lesions and surrounding collagenous lesions from all surgical specimens with pterygia. In contrast, no immunoreactivity to D-β-Asp-containing proteins was seen in pterygium-free specimens. D-β-Asp-containing proteins are produced in organs as they are affected by the aging process. In addition, conversion of L- to D-aspartyl residues is accelerated by ultraviolet (UV) irradiation. Since pterygia can form due to aging or UV exposure, it is reasonable to find D-β-Asp-containing proteins in specimens with pterygia. Furthermore, since D-β-Asp is a non-native amino acid, D-β-Asp-containing proteins may be recognized as allogeneic antigens. Therefore, D-β-Asp-containing proteins in pterygia may responsible for the fibrovascular changes seen in the disorder.

  11. Studies on the protein and sulfur amino acid requirements of young bobwhite quail

    Science.gov (United States)

    Serafin, J.A.

    1977-01-01

    Four experiments were conducted with purified diets to examine the influence of protein level and to estimate the sulfur amino acid (S.A.A.) requirement of young Bobwhite quail (Colinus virginianus). These studies demonstrated (I) that 26% protein was sufficient for rapid growth when the diet was supplemented with methionine; (2) that diets containing higher levels of protein (29.3% and 31.3%) failed to support satisfactory growth unless they contained supplemental methionine; and (3) that young Bobwhite quail require no more than 1.0% sulfur-containing amino acids for optimal growth and efficiency of feed utilization. A fifth experiment was conducted to examine the protein and S.A.A. requirements of young Bobwhite quail using practical rations and to compare results with those obtained with purified diets. Diets containing 24%, 26% and 28% protein were supplied with and without supplemental methionine in a five week study. Results showed significant growth responses to protein and supplemental methionine. Responses showed that Bobwhite quail require no more than 26% protein for maximum growth and efficiency of feed utilization when the S.A.A. level of the diet was approximately 1.0%. The results were in close agreement with those obtained with purified diets. These findings define more precisely than had been known the quantitative requirements of young Bobwhite quail for protein and for the S.A.A. necessary for optimal growth.

  12. Identification of β-phenylalanine as a non-protein amino acid in cultivated rice, Oryza sativa.

    Science.gov (United States)

    Yokoo, Takayuki; Takata, Ryo; Yan, Jian; Matsumoto, Fuka; Teraishi, Masayoshi; Okumoto, Yutaka; Jander, Georg; Mori, Naoki

    2015-01-01

    Non-protein amino acids, often analogs of the standard 20 protein amino acids, have been discovered in many plant species. Recent research with cultivated rice (Oryza sativa) identified (3R)-β-tyrosine, as well as a tyrosine amino mutase that synthesizes (3R)-β-tyrosine from the protein amino acid (2S)-α-tyrosine. Gas chromatography-mass spectrometry (GC-MS) assays and comparison to an authentic standard showed that β-phenylalanine is also a relatively abundant non-protein amino acid in rice leaves and that its biosynthesis occurs independently from that of β-tyrosine.

  13. The up-regulation of glial fibrillary acidic protein media through P38-mitogen activated protein kinase pathway after spinal cord injury%脊髓损伤后通过P38丝裂素活化蛋白激酶途径上调胶质纤维酸性蛋白的表达

    Institute of Scientific and Technical Information of China (English)

    于春雷; 王翀昊; 张弘; 刘长江; 腾宇飞

    2006-01-01

    目的 探讨脊髓损伤(SCI)后受损的脊髓组织与其周围组织中胶质纤维酸性蛋白(glial fibrillary acidic protein,GFAP)和P38丝裂素活化蛋白激酶(P38-mitogen activated protein kinase,P38MAPK)的表达情况.方法 将80只大鼠随机分成10组:正常对照组、损伤后1 d组、损伤后4 d组、损伤后7 d组、损伤后14 d组、损伤后应用SB203580药物1 d组、4 d组、7 d组、14 d组及健康大鼠给药组,每组均8只.应用westerblot技术检测各组损伤组织及损伤周围组织的P38MAPK及GFAP的表达.结果 受损脊髓组织从伤后第1 d P38MAPK、GFAP开始上升,持续到第7 d开始回落,第7 d是表达最高峰.损伤组织P38MAPK及GFAP的表达具有明显正相关性(r=0.854,P<0.05),损伤周围组织P38MAPK及GFAP的表达无明显相关性(r=0.554,P>0.05).结论 损伤组织GFAP表达上调是通过P38MAPK介导的,损伤周围组织GFAP上调与P38MAPK表达无关.

  14. Polymorphism of the 86th amino acid in CX26 protein and hereditary deafness

    Institute of Scientific and Technical Information of China (English)

    Xi Shi; Mingyang Shi; Yuehua Qiao; Shiwei Qiu; Fendong Yan; Lizhang Shi; Yili Xuan; Wei Zhuang; Yingli Bei; Hanli Yao; Na Yuan

    2016-01-01

    Objective:To investigate the membrane localization function of the CX26 protein when its 86th amino acid is Thr, Ser or Arg, and its relations to deafness. Methods:CX26-GFP prote