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Sample records for betaine-homocysteine s-methyltransferase bhmt

  1. Structure-activity study of new inhibitors of human betaine-homocysteine S-methyltransferase

    Czech Academy of Sciences Publication Activity Database

    Vaněk, Václav; Buděšínský, Miloš; Kabeleová, Petra; Šanda, Miloslav; Kožíšek, Milan; Hančlová, Ivona; Mládková, Jana; Brynda, Jiří; Rosenberg, Ivan; Koutmos, M.; Garrow, T. A.; Jiráček, Jiří

    2009-01-01

    Roč. 52, č. 12 (2009), s. 3652-3665 ISSN 0022-2623 R&D Projects: GA MŠk 1M0508 Grant - others:GA MŠk(CZ) LC06077; NIH(US) R01TW0052501 Program:LC Institutional research plan: CEZ:AV0Z40550506 Keywords : BHMT * betain * homocysteine * methionine * inhibitor Subject RIV: CE - Biochemistry Impact factor: 4.802, year: 2009

  2. Dissecting the Catalytic Mechanism of Betaine-Homocysteine S-Methyltransferase Using Intrinsic Tryptophan Fluorescence and Site-Directed Mutagenesis

    Energy Technology Data Exchange (ETDEWEB)

    Castro, C.; Gratson, A.A.; Evans, J.C.; Jiracek, J.; Collinsova, M.; Ludwig, M.L.; Garrow, T.A. (ASCR); (UIUC); (Michigan)

    2010-03-05

    Betaine-homocysteine S-methyltransferase (BHMT) is a zinc-dependent enzyme that catalyzes the transfer of a methyl group from glycine betaine (Bet) to homocysteine (Hcy) to form dimethylglycine (DMG) and methionine (Met). Previous studies in other laboratories have indicated that catalysis proceeds through the formation of a ternary complex, with a transition state mimicked by the inhibitor S-({delta}-carboxybutyl)-l-homocysteine (CBHcy). Using changes in intrinsic tryptophan fluorescence to determine the affinity of human BHMT for substrates, products, or CBHcy, we now demonstrate that the enzyme-substrate complex reaches its transition state through an ordered bi-bi mechanism in which Hcy is the first substrate to bind and Met is the last product released. Hcy, Met, and CBHcy bind to the enzyme to form binary complexes with K{sub d} values of 7.9, 6.9, and 0.28 {micro}M, respectively. Binary complexes with Bet and DMG cannot be detected with fluorescence as a probe, but Bet and DMG bind tightly to BHMT-Hcy to form ternary complexes with K{sub d} values of 1.1 and 0.73 {micro}M, respectively. Mutation of each of the seven tryptophan residues in human BHMT provides evidence that the enzyme undergoes two distinct conformational changes that are reflected in the fluorescence of the enzyme. The first is induced when Hcy binds, and the second, when Bet binds. As predicted by the crystal structure of BHMT, the amino acids Trp44 and Tyr160 are involved in binding Bet, and Glu159 in binding Hcy. Replacing these residues by site-directed mutagenesis significantly reduces the catalytic efficiency (V{sub max}/K{sub m}) of the enzyme. Replacing Tyr77 with Phe abolishes enzyme activity.

  3. Double-Headed Sulfur-Linked Amino Acids As First Inhibitors for Betaine-Homocysteine S-Methyltransferase 2

    Czech Academy of Sciences Publication Activity Database

    Mládková, Jana; Vaněk, Václav; Buděšínský, Miloš; Elbert, Tomáš; Demianova, Zuzana; Garrow, T. A.; Jiráček, Jiří

    2012-01-01

    Roč. 55, č. 15 (2012), s. 6822-6831 ISSN 0022-2623 R&D Projects: GA ČR GA203/09/1919; GA ČR(CZ) GAP207/10/1277 Institutional support: RVO:61388963 Keywords : betaine * homocysteine * methionine * BHMT * inhibitor Subject RIV: CE - Biochemistry Impact factor: 5.614, year: 2012

  4. Inhibition of betaine-homocysteine S-methyltransferase causes hyperhomocysteinemia in mice

    Czech Academy of Sciences Publication Activity Database

    Collinsová, Michaela; Straková, J.; Jiráček, Jiří; Garrow, T. A.

    2006-01-01

    Roč. 136, č. 6 (2006), s. 1493-1497 ISSN 0022-3166 R&D Projects: GA AV ČR(CZ) IAA4055302 Grant - others:NIH(US) DK52501 Institutional research plan: CEZ:AV0Z40550506 Keywords : betaine * homocystein e * dimethylsulfoniopropionate Subject RIV: CE - Biochemistry Impact factor: 4.009, year: 2006

  5. The development of a new class of inhibitors for betaine-homocysteine S-methyltransferase

    Czech Academy of Sciences Publication Activity Database

    Pícha, Jan; Vaněk, Václav; Buděšínský, Miloš; Mládková, Jana; Garrow, T. A.; Jiráček, Jiří

    2013-01-01

    Roč. 65, July (2013), s. 256-275 ISSN 0223-5234 R&D Projects: GA ČR(CZ) GAP207/10/1277 Institutional support: RVO:61388963 Keywords : BHMT * inhibitor * homocysteine * phosphonate * phosphinate * amino acid derivative * bioisostere * S-alkylated homocysteine Subject RIV: CE - Biochemistry Impact factor: 3.432, year: 2013

  6. S-alkylated homocysteine derivatives: New inhibitors of human betaine-homocysteine S-methyltransferase

    Czech Academy of Sciences Publication Activity Database

    Jiráček, Jiří; Collinsová, Michaela; Rosenberg, Ivan; Buděšínský, Miloš; Protivínská, Eva; Netušilová, Hana; Garrow, T. A.

    2006-01-01

    Roč. 49, č. 13 (2006), s. 3982-3989 ISSN 0022-2623 R&D Projects: GA AV ČR(CZ) IAA4055302 Grant - others:NIH(US) DK52501; NIH(US) R01TW0052501; IARS(US) 50-352 Institutional research plan: CEZ:AV0Z40550506 Keywords : BHMT * S-alkylated homocystein e * inhibitor Subject RIV: CE - Biochemistry Impact factor: 5.115, year: 2006

  7. Specific potassium ion interactions facilitate homocysteine binding to betaine-homocysteine S-methyltransferase

    Czech Academy of Sciences Publication Activity Database

    Mládková, Jana; Hladílková, Jana; Diamond, C. E.; Tryon, K.; Yamada, K.; Garrow, T. A.; Jungwirth, Pavel; Koutmos, M.; Jiráček, Jiří

    2014-01-01

    Roč. 82, č. 10 (2014), s. 2552-2564 ISSN 0887-3585 R&D Projects: GA ČR(CZ) GAP207/10/1277; GA ČR GBP208/12/G016 Institutional support: RVO:61388963 Keywords : BHMT * homocysteine * potassium * crystal structure * molecular dynamics * simulations * enzyme kinetics Subject RIV: CE - Biochemistry Impact factor: 2.627, year: 2014

  8. Downregulation of hepatic betaine:homocysteine methyltransferase (BHMT) expression in taurine-deficient mice is reversed by taurine supplementation in vivo.

    Science.gov (United States)

    Jurkowska, Halina; Niewiadomski, Julie; Hirschberger, Lawrence L; Roman, Heather B; Mazor, Kevin M; Liu, Xiaojing; Locasale, Jason W; Park, Eunkyue; Stipanuk, Martha H

    2016-03-01

    The cysteine dioxygenase (Cdo1)-null and the cysteine sulfinic acid decarboxylase (Csad)-null mouse are not able to synthesize hypotaurine/taurine by the cysteine/cysteine sulfinate pathway and have very low tissue taurine levels. These mice provide excellent models for studying the effects of taurine on biological processes. Using these mouse models, we identified betaine:homocysteine methyltransferase (BHMT) as a protein whose in vivo expression is robustly regulated by taurine. BHMT levels are low in liver of both Cdo1-null and Csad-null mice, but are restored to wild-type levels by dietary taurine supplementation. A lack of BHMT activity was indicated by an increase in the hepatic betaine level. In contrast to observations in liver of Cdo1-null and Csad-null mice, BHMT was not affected by taurine supplementation of primary hepatocytes from these mice. Likewise, CSAD abundance was not affected by taurine supplementation of primary hepatocytes, although it was robustly upregulated in liver of Cdo1-null and Csad-null mice and lowered to wild-type levels by dietary taurine supplementation. The mechanism by which taurine status affects hepatic CSAD and BHMT expression appears to be complex and to require factors outside of hepatocytes. Within the liver, mRNA abundance for both CSAD and BHMT was upregulated in parallel with protein levels, indicating regulation of BHMT and CSAD mRNA synthesis or degradation.

  9. Combining combinatorial chemistry and affinity chromatography: highly selective inhibitors of human betaine:homocysteine S-methyltransferase

    Czech Academy of Sciences Publication Activity Database

    Collinsová, Michaela; Castro, C.; Garrow, T. A.; Yiotakis, A.; Dive, V.; Jiráček, Jiří

    2003-01-01

    Roč. 10, - (2003), s. 113-122 ISSN 1074-5521 R&D Projects: GA AV ČR IAB4055003 Institutional research plan: CEZ:AV0Z4055905 Keywords : BHMT * inhibitor * phosphinic Subject RIV: CE - Biochemistry Impact factor: 6.129, year: 2003

  10. Quantification of homocysteine-related metabolites and the role of betaine-homocysteine S-methyltransferase in HepG2 cells

    Czech Academy of Sciences Publication Activity Database

    Kořínek, M.; Šístek, V.; Mládková, Jana; Mikeš, P.; Jiráček, Jiří; Selicharová, Irena

    2013-01-01

    Roč. 27, č. 1 (2013), s. 111-121 ISSN 0269-3879 R&D Projects: GA ČR(CZ) GAP207/10/1277 Institutional support: RVO:61388963 Keywords : homocysteine * BHMT * LC-MS/MS * HepG2 * metabolites Subject RIV: CE - Biochemistry Impact factor: 1.662, year: 2013

  11. Dissecting the Catalytic Mechanism of Betaine - Homocysteine S-Methyltransferase by Use of Intrinsic Tryptophan Fluorescence and Site-Directed Mutagenesis

    Czech Academy of Sciences Publication Activity Database

    Castro, C.; Gratson, A. A.; Evans, J. C.; Jiráček, Jiří; Collinsová, Michaela; Ludwig, M. L.; Garrow, T. A.

    2004-01-01

    Roč. 43, č. 18 (2004), s. 5341-5351 ISSN 0006-2960 R&D Projects: GA AV ČR IAA4055302 Grant - others:NIH(US) GM16429; NIH(US) DK52501; Illinois Agricultural Experimental Station(US) ILLU-698-352 Institutional research plan: CEZ:AV0Z4055905 Keywords : BHMT * CBHcy * fluorescence Subject RIV: CE - Biochemistry Impact factor: 4.008, year: 2004

  12. Analogy tranzitního stavu jako inhibitory lidského enzymu betain:homocystein S-methyltransferasy

    Czech Academy of Sciences Publication Activity Database

    Netušilová, Hana; Buděšínský, Miloš; Rosenberg, Ivan; Jiráček, Jiří

    2004-01-01

    Roč. 98, č. 5 (2004), s. 295 ISSN 0009-2770. [Mezioborové setkání mladých biologů, biochemiků a chemiků /4./. 09.06.2004-12.06.2004, Žďárské vrchy] R&D Projects: GA ČR GA203/01/1166; GA AV ČR IAA4055302 Keywords : betain: homocystein e S-methyltransferase Subject RIV: CC - Organic Chemistry

  13. Development of first inhibitors for betaine-homocysteine S-methyltransferase 2

    Czech Academy of Sciences Publication Activity Database

    Mládková, J.; Vaněk, Václav; Elbert, Tomáš; Buděšínský, Miloš; Jiráček, Jiří

    2012-01-01

    Roč. 106, - (2012), s894-s894 ISSN 0009-2770. [EuCheMS Chemistry Congress /4./. 26.08.2012-30.08.2012, Prague] Institutional research plan: CEZ:AV0Z40550506 Keywords : enzyme catalysis * inhibitors * alkylation Subject RIV: CC - Organic Chemistry

  14. Development of novel highly selective phosphinic pseudopeptide inhibitors of Zn-metalloenzyme betaine: homocysteine S-methyltransferase

    Czech Academy of Sciences Publication Activity Database

    Collinsová, Michaela

    2002-01-01

    Roč. 96, č. 4 (2002), s. 210-211 ISSN 0009-2770. [Sigma-Aldrich konference mladých chemiků, biochemiků a molekulárních biologů. 22.05.2002-25.05.2002, Velké Meziříčí] Institutional research plan: CEZ:AV0Z4055905 Keywords : phosphinic pseudopeptides Subject RIV: CE - Biochemistry

  15. Effects of hyperhomocysteinemia and betaine-homocysteine S-methyltransferase inhibition on hepatocyte metabolites and the proteome

    Czech Academy of Sciences Publication Activity Database

    Selicharová, Irena; Kořínek, M.; Demianova, Zuzana; Chrudinová, Martina; Mládková, Jana; Jiráček, Jiří

    2013-01-01

    Roč. 1834, č. 8 (2013), s. 1596-1606 ISSN 1570-9639 R&D Projects: GA ČR(CZ) GAP207/10/1277 Institutional support: RVO:61388963 Keywords : apolipoprotein * fibrinogen * one-carbon metabolism * S-Adenosylmethionine * two-dimensional electrophoresis Subject RIV: CE - Biochemistry Impact factor: 3.191, year: 2013

  16. Association between BHMT gene rs3733890 polymorphism and cancer risk: evidence from a meta-analysis

    Directory of Open Access Journals (Sweden)

    Xu Y

    2016-08-01

    Full Text Available Yue Xu,1,* Cunye Yan,2,* Zongyao Hao,1 Jun Zhou,1 Song Fan,1 Sheng Tai,1 Cheng Yang,1 Li Zhang,1 Chaozhao Liang1 1Department of Urology, The First Affiliated Hospital of Anhui Medical University and Institute of Urology, 2First School of Clinical Medicine, Anhui Medical University, Hefei, Anhui, People’s Republic of China *These authors contributed equally to this work Background and objective: The gene betaine-homocysteine methyltransferase (BHMT has drawn much attention during the past decades. An increasing number of clinical and genetic investigations have supposed that BHMT rs3733890 polymorphism might be associated with risk of breast cancer and ovarian cancer. As no consistent conclusion has been achieved, we conducted an up-to-date summary of BHMT rs3733890 polymorphism and cancer risk through a meta-analysis. Materials and methods: The articles were collected from PubMed, Google Scholar, and CNKI (Chinese databases up to December 2015. Then, the correlations were determined by reading the titles and abstracts and by further reading the full text to filter the unqualified articles. Odds ratio (OR and the corresponding 95% confidence intervals (CI were used to assess the results. Results: Among 187 articles collected in the analysis, seven studies with a total of 2,832 cases and 3,958 controls were included for evaluation of the association between BHMT rs3733890 polymorphism and susceptibility of cancer risk. The heterogeneity test showed no significant differences. Furthermore, we found that BHMT –742G>A polymorphism in case and control groups showed no statistically significant association with susceptibility in various cancer types except for uterine cervical cancer (A vs G: OR =0.641, 95% CI =0.445–0.923, P=0.017; AA+AG vs GG: OR =0.579, 95% CI =0.362–0.924, P=0.022. In addition, no statistically significant association was uncovered when stratification analyses were conducted by ethnicity and genotyping methods. Conclusion

  17. Maternal Folate Status and the BHMT c.716G>A Polymorphism Affect the Betaine Dimethylglycine Pathway during Pregnancy

    Directory of Open Access Journals (Sweden)

    Jose M. Colomina

    2016-10-01

    Full Text Available The effect of the betaine: homocysteine methyltransferase BHMT c.716G>A (G: guanosine; A: adenosine single nucleotide polymorphism (SNP on the BHMT pathway is unknown during pregnancy. We hypothesised that it impairs betaine to dimethylglycine conversion and that folate status modifies its effect. We studied 612 women from the Reus Tarragona Birth Cohort from ≤12 gestational weeks (GW throughout pregnancy. The frequency of the variant BHMT c.716A allele was 30.8% (95% confidence interval (CI: 28.3, 33.5. In participants with normal-high plasma folate status (>13.4 nmol/L, least square geometric mean [95% CI] plasma dimethylglycine (pDMG, µmol/L was lower in the GA (2.35 [2.23, 2.47] versus GG (2.58 [2.46, 2.70] genotype at ≤12 GW (p < 0.05 and in the GA (2.08 [1.97, 2.19] and AA (1.94 [1.75, 2.16] versus GG (2.29 [2.18, 2.40] genotypes at 15 GW (p < 0.05. No differences in pDMG between genotypes were observed in participants with possible folate deficiency (≤13.4 nmol/L (p for interactions at ≤12 GW: 0.023 and 15 GW: 0.038. PDMG was lower in participants with the AA versus GG genotype at 34 GW (2.01 [1.79, 2.25] versus 2.44 [2.16, 2.76] and at labour, 2.51 [2.39, 2.64] versus 3.00 [2.84, 3.18], (p < 0.01. Possible deficiency compared to normal-high folate status was associated with higher pDMG in multiple linear regression analysis (β coefficients [SEM] ranging from 0.07 [0.04], p < 0.05 to 0.20 [0.04], p < 0.001 in models from early and mid-late pregnancy and the AA compared to GG genotype was associated with lower pDMG (β coefficients [SEM] ranging from −0.11 [0.06], p = 0.055 to −0.23 [0.06], p < 0.001. Conclusion: During pregnancy, the BHMT pathway is affected by folate status and by the variant BHMT c.716A allele.

  18. Structural Basis of Substrate Recognition in Thiopurine S-Methyltransferase

    Energy Technology Data Exchange (ETDEWEB)

    Peng, Yi; Feng, Qiping; Wilk, Dennis; Adjei, Araba A.; Salavaggione, Oreste E.; Weinshilboum, Richard M.; Yee, Vivien C. (Case Western); (MCCM)

    2008-09-23

    Thiopurine S-methyltransferase (TPMT) modulates the cytotoxic effects of thiopurine prodrugs such as 6-mercaptopurine by methylating them in a reaction using S-adenosyl-l-methionine as the donor. Patients with TPMT variant allozymes exhibit diminished levels of protein and/or enzyme activity and are at risk for thiopurine drug-induced toxicity. We have determined two crystal structures of murine TPMT, as a binary complex with the product S-adenosyl-l-homocysteine and as a ternary complex with S-adenosyl-l-homocysteine and the substrate 6-mercaptopurine, to 1.8 and 2.0 {angstrom} resolution, respectively. Comparison of the structures reveals that an active site loop becomes ordered upon 6-mercaptopurine binding. The positions of the two ligands are consistent with the expected S{sub N}2 reaction mechanism. Arg147 and Arg221, the only polar amino acids near 6-mercaptopurine, are highlighted as possible participants in substrate deprotonation. To probe whether these residues are important for catalysis, point mutants were prepared in the human enzyme. Substitution of Arg152 (Arg147 in murine TPMT) with glutamic acid decreases V{sub max} and increases K{sub m} for 6-mercaptopurine but not K{sub m} for S-adenosyl-l-methionine. Substitution at this position with alanine or histidine and similar substitutions of Arg226 (Arg221 in murine TPMT) result in no effect on enzyme activity. The double mutant Arg152Ala/Arg226Ala exhibits a decreased V{sub max} and increased K{sub m} for 6-mercaptopurine. These observations suggest that either Arg152 or Arg226 may participate in some fashion in the TPMT reaction, with one residue compensating when the other is altered, and that Arg152 may interact with substrate more directly than Arg226, consistent with observations in the murine TPMT crystal structure.

  19. Effects of betaine supplementation and choline deficiency on folate deficiency-induced hyperhomocysteinemia in rats.

    Science.gov (United States)

    Liu, Ying; Liu, Yi-qun; Morita, Tatsuya; Sugiyama, Kimio

    2012-01-01

    The effect of betaine status on folate deficiency-induced hyperhomocysteinemia was investigated to determine whether folate deficiency impairs homocysteine removal not only by the methionine synthase (MS) pathway but also by the betaine-homocysteine S-methyltransferase (BHMT) pathway. For this purpose, we investigated the effect of dietary supplementation with betaine at a high level (1%) in rats fed a folate-deprived 10% casein diet (10C) and 20% casein diet (20C). We also investigated the effect of choline deprivation on folate deficiency-induced hyperhomocysteinemia in rats fed 20C. Supplementation of folate-deprived 10C and 20C with 1% betaine significantly suppressed folate deprivation-induced hyperhomocysteinemia, but the extent of suppression was partial or limited, especially in rats fed 10C, the suppression of plasma homocysteine increment being 48.5% in rats fed 10C and 69.7% in rats fed 20C. Although betaine supplementation greatly increased hepatic betaine concentration and BHMT activity, these increases did not fully explain why the effect of betaine supplementation was partial or limited. Folate deprivation markedly increased the hepatic concentration of N,N-dimethylglycine (DMG), a known inhibitor of BHMT, and there was a significant positive correlation between hepatic DMG concentration and plasma homocysteine concentration, suggesting that folate deficiency increases hepatic DMG concentration and thereby depresses BHMT reaction, leading to interference with the effect of betaine supplementation. Choline deprivation did not increase plasma homocysteine concentration in rats fed 20C, but it markedly enhanced plasma homocysteine concentration when rats were fed folate-deprived 20C. This indicates that choline deprivation reinforced folate deprivation-induced hyperhomocysteinemia. Increased hepatic DMG concentration was also associated with such an effect. These results support the concept that folate deficiency impairs homocysteine metabolism not only

  20. Betaine-homocysteine methyltransferase: zinc in a distorted barrel

    Czech Academy of Sciences Publication Activity Database

    Evans, J. C.; Huddler, D. P.; Jiráček, Jiří; Castro, C.; Millian, N. S.; Garrow, T. A.; Ludwig, M. L.

    2002-01-01

    Roč. 10, - (2002), s. 1159-1171 ISSN 0969-2126 R&D Projects: GA AV ČR IAB4055003 Grant - others:NIH(US) GM16429; NIH(US) DK52501 Institutional research plan: CEZ:AV0Z4055905 Keywords : homocysteine Subject RIV: CE - Biochemistry Impact factor: 6.030, year: 2002

  1. Suppression Effects of Betaine-Enriched Spinach on Hyperhomocysteinemia Induced by Guanidinoacetic Acid and Choline Deficiency in Rats

    Directory of Open Access Journals (Sweden)

    Yi-Qun Liu

    2014-01-01

    Full Text Available Betaine is an important natural component of rich food sources, especially spinach. Rats were fed diets with betaine or spinach powder at the same level of betaine for 10 days to investigate the dose-dependent effects of spinach powder supplementation on hyperhomocysteinemia induced by guanidinoacetic acid (GAA addition and choline deprivation. The GAA-induced hyperhomocysteinemia in rats fed 25% casein diet (25C was significantly suppressed by supplementation with betaine or spinach, and it was completely suppressed by taking 11.0% spinach supplementation. The choline deprivation-induced enhancement of plasma homocysteine concentration in rats fed 25% soybean protein diet (25S was markedly suppressed by 3.82% spinach. Supplementation with betaine or spinach partially prevented the effects of GAA on hepatic concentrations of methionine metabolites. The decrease in activity of betaine-homocysteine S-methyltransferase (BHMT and cystathionine β-synthase (CBS in GAA-induced hyperhomocysteinemia was recovered by supplementation with betaine or spinach. Supplementation with betaine or spinach did not affect BHMT activity, whereas it partially restored CBS activity in choline-deprived 25S. The results indicated that betaine or spinach could completely suppress the hyperhomocysteinemia induced by choline deficiency resulting from stimulating the homocysteine removal by both remethylation and cystathionine formation.

  2. Metazoan Remaining Genes for Essential Amino Acid Biosynthesis: Sequence Conservation and Evolutionary Analyses

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    Igor R. Costa

    2014-12-01

    Full Text Available Essential amino acids (EAA consist of a group of nine amino acids that animals are unable to synthesize via de novo pathways. Recently, it has been found that most metazoans lack the same set of enzymes responsible for the de novo EAA biosynthesis. Here we investigate the sequence conservation and evolution of all the metazoan remaining genes for EAA pathways. Initially, the set of all 49 enzymes responsible for the EAA de novo biosynthesis in yeast was retrieved. These enzymes were used as BLAST queries to search for similar sequences in a database containing 10 complete metazoan genomes. Eight enzymes typically attributed to EAA pathways were found to be ubiquitous in metazoan genomes, suggesting a conserved functional role. In this study, we address the question of how these genes evolved after losing their pathway partners. To do this, we compared metazoan genes with their fungal and plant orthologs. Using phylogenetic analysis with maximum likelihood, we found that acetolactate synthase (ALS and betaine-homocysteine S-methyltransferase (BHMT diverged from the expected Tree of Life (ToL relationships. High sequence conservation in the paraphyletic group Plant-Fungi was identified for these two genes using a newly developed Python algorithm. Selective pressure analysis of ALS and BHMT protein sequences showed higher non-synonymous mutation ratios in comparisons between metazoans/fungi and metazoans/plants, supporting the hypothesis that these two genes have undergone non-ToL evolution in animals.

  3. Paradoxical elevated thiopurine S-methyltransferase activity after pancytopenia during azathioprine therapy: potential influence of red blood cell age

    NARCIS (Netherlands)

    de Boer, Nanne K. H.; van Bodegraven, Adriaan A.; de Graaf, Peer; van der Hulst, Rene W. M.; Zoetekouw, Lida; van Kuilenburg, André B. P.

    2008-01-01

    There is an increased risk of developing bone marrow depression and infections during azathioprine therapy for inflammatory bowel disease. Patients with low or absent thiopurine S-methyltransferase (TPMT) activity have an increased risk of developing myelotoxicity. We describe a patient who

  4. Association between TPMT*3C and decreased thiopurine S-methyltransferase activity in patients with neuromyelitis optica spectrum disorders in China.

    Science.gov (United States)

    Gong, Xiaoqing; Mei, Shenghui; Li, Xindi; Li, Xingang; Zhou, Heng; Liu, Yonghong; Zhou, Anna; Yang, Li; Zhao, Zhigang; Zhang, Xinghu

    2018-06-01

    Thiopurines are effective drugs in treating neuromyelitis optica spectrum disorders and other diseases. Thiopurines' toxicity is mainly imputed to thiopurine S-methyltransferase activity. In Chinese population, the most common and important variation of thiopurine S-methyltransferase is TPMT*3C (rs1142345). This study aims to reveal the association between thiopurine S-methyltransferase activity and genetic polymorphisms of thiopurine S-methyltransferase in patients with neuromyelitis optica spectrum disorders in China. A liquid chromatography tandem mass/mass method was used to evaluate the thiopurine S-methyltransferase activity by using 6-mercapthioprine as the substrate in human erythrocyte haemolysate via 1 h incubation at 37 °C to form its methylated product 6-methylmercaptopurine. The amount of 6-methylmercaptopurine was adjusted by haematocrit and normalized to 8 × 10 8 erythrocytes. The selected polymorphisms of thiopurine S-methyltransferase were identified using MassARRAY system (Sequenom) and multiple SNaPshot technique. In 69 patients with neuromyelitis optica spectrum disorders, thiopurine S-methyltransferase activity was 80.29-154.53 (127.51 ± 16.83) pmol/h/8 × 10 8 erythrocytes. TPMT*3C (rs1142345) was associated with lower thiopurine S-methyltransferase activity (BETA = -25.37, P = 0.011). Other selected variants were not associated with thiopurine S-methyltransferase activity. TPMT*3C affects TPMT activity in Chinese patients with neuromyelitis optica spectrum disorders. Further studies are warranted to confirm the results. TPRs = thiopurines; NMOSD = neuromyelitis optica spectrum disorders; TPMT = thiopurine S-methyltransferase; LC-MS/MS = liquid chromatography tandem mass/mass; 6-MMP = 6-methylmercaptopurine; IS = internal standard; SNP = single nucleotide polymorphism; MAF = minor allele frequency; HWE = Hardy-Weinberg equilibrium; BETA = regression coefficients; UTR-3 = untranslated region 3.

  5. Determination of thiopurine S-methyltransferase activity by hydrophilic interaction liquid chromatography hyphenated with mass spectrometry.

    Science.gov (United States)

    Pecher, Daniel; Dokupilová, Svetlana; Zelinková, Zuzana; Peppelenbosch, Maikel; Mikušová, Veronika; Mikuš, Peter

    2017-08-05

    Thiopurine S-methyltransferase (TPMT) plays an important role in the metabolism of thiopurines used in the therapy of inflammatory bowel diseases (IBD). In this work a new progressive method for the determination of TPMT activity in red blood cells lysates was developed. Analysis was carried out by means of hydrophilic interaction liquid chromatography (HILIC) hyphenated with mass spectrometry (MS). In comparison with reversed-phase high-performance liquid chromatography (RP-HPLC), that has been typically applied in determination of TPMT activity, the HILIC significantly improved the analytical signal provided by MS, shortened analysis time, and improved chromatographic resolution. The HILIC-HPLC-MS method was optimized and validated, providing favorable parameters of detection and quantitation limits (5.5 and 16.5pmol/mL, respectively), linearity (coefficient of determination 0.9999 in the range of 0.01-1.0nmol/mL), recovery and precision (93.25-100.37% with RSD 1.06-1.32% in the whole concentration range of QC samples). Moreover, in contrast to the conventional RP-HPLC-UV approach, the complex phenotype TPMT profiles can be reliably and without interferences monitored using the HILIC-HPLC-MS method. Such advanced monitoring can provide valuable detail information on the thiopurines (e.g. evaluating ratio of methylated and non-methylated 6-mercaptopurine) and, by that, TPMT action in biological systems before and during the therapy of IBD. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. The Roles of Two miRNAs in Regulating the Immune Response of Sea Cucumber.

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    Zhang, Pengjuan; Li, Chenghua; Zhang, Ran; Zhang, Weiwei; Jin, Chunhua; Wang, Lingling; Song, Linsheng

    2015-12-01

    MicroRNAs (miRNAs) have emerged as key regulators in many pathological processes by suppressing the transcriptional and post-transcriptional expression of target genes. MiR-2008 was previously found to be significantly up-regulated in diseased sea cucumber Apostichopus japonicus by high-through sequencing, whereas the reads of miR-137, a well-documented tumor repressor, displayed no significant change. In the present study, we found that miR-137 expression was slightly attenuated and miR-2008 was significantly enhanced after Vibrio splendidus infection or Lipopolysaccharides application. Further target screening and dual-luciferase reporter assay revealed that the two important miRNAs shared a common target gene of betaine-homocysteine S-methyltransferase (AjBHMT), which exhibited noncorrelated messenger RNA and protein expression patterns after bacterial challenge. In order to fully understand their regulatory mechanisms, we conducted the functional experiments in vitro and in vivo. The overexpression of miR-137 in sea cucumber or primary coelomocytes significantly decreased, whereas the inhibition of miR-137 increased the mRNA and protein expression levels of AjBHMT. In contrast, miR-2008 overexpression and inhibition showed no effect on AjBHMT mRNA levels, but the concentration of AjBHMT protein displayed significant changes both in vitro and in vivo. Consistently, the homocysteine (Hcy) contents were also accordingly altered in the aberrant expression analysis of both miRNAs, consistent with the results of the AjBHMT silencing assay in vitro and in vivo. More importantly, small interfering RNA mediated AjBHMT knockdown and Hcy exposure analyses both significantly increased reactive oxygen species (ROS) production and decreased the number of surviving invasive pathogen in sea cucumber coelomocytes. Taken together, these findings confirmed the differential roles of sea cucumber miR-137 and miR-2008 in regulating the common target AjBHMT to promote ROS production

  7. Evaluation of prescriber responses to pharmacogenomics clinical decision support for thiopurine S-methyltransferase testing.

    Science.gov (United States)

    Ubanyionwu, Samuel; Formea, Christine M; Anderson, Benjamin; Wix, Kelly; Dierkhising, Ross; Caraballo, Pedro J

    2018-02-15

    Results of a study of prescribers' responses to a pharmacogenomics-based clinical decision support (CDS) alert designed to prompt thiopurine S -methyltransferase (TPMT) status testing are reported. A single-center, retrospective, chart review-based study was conducted to evaluate prescriber compliance with a pretest CDS alert that warned of potential thiopurine drug toxicity resulting from deficient TPMT activity due to TPMT gene polymorphism. The CDS alert was triggered when prescribers ordered thiopurine drugs for patients whose records did not indicate TPMT status or when historical thiopurine use was documented in the electronic health record. The alert pop-up also provided a link to online educational resources to guide thiopurine dosing calculations. During the 9-month study period, 500 CDS alerts were generated: in 101 cases (20%), TPMT phenotyping or TPMT genotyping was ordered; in 399 cases (80%), testing was not ordered. Multivariable regression analysis indicated that documentation of historical thiopurine use was the only independent predictor of test ordering. Among the 99 patients tested subsequent to CDS alerts, 70 (71%) had normal TPMT activity, 29 (29%) had intermediate activity, and none had deficient activity. The online resources provided thiopurine dosing recommendations applicable to 24 patients, but only 3 were prescribed guideline-supported doses after CDS alerts. The pretest CDS rule resulted in a large proportion of neglected alerts due to poor alerting accuracy and consequent alert fatigue. Prescriber usage of online thiopurine dosing resources was low. Copyright © 2018 by the American Society of Health-System Pharmacists, Inc. All rights reserved.

  8. A Possible Mechanism: Vildagliptin Prevents Aortic Dysfunction through Paraoxonase and Angiopoietin-Like 3

    Directory of Open Access Journals (Sweden)

    Qian Zhang

    2018-01-01

    Full Text Available The collected data have revealed the beneficial effects of dipeptidyl peptidase-4 (DPP-4 inhibitors on the vascular endothelium, including vildagliptin. However, the involved mechanisms are not yet clear. In this study, Sprague-Dawley rats were randomly divided into the following four groups: control, diabetic, diabetic + low-dose vildagliptin (10 mg/kg/d, and diabetic + high-dose vildagliptin (20 mg/kg/d. The diabetic model was created by feeding a high-fat diet for four weeks and injection of streptozotocin. Then, vildagliptin groups were given oral vildagliptin for twelve weeks, and the control and diabetic groups were given the same volume of saline. The metabolic parameters, endothelial function, and whole genome expression in the aorta were examined. After 12 weeks of treatment, vildagliptin groups showed significantly reduced blood glucose, blood total cholesterol, and attenuated endothelial dysfunction. Notably, vildagliptin may inhibit angiopoietin-like 3 (Angptl3 and betaine-homocysteine S-methyltransferase (Bhmt expression and activated paraoxonase-1 (Pon1 in the aorta of diabetic rats. These findings may demonstrate the vasoprotective pathway of vildagliptin in vivo.

  9. Comparative transcriptome analysis on the alteration of gene expression in ayu (Plecoglossus altivelis) larvae associated with salinity change.

    Science.gov (United States)

    Lu, Xin-Jiang; Zhang, Hao; Yang, Guan-Jun; Li, Ming-Yun; Chen, Jiong

    2016-05-18

    Ayu (Plecoglossus altivelis) fish, which are an amphidromous species distributed in East Asia, live in brackish water (BW) during their larval stage and in fresh water (FW) during their adult stage. In this study, we found that FW-acclimated ayu larvae exhibited a slower growth ratio compared with that of BW-acclimated larvae. However, the mechanism underlying FW acclimation on growth suppression is poorly known. We employed transcriptome analysis to investigate the differential gene expression of FW acclimation by RNA sequencing. We identified 158 upregulated and 139 downregulated transcripts in FW-acclimated ayu larvae compared with that in BW-acclimated larvae. As determined by Gene Ontology annotation and Kyoto Encyclopedia of Genes and Genomes pathway mapping, functional annotation of the genes covered diverse biological functions and processes, and included neuroendocrinology, osmotic regulation, energy metabolism, and the cytoskeleton. Transcriptional expression of several differentially expressed genes in response to FW acclimation was further confirmed by real-time quantitative PCR. In accordance with transcriptome analysis, iodothyronine deiodinase (ID), pro-opiomelanocortin (POMC), betaine-homocysteine S-methyltransferase 1(BHMT), fructose-bisphosphate aldolase B (aldolase B), tyrosine aminotransferase (TAT), and Na(+)-K(+) ATPase (NKA) were upregulated after FW acclimation. Furthermore, the mRNA expressions of b-type natriuretic peptide (BNP) and transgelin were downregulated after FW acclimation. Our data indicate that FW acclimation reduced the growth rate of ayu larvae, which might result from the expression alteration of genes related to endocrine hormones, energy metabolism, and direct osmoregulation.

  10. The Metabolic Burden of Methyl Donor Deficiency with Focus on the Betaine Homocysteine Methyltransferase Pathway

    Directory of Open Access Journals (Sweden)

    Rima Obeid

    2013-09-01

    Full Text Available Methyl groups are important for numerous cellular functions such as DNA methylation, phosphatidylcholine synthesis, and protein synthesis. The methyl group can directly be delivered by dietary methyl donors, including methionine, folate, betaine, and choline. The liver and the muscles appear to be the major organs for methyl group metabolism. Choline can be synthesized from phosphatidylcholine via the cytidine-diphosphate (CDP pathway. Low dietary choline loweres methionine formation and causes a marked increase in S-adenosylmethionine utilization in the liver. The link between choline, betaine, and energy metabolism in humans indicates novel functions for these nutrients. This function appears to goes beyond the role of the nutrients in gene methylation and epigenetic control. Studies that simulated methyl-deficient diets reported disturbances in energy metabolism and protein synthesis in the liver, fatty liver, or muscle disorders. Changes in plasma concentrations of total homocysteine (tHcy reflect one aspect of the metabolic consequences of methyl group deficiency or nutrient supplementations. Folic acid supplementation spares betaine as a methyl donor. Betaine is a significant determinant of plasma tHcy, particularly in case of folate deficiency, methionine load, or alcohol consumption. Betaine supplementation has a lowering effect on post-methionine load tHcy. Hypomethylation and tHcy elevation can be attenuated when choline or betaine is available.

  11. Comparative transcriptome analysis on the alteration of gene expression in ayu (Plecoglossus altivelis larvae associated with salinity change

    Directory of Open Access Journals (Sweden)

    Xin-Jiang LU

    2016-05-01

    Full Text Available Ayu (Plecoglossus altivelis fish, which are an amphidromous species distributed in East Asia, live in brackish water (BW during their larval stage and in fresh water (FW during their adult stage. In this study, we found that FW-acclimated ayu larvae exhibited a slower growth ratio compared with that of BW-acclimated larvae. However, the mechanism underlying FW acclimation on growth suppression is poorly known. We employed transcriptome analysis to investigate the differential gene expression of FW acclimation by RNA sequencing. We identified 158 upregulated and 139 downregulated transcripts in FW-acclimated ayu larvae compared with that in BW-acclimated larvae. As determined by Gene Ontology annotation and Kyoto Encyclopedia of Genes and Genomes pathway mapping, functional annotation of the genes covered diverse biological functions and processes, and included neuroendocrinology, osmotic regulation, energy metabolism, and the cytoskeleton. Transcriptional expression of several differentially expressed genes in response to FW acclimation was further confirmed by real-time quantitative PCR. In accordance with transcriptome analysis, iodothyronine deiodinase (ID, pro-opiomelanocortin (POMC, betaine-homocysteine S-methyltransferase 1(BHMT, fructose-bisphosphate aldolase B (aldolase B, tyrosine aminotransferase (TAT, and Na+-K+ ATPase (NKA were upregulated after FW acclimation. Furthermore, the mRNA expressions of b-type natriuretic peptide (BNP and transgelin were downregulated after FW acclimation. Our data indicate that FW acclimation reduced the growth rate of ayu larvae, which might result from the expression alteration of genes related to endocrine hormones, energy metabolism, and direct osmoregulation.

  12. Methylation of 6-mercaptopurine and 6-thioguanine by thiopurine S-methyltransferase. A comparison of activity in red blood cell samples of 199 blood donors.

    Science.gov (United States)

    Kröplin, T; Iven, H

    2000-07-01

    To compare 6-mercaptopurine (6-MP) and 6-thioguanine (6-TG) as substrates for the methylation reaction catalysed by the enzyme thiopurine S-methyltransferase (TPMT). TPMT activity in haemolysed red blood cells of healthy blood donors was determined twice, using the same experimental setting and equal molar concentrations of 6-TG and 6-MP as substrates. After extraction, the reaction products 6-methyl-TG and 6-methyl-MP were quantified using specific high-performance liquid chromatography procedures. The medians of the TPMT activities from 199 blood donors were 54.4 nmol 6-MTG g(-1)Hb h(-1) when measured with 6-TG as the substrate and 35.8 nmol 6-MMP g(-1) Hb h(-1) when measured with 6-MP. The correlation coefficient for the 199 pairs of values was 0.8695. On average, TPMT activity was 34% lower with 6-MP as substrate than with 6-TG as substrate.

  13. Combining combinatorial chemistry and affinity chromatography protocols for systematically probing protein-ligand interactions: application to the development of highly selective phosphinic inhibitors of human bataine: homocysteine S-methyltransferase

    Czech Academy of Sciences Publication Activity Database

    Collinsová, Michaela; Garrow, T. A.; Castro, C.; Dive, V.; Yiotakis, A.; Jiráček, Jiří

    2002-01-01

    Roč. 8, - (2002), s. S221 ISSN 1075-2617. [European Peptide Symposium /27./. 31.08.2002-06.09.2002, Sorrento] Institutional research plan: CEZ:AV0Z4055905 Keywords : homocysteine S-methyltransferase Subject RIV: CE - Biochemistry

  14. Inhibition of thiopurine S-methyltransferase activity by impurities in commercially available substrates: a factor for differing results of TPMT measurements.

    Science.gov (United States)

    Kröplin, T; Fischer, C; Iven, H

    1999-06-01

    Thiopurine S-methyltransferase (TPMT) activity, when measured in red blood cells (RBC) with a recently published TPMT activity assay using 6-thioguanine (6-TG) as substrate, could not be reproduced in another laboratory. We investigated factors which could influence the results of the TPMT activity measurement. We tested twelve 6-TG and four 6-mercaptopurine (6-MP) compounds from different suppliers as substrates and determined the enzyme kinetic parameters Km and Vmax. Furthermore, we studied the influence of different 6-TG compounds on the affinity of the methyl donor S-adenosyl-L-methionine (SAM) to the TPMT enzyme. All 6-TG products were of equal purity (declared >98% by the supplier): this was ascertained by HPLC. However, the rate of methylation obtained following incubation with 6-TG from different suppliers ranged from 10% to 100% when incubated with the same RBC lysate. The lowest apparent Km value for a 6-TG was 22.3 micromol x l(-1), while the product with the highest methylation rate showed a Km of 156 micromol x l(-1). From these results we assume that there is a contaminant in some 6-TG products, which acts as a strong inhibitor of TPMT activity. Compounds possibly used for the synthesis of 6-TG (guanine, pyridine, 6-chloroguanine) did not affect the methylation rate. Thioxanthine, which is known to be a strong inhibitor of TPMT when added to the assay system to give a 2% contamination, reduced TPMT activity from 100% to 72%. Using 6-MP from different suppliers as substrate resulted in Km values ranging from 110 to 162 micromol x l(-1) and Vmox values ranging from 54 to 68 nmol 6-MMP x g(-1)Hb x h(-1). The Km value for the methyl donor SAM was similar to and independent from the thiopurine substrates tested (range 4.9-11 micromol-l(-1) SAM). In contrast to other investigators, we found non-enzymatic S-methylation, which was negligible under our assay conditions (3% with 128 micromol x l(-1) SAM), but could become relevant in experiments using higher

  15. Betaine reduces hepatic lipidosis induced by carbon tetrachloride in Sprague-Dawley rats.

    Science.gov (United States)

    Junnila, M; Barak, A J; Beckenhauer, H C; Rahko, T

    1998-10-01

    Carbon tetrachloride-injected rats were given liquid diets with and without betaine for 7 d. Hepatic lipidosis was induced by 4 daily injections of carbon tetrachloride (CCl4). Animals were killed and their livers and blood taken for analysis of betaine, S-adenosylmethionine (SAM), betaine homocysteine methyltransferase (BHMT), triglyceride, alanine aminotransferase and aspartate aminotransferase. Liver samples were also processed and stained for histological examination. Supplemental betaine reduced triglyceride in the liver and centrilobular hepatic lipidosis induced by the CCl4 injections. In both the control and experimental groups receiving betaine, liver betaine, BHMT and SAM were significantly higher than in their respective groups not receiving betaine. This study provides evidence that betaine protects the liver against CCl4-induced lipidosis and may be a useful therapeutic and prophylactic agent in ameliorating the harmful effects of CCl4.

  16. Dimethylglycine accumulates in uremia and predicts elevated plasma homocysteine concentrations.

    Science.gov (United States)

    McGregor, D O; Dellow, W J; Lever, M; George, P M; Robson, R A; Chambers, S T

    2001-06-01

    Hyperhomocysteinemia is a risk factor for atherosclerosis that is common in chronic renal failure (CRF), but its cause is unknown. Homocysteine metabolism is linked to betaine-homocysteine methyl transferase (BHMT), a zinc metalloenzyme that converts glycine betaine (GB) to N,N dimethylglycine (DMG). DMG is a known feedback inhibitor of BHMT. We postulated that DMG might accumulate in CRF and contribute to hyperhomocysteinemia by inhibiting BHMT activity. Plasma and urine concentrations of GB and DMG were measured in 33 dialysis patients (15 continuous ambulatory peritoneal dialysis and 18 hemodialysis), 33 patients with CRF, and 33 age-matched controls. Concentrations of fasting plasma total homocysteine (tHcy), red cell and serum folate, vitamins B(6) and B(12), serum zinc, and routine biochemistry were also measured. Groups were compared, and determinants of plasma tHcy were identified by correlations and stepwise linear regression. Plasma DMG increased as renal function declined and was twofold to threefold elevated in dialysis patients. Plasma GB did not differ between groups. The fractional excretion of GB (FE(GB)) was increased tenfold, and FED(MG) was doubled in CRF patients compared with controls. Plasma tHcy correlated positively with plasma DMG, the plasma DMG:GB ratio, plasma creatinine, and FE(GB) and negatively with serum folate, zinc, and plasma GB. In the multiple regression model, only plasma creatinine, plasma DMG, or the DMG:GB ratio was independent predictors of tHcy. DMG accumulates in CRF and independently predicts plasma tHcy concentrations. These findings suggest that reduced BHMT activity is important in the pathogenesis of hyperhomocysteinemia in CRF.

  17. Elevated dimethylglycine in blood of children with congenital heart defects and their mothers.

    Science.gov (United States)

    Alsayed, Ranwa; Al Quobaili, Faizeh; Srour, Samir; Geisel, Jürgen; Obeid, Rima

    2013-08-01

    Congenital Heart Defects (CHD) may be related to nutritional deficiencies affecting the methylation cycle. We aimed to study the metabolic markers of the betaine homocysteine methyl transferase (BHMT) pathway in children with CHD and their mothers compared to children without CHD and their mothers. Children with CHD (n=105, age DMG). Children with CHD had higher plasma SAM (131 vs. 100 nmol/L) and DMG (8.7 vs. 6.0 μmol/L) and lower betaine/DMG ratio (7.5 vs. 10.2) compared to the controls. Mothers of CHD children showed also higher DMG (6.1 vs. 4.1 µmol/L) and lower betaine/DMG ratio compared with the mothers of the controls. Higher SAM levels were related to higher cystathionine, MMA, betaine, choline, and DMG. MMA elevation in the patients was related to higher HCY, SAM, betaine and DMG. Elevated DMG in CHD children and their mothers compared to the controls can indicate upregulation of the BHMT pathway in this disease group. Nutritional factors are related to metabolic imbalance during pregnancy that may be related to worse birth outcome. Copyright © 2013 Elsevier Inc. All rights reserved.

  18. Choline and methionine differentially alter methyl carbon metabolism in bovine neonatal hepatocytes.

    Science.gov (United States)

    Chandler, Tawny L; White, Heather M

    2017-01-01

    Intersections in hepatic methyl group metabolism pathways highlights potential competition or compensation of methyl donors. The objective of this experiment was to examine the expression of genes related to methyl group transfer and lipid metabolism in response to increasing concentrations of choline chloride (CC) and DL-methionine (DLM) in primary neonatal hepatocytes that were or were not exposed to fatty acids (FA). Primary hepatocytes isolated from 4 neonatal Holstein calves were maintained as monolayer cultures for 24 h before treatment with CC (61, 128, 2028, and 4528 μmol/L) and DLM (16, 30, 100, 300 μmol/L), with or without a 1 mmol/L FA cocktail in a factorial arrangement. After 24 h of treatment, media was collected for quantification of reactive oxygen species (ROS) and very low-density lipoprotein (VLDL), and cell lysates were collected for quantification of gene expression. No interactions were detected between CC, DLM, or FA. Both CC and DLM decreased the expression of methionine adenosyltransferase 1A (MAT1A). Increasing CC did not alter betaine-homocysteine S-methyltranferase (BHMT) but did increase 5-methyltetrahydrofolate-homocysteine methyltransferase (MTR) and methylenetetrahydrofolate reductase (MTHFR) expression. Increasing DLM decreased expression of BHMT and MTR, but did not affect MTHFR. Expression of both phosphatidylethanolamine N-methyltransferase (PEMT) and microsomal triglyceride transfer protein (MTTP) were decreased by increasing CC and DLM, while carnitine palmitoyltransferase 1A (CPT1A) was unaffected by either. Treatment with FA decreased the expression of MAT1A, MTR, MTHFR and tended to decrease PEMT but did not affect BHMT and MTTP. Treatment with FA increased CPT1A expression. Increasing CC increased secretion of VLDL and decreased the accumulation of ROS in media. Within neonatal bovine hepatocytes, choline and methionine differentially regulate methyl carbon pathways and suggest that choline may play a critical role in

  19. Dynamic regulation of hepatic lipid droplet properties by diet.

    Science.gov (United States)

    Crunk, Amanda E; Monks, Jenifer; Murakami, Aya; Jackman, Matthew; Maclean, Paul S; Ladinsky, Mark; Bales, Elise S; Cain, Shannon; Orlicky, David J; McManaman, James L

    2013-01-01

    Cytoplasmic lipid droplets (CLD) are organelle-like structures that function in neutral lipid storage, transport and metabolism through the actions of specific surface-associated proteins. Although diet and metabolism influence hepatic CLD levels, how they affect CLD protein composition is largely unknown. We used non-biased, shotgun, proteomics in combination with metabolic analysis, quantitative immunoblotting, electron microscopy and confocal imaging to define the effects of low- and high-fat diets on CLD properties in fasted-refed mice. We found that the hepatic CLD proteome is distinct from that of CLD from other mammalian tissues, containing enzymes from multiple metabolic pathways. The hepatic CLD proteome is also differentially affected by dietary fat content and hepatic metabolic status. High fat feeding markedly increased the CLD surface density of perilipin-2, a critical regulator of hepatic neutral lipid storage, whereas it reduced CLD levels of betaine-homocysteine S-methyltransferase, an enzyme regulator of homocysteine levels linked to fatty liver disease and hepatocellular carcinoma. Collectively our data demonstrate that the hepatic CLD proteome is enriched in metabolic enzymes, and that it is qualitatively and quantitatively regulated by diet and metabolism. These findings implicate CLD in the regulation of hepatic metabolic processes, and suggest that their properties undergo reorganization in response to hepatic metabolic demands.

  20. A Protein Extract from Chicken Reduces Plasma Homocysteine in Rats

    Directory of Open Access Journals (Sweden)

    Vegard Lysne

    2015-06-01

    Full Text Available The present study aimed to evaluate effects of a water-soluble protein fraction of chicken (CP, with a low methionine/glycine ratio, on plasma homocysteine and metabolites related to homocysteine metabolism. Male Wistar rats were fed either a control diet with 20% w/w casein as the protein source, or an experimental diet where 6, 14 or 20% w/w of the casein was replaced with the same amount of CP for four weeks. Rats fed CP had reduced plasma total homocysteine level and markedly increased levels of the choline pathway metabolites betaine, dimethylglycine, sarcosine, glycine and serine, as well as the transsulfuration pathway metabolites cystathionine and cysteine. Hepatic mRNA level of enzymes involved in homocysteine remethylation, methionine synthase and betaine-homocysteine S-methyltransferase, were unchanged, whereas cystathionine gamma-lyase of the transsulfuration pathway was increased in the CP treated rats. Plasma concentrations of vitamin B2, folate, cobalamin, and the B-6 catabolite pyridoxic acid were increased in the 20% CP-treated rats. In conclusion, the CP diet was associated with lower plasma homocysteine concentration and higher levels of serine, choline oxidation and transsulfuration metabolites compared to a casein diet. The status of related B-vitamins was also affected by CP.

  1. Transplacental exposure to inorganic arsenic at a hepatocarcinogenic dose induces fetal gene expression changes in mice indicative of aberrant estrogen signaling and disrupted steroid metabolism

    International Nuclear Information System (INIS)

    Liu Jie; Xie Yaxiong; Cooper, Ryan; Ducharme, Danica M.K.; Tennant, Raymond; Diwan, Bhalchandra A.; Waalkes, Michael P.

    2007-01-01

    Exposure to inorganic arsenic in utero in C3H mice produces hepatocellular carcinoma in male offspring when they reach adulthood. To help define the molecular events associated with the fetal onset of arsenic hepatocarcinogenesis, pregnant C3H mice were given drinking water containing 0 (control) or 85 ppm arsenic from day 8 to 18 of gestation. At the end of the arsenic exposure period, male fetal livers were removed and RNA isolated for microarray analysis using 22K oligo chips. Arsenic exposure in utero produced significant (p < 0.001) alterations in expression of 187 genes, with approximately 25% of aberrantly expressed genes related to either estrogen signaling or steroid metabolism. Real-time RT-PCR on selected genes confirmed these changes. Various genes controlled by estrogen, including X-inactive-specific transcript, anterior gradient-2, trefoil factor-1, CRP-ductin, ghrelin, and small proline-rich protein-2A, were dramatically over-expressed. Estrogen-regulated genes including cytokeratin 1-19 and Cyp2a4 were over-expressed, although Cyp3a25 was suppressed. Several genes involved with steroid metabolism also showed remarkable expression changes, including increased expression of 17β-hydroxysteroid dehydrogenase-7 (HSD17β7; involved in estradiol production) and decreased expression of HSD17β5 (involved in testosterone production). The expression of key genes important in methionine metabolism, such as methionine adenosyltransferase-1a, betaine-homocysteine methyltransferase and thioether S-methyltransferase, were suppressed. Thus, exposure of mouse fetus to inorganic arsenic during a critical period in development significantly alters the expression of various genes encoding estrogen signaling and steroid or methionine metabolism. These alterations could disrupt genetic programming at the very early life stage, which could impact tumor formation much later in adulthood

  2. Azathioprine-associated acute myeloid leukemia in a patient with Crohn's disease and thiopurine S-methyltransferase deficiency

    DEFF Research Database (Denmark)

    Yenson, P.R.; Forrest, D.; Schmiegelow, K.

    2008-01-01

    risk of hematologic toxicity and leukemogenesis. We present such a patient who was a slow metabolizer for azathioprine, and developed a rapidly lethal form acute myeloid leukemia after relatively low dose exposure to the drug. There was prominent hemophagocytic activity in the bone marrow...

  3. Liver proteomics in progressive alcoholic steatosis

    Energy Technology Data Exchange (ETDEWEB)

    Fernando, Harshica [Department of Pathology, The University of Texas Medical Branch, Galveston, TX 77555 (United States); Wiktorowicz, John E.; Soman, Kizhake V. [Department of Biochemistry and Molecular Biology, The University of Texas Medical Branch, Galveston, TX 77555 (United States); Kaphalia, Bhupendra S.; Khan, M. Firoze [Department of Pathology, The University of Texas Medical Branch, Galveston, TX 77555 (United States); Shakeel Ansari, G.A., E-mail: sansari@utmb.edu [Department of Pathology, The University of Texas Medical Branch, Galveston, TX 77555 (United States)

    2013-02-01

    Fatty liver is an early stage of alcoholic and nonalcoholic liver disease (ALD and NALD) that progresses to steatohepatitis and other irreversible conditions. In this study, we identified proteins that were differentially expressed in the livers of rats fed 5% ethanol in a Lieber–DeCarli diet daily for 1 and 3 months by discovery proteomics (two-dimensional gel electrophoresis and mass spectrometry) and non-parametric modeling (Multivariate Adaptive Regression Splines). Hepatic fatty infiltration was significantly higher in ethanol-fed animals as compared to controls, and more pronounced at 3 months of ethanol feeding. Discovery proteomics identified changes in the expression of proteins involved in alcohol, lipid, and amino acid metabolism after ethanol feeding. At 1 and 3 months, 12 and 15 different proteins were differentially expressed. Of the identified proteins, down regulation of alcohol dehydrogenase (− 1.6) at 1 month and up regulation of aldehyde dehydrogenase (2.1) at 3 months could be a protective/adaptive mechanism against ethanol toxicity. In addition, betaine-homocysteine S-methyltransferase 2 a protein responsible for methionine metabolism and previously implicated in fatty liver development was significantly up regulated (1.4) at ethanol-induced fatty liver stage (1 month) while peroxiredoxin-1 was down regulated (− 1.5) at late fatty liver stage (3 months). Nonparametric analysis of the protein spots yielded fewer proteins and narrowed the list of possible markers and identified D-dopachrome tautomerase (− 1.7, at 3 months) as a possible marker for ethanol-induced early steatohepatitis. The observed differential regulation of proteins have potential to serve as biomarker signature for the detection of steatosis and its progression to steatohepatitis once validated in plasma/serum. -- Graphical abstract: The figure shows the Hierarchial cluster analysis of differentially expressed protein spots obtained after ethanol feeding for 1 (1–3

  4. Enhanced proteolysis of thiopurine S-methyltransferase (TPMT) encoded by mutant alleles in humans (TPMT∗3A, TPMT∗2): Mechanisms for the genetic polymorphism of TPMT activity

    OpenAIRE

    Tai, Hung-Liang; Krynetski, Eugene Y.; Schuetz, Erin G.; Yanishevski, Yuri; Evans, William E.

    1997-01-01

    TPMT is a cytosolic enzyme that catalyzes the S-methylation of aromatic and heterocyclic sulfhydryl compounds, including medications such as mercaptopurine and thioguanine. TPMT activity exhibits autosomal codominant genetic polymorphism, and patients inheriting TPMT deficiency are at high risk of potentially fatal hematopoietic toxicity. The most prevalent mutant alleles associated with TPMT deficiency in humans have been cloned and characterized (TPMT∗2 and TPMT∗3A), but the mechanisms for ...

  5. Gender and single nucleotide polymorphisms in MTHFR, BHMT, SPTLC1, CRBP2R, and SCARB1 are significant predictors of plasma homocysteine normalized by RBC folate in healthy adults.

    Science.gov (United States)

    Using linear regression models, we studied the main and two-way interaction effects of the predictor variables gender, age, BMI, and 64 folate/vitamin B-12/homocysteine/lipid/cholesterol-related single nucleotide polymorphisms (SNP) on log-transformed plasma homocysteine normalized by red blood cell...

  6. Evaluations of the trans-sulfuration pathway in multiple liver toxicity studies

    International Nuclear Information System (INIS)

    Schnackenberg, Laura K.; Chen Minjun; Sun, Jinchun; Holland, Ricky D.; Dragan, Yvonne; Tong Weida; Welsh, William; Beger, Richard D.

    2009-01-01

    Drug-induced liver injury has been associated with the generation of reactive metabolites, which are primarily detoxified via glutathione conjugation. In this study, it was hypothesized that molecules involved in the synthesis of glutathione would be diminished to replenish the glutathione depleted through conjugation reactions. Since S-adenosylmethionine (SAMe) is the primary source of the sulfur atom in glutathione, UPLC/MS and NMR were used to evaluate metabolites involved with the transulfuration pathway in urine samples collected during studies of eight liver toxic compounds in Sprague-Dawley rats. Urinary levels of creatine were increased on day 1 or day 2 in 8 high dose liver toxicity studies. Taurine concentration in urine was increased in only 3 of 8 liver toxicity studies while SAMe was found to be reduced in 4 of 5 liver toxicity studies. To further validate the results from the metabonomic studies, microarray data from rat liver samples following treatment with acetaminophen was obtained from the Gene Expression Omnibus (GEO) database. Some genes involved in the trans-sulfuration pathway, including guanidinoacetate N-methyltransferase, glycine N-methyltransferase, betaine-homocysteine methyltransferase and cysteine dioxygenase were found to be significantly decreased while methionine adenosyl transferase II, alpha increased at 24 h post-dosing, which is consistent with the SAMe and creatine findings. The metabolic and transcriptomic results show that the trans-sulfuration pathway from SAMe to glutathione was disturbed due to the administration of heptatotoxicants

  7. Lean Body Mass Harbors Sensing Mechanisms that Allow Safeguarding of Methionine Homeostasis

    Directory of Open Access Journals (Sweden)

    Yves Ingenbleek

    2017-09-01

    Full Text Available Protein-depleted states generate allosteric inhibition of liver cystathionine β-synthase (CBS, which governs the first enzymatic step of the transsulfuration cascade, resulting in upstream accretion of homocysteine (Hcy in body fluids. A similar Hcy increase may arise from normal hepatocytes undergoing experimentally-induced impairment of betaine-homocysteine methyltransferase (BHTM activity or from components of lean body mass (LBM submitted to any inflammatory disorder. LBM comprises a composite agglomeration of extrarenal tissues characterized by naturally occurring BHTM inactivity. As a result of cellular injury, LBM releases high concentrations of Hcy into the extracellular space, contrasting with the disruption of normal remethylation pathways. Hyperhomocysteinemia acts as a biomarker, reflecting the severity of insult and operating as an alarm signal. Elevated Hcy levels constitute a precursor pool recognized by a CBS coding region that reacts to meet increased methionine requirements in LBM tissues, using its enhanced production in hepatocytes. Preservation of methionine homeostasis benefits from its high metabolic priority and survival value.

  8. Lean Body Mass Harbors Sensing Mechanisms that Allow Safeguarding of Methionine Homeostasis.

    Science.gov (United States)

    Ingenbleek, Yves

    2017-09-20

    Protein-depleted states generate allosteric inhibition of liver cystathionine β-synthase (CBS), which governs the first enzymatic step of the transsulfuration cascade, resulting in upstream accretion of homocysteine (Hcy) in body fluids. A similar Hcy increase may arise from normal hepatocytes undergoing experimentally-induced impairment of betaine-homocysteine methyltransferase (BHTM) activity or from components of lean body mass (LBM) submitted to any inflammatory disorder. LBM comprises a composite agglomeration of extrarenal tissues characterized by naturally occurring BHTM inactivity. As a result of cellular injury, LBM releases high concentrations of Hcy into the extracellular space, contrasting with the disruption of normal remethylation pathways. Hyperhomocysteinemia acts as a biomarker, reflecting the severity of insult and operating as an alarm signal. Elevated Hcy levels constitute a precursor pool recognized by a CBS coding region that reacts to meet increased methionine requirements in LBM tissues, using its enhanced production in hepatocytes. Preservation of methionine homeostasis benefits from its high metabolic priority and survival value.

  9. Metabolism of S-adenosylmethionine in rat hepatocytes: transfer of methyl group from S-adenosylmethionine by methyltransferase reactions

    International Nuclear Information System (INIS)

    Tsukada, K.; Abe, T.; Kuwahata, T.; Mitsui, K.

    1985-01-01

    Treatment of rats with a methionine diet leads not only to a marked increase of S-adenosylmethionine synthetase in liver, but also to the increase of glycine, guanidoacetate and betaine-homocysteine methyltransferases. The activity of tRNA methyltransferase decreased with the increased amounts of methionine in the diets. However, the activities of phospholipids and S-adenosylmethionine-homocysteine methyltransferases did not show any significant change. When hepatocarcinogenesis induced by 2-fluorenylacetamide progresses, the activities of glycine and guanidoacetate methyltransferases in rat liver decreased, and could not be detected in tumorous areas 8 months after treatment. The levels of S-adenosylmethionine in the liver also decreased to levels of one-fifth of control animals at 8 months. The uptake and metabolism of [methyl- 3 H]-methionine and -S-adenosylmethionine have been investigated by in vivo and isolated hepatocytes. The uptake of methionine and transfer of methyl group to phospholipid in the cells by methionine were remarkably higher than those by S-adenosylmethionine. These results indicate that phospholipids in hepatocytes accept methyl group from S-adenosylmethionine immediately, when it is synthesized from methionine, before mixing its pool in the cells. 39 references, 1 figure, 2 tables

  10. Differential proteomic and tissue expression analyses identify valuable diagnostic biomarkers of hepatocellular differentiation and hepatoid adenocarcinomas.

    Science.gov (United States)

    Reis, Henning; Padden, Juliet; Ahrens, Maike; Pütter, Carolin; Bertram, Stefanie; Pott, Leona L; Reis, Anna-Carinna; Weber, Frank; Juntermanns, Benjamin; Hoffmann, Andreas-C; Eisenacher, Martin; Schlaak, Joörg F; Canbay, Ali; Meyer, Helmut E; Sitek, Barbara; Baba, Hideo A

    2015-10-01

    The exact discrimination of lesions with true hepatocellular differentiation from secondary tumours and neoplasms with hepatocellular histomorphology like hepatoid adenocarcinomas (HAC) is crucial. Therefore, we aimed to identify ancillary protein biomarkers by using complementary proteomic techniques (2D-DIGE, label-free MS). The identified candidates were immunohistochemically validated in 14 paired samples of hepatocellular carcinoma (HCC) and non-tumourous liver tissue (NT). The candidates and HepPar1/Arginase1 were afterwards tested for consistency in a large cohort of hepatocellular lesions and NT (n = 290), non-hepatocellular malignancies (n = 383) and HAC (n = 13). Eight non-redundant, differentially expressed proteins were suitable for further immunohistochemical validation and four (ABAT, BHMT, FABP1, HAOX1) for further evaluation. Sensitivity and specificity rates for HCC/HAC were as follows: HepPar1 80.2%, 94.3% / 80.2%, 46.2%; Arginase1 82%, 99.4% / 82%, 69.2%; BHMT 61.4%, 93.8% / 61.4%, 100%; ABAT 84.4%, 33.7% / 84.4%, 30.8%; FABP1 87.2%, 95% / 87.2%, 69.2%; HAOX1 95.5%, 36.3% / 95.5%, 46.2%. The best 2×/3× biomarker panels for the diagnosis of HCC consisted of Arginase1/HAOX1 and BHMT/Arginase1/HAOX1 and for HAC consisted of Arginase1/FABP1 and BHMT/Arginase1/FABP1. In summary, we successfully identified, validated and benchmarked protein biomarker candidates of hepatocellular differentiation. BHMT in particular exhibited superior diagnostic characteristics in hepatocellular lesions and specifically in HAC. BHMT is therefore a promising (panel based) biomarker candidate in the differential diagnostic process of lesions with hepatocellular aspect.

  11. Dietary intake of S-(alpha-carboxybutyl)-DL-homocysteine induces hyperhomocysteinemia in rats

    Czech Academy of Sciences Publication Activity Database

    Straková, J.; Williams, K. T.; Gupta, S.; Schalinske, K. L.; Kruger, W. D.; Rozen, R.; Jiráček, Jiří; Li, L.; Garrow, T. A.

    2010-01-01

    Roč. 30, č. 7 (2010), s. 492-500 ISSN 0271-5317 R&D Projects: GA ČR(CZ) GAP207/10/1277 Grant - others:NIH(US) DK52501 Institutional research plan: CEZ:AV0Z40550506 Keywords : BHMT * homocysteine * rat * betaine Subject RIV: CC - Organic Chemistry Impact factor: 2.092, year: 2010

  12. Regulation of homocysteine metabolism and methylation in human and mouse tissues

    Science.gov (United States)

    Chen, Natalie C.; Yang, Fan; Capecci, Louis M.; Gu, Ziyu; Schafer, Andrew I.; Durante, William; Yang, Xiao-Feng; Wang, Hong

    2010-01-01

    Hyperhomocysteinemia is an independent risk factor for cardiovascular disease. Homocysteine (Hcy) metabolism involves multiple enzymes; however, tissue Hcy metabolism and its relevance to methylation remain unknown. Here, we established gene expression profiles of 8 Hcy metabolic and 12 methylation enzymes in 20 human and 19 mouse tissues through bioinformatic analysis using expression sequence tag clone counts in tissue cDNA libraries. We analyzed correlations between gene expression, Hcy, S-adenosylhomocysteine (SAH), and S-adenosylmethionine (SAM) levels, and SAM/SAH ratios in mouse tissues. Hcy metabolic and methylation enzymes were classified into two types. The expression of Type 1 enzymes positively correlated with tissue Hcy and SAH levels. These include cystathionine β-synthase, cystathionine-γ-lyase, paraxonase 1, 5,10-methylenetetrahydrofolate reductase, betaine:homocysteine methyltransferase, methionine adenosyltransferase, phosphatidylethanolamine N-methyltransferases and glycine N-methyltransferase. Type 2 enzyme expressions correlate with neither tissue Hcy nor SAH levels. These include SAH hydrolase, methionyl-tRNA synthase, 5-methyltetrahydrofolate:Hcy methyltransferase, S-adenosylmethionine decarboxylase, DNA methyltransferase 1/3a, isoprenylcysteine carboxyl methyltransferases, and histone-lysine N-methyltransferase. SAH is the only Hcy metabolite significantly correlated with Hcy levels and methylation enzyme expression. We established equations expressing combined effects of methylation enzymes on tissue SAH, SAM, and SAM/SAH ratios. Our study is the first to provide panoramic tissue gene expression profiles and mathematical models of tissue methylation regulation.—Chen, N. C., Yang, F., Capecci, L. M., Gu, Z., Schafer, A. I., Durante, W., Yang, X.-F., Wang, H. Regulation of homocysteine metabolism and methylation in human and mouse tissues. PMID:20305127

  13. Differential gene expression in the liver of the African lungfish, Protopterus annectens, after 6 months of aestivation in air or 1 day of arousal from 6 months of aestivation.

    Directory of Open Access Journals (Sweden)

    Kum C Hiong

    Full Text Available The African lungfish, Protopterus annectens, can undergo aestivation during drought. Aestivation has three phases: induction, maintenance and arousal. The objective of this study was to examine the differential gene expression in the liver of P. annectens after 6 months (the maintenance phase of aestivation as compared with the freshwater control, or after 1 day of arousal from 6 months aestivation as compared with 6 months of aestivation using suppression subtractive hybridization. During the maintenance phase of aestivation, the mRNA expression of argininosuccinate synthetase 1 and carbamoyl phosphate synthetase III were up-regulated, indicating an increase in the ornithine-urea cycle capacity to detoxify ammonia to urea. There was also an increase in the expression of betaine homocysteine-S-transferase 1 which could reduce and prevent the accumulation of hepatic homocysteine. On the other hand, the down-regulation of superoxide dismutase 1 expression could signify a decrease in ROS production during the maintenance phase of aestivation. In addition, the maintenance phase was marked by decreases in expressions of genes related to blood coagulation, complement fixation and iron and copper metabolism, which could be strategies used to prevent thrombosis and to conserve energy. Unlike the maintenance phase of aestivation, there were increases in expressions of genes related to nitrogen, carbohydrate and lipid metabolism and fatty acid transport after 1 day of arousal from 6 months aestivation. There were also up-regulation in expressions of genes that were involved in the electron transport system and ATP synthesis, indicating a greater demand for metabolic energy during arousal. Overall, our results signify the importance of sustaining a low rate of waste production and conservation of energy store during the maintenance phase, and the dependence on internal energy store for repair and structural modification during the arousal phase, of

  14. Cobalamin-Independent Methionine Synthase (MetE): A Face-to-Face Double Barrel that Evolved by Gene Duplication

    Energy Technology Data Exchange (ETDEWEB)

    Pejcha, Robert; Ludwig, Martha L. (Michigan)

    2010-03-08

    Cobalamin-independent methionine synthase (MetE) catalyzes the transfer of a methyl group from methyltetrahydrofolate to L-homocysteine (Hcy) without using an intermediate methyl carrier. Although MetE displays no detectable sequence homology with cobalamin-dependent methionine synthase (MetH), both enzymes require zinc for activation and binding of Hcy. Crystallographic analyses of MetE from T. maritima reveal an unusual dual-barrel structure in which the active site lies between the tops of the two ({beta}{alpha}){sub 8} barrels. The fold of the N-terminal barrel confirms that it has evolved from the C-terminal polypeptide by gene duplication; comparisons of the barrels provide an intriguing example of homologous domain evolution in which binding sites are obliterated. The C-terminal barrel incorporates the zinc ion that binds and activates Hcy. The zinc-binding site in MetE is distinguished from the (Cys){sub 3}Zn site in the related enzymes, MetH and betaine-homocysteine methyltransferase, by its position in the barrel and by the metal ligands, which are histidine, cysteine, glutamate, and cysteine in the resting form of MetE. Hcy associates at the face of the metal opposite glutamate, which moves away from the zinc in the binary E {center_dot} Hcy complex. The folate substrate is not intimately associated with the N-terminal barrel; instead, elements from both barrels contribute binding determinants in a binary complex in which the folate substrate is incorrectly oriented for methyl transfer. Atypical locations of the Hcy and folate sites in the C-terminal barrel presumably permit direct interaction of the substrates in a ternary complex. Structures of the binary substrate complexes imply that rearrangement of folate, perhaps accompanied by domain rearrangement, must occur before formation of a ternary complex that is competent for methyl transfer.

  15. SULPHUR-CONTAINING AMINO ACIDS METABOLISM IN EXPERIMENTAL HYPER- AND HYPOTHYROIDISM IN RATS.

    Science.gov (United States)

    Nechiporuk, V; Zaichko, N; Korda, М; Melnyk, A; Koloshko, O

    2017-10-01

    Hyper- and hypothyroidism are some of the most common endocrinopathies that cause many metabolic disorders including amino acids metabolism. However, a specific molecular mechanism of thyroid hormones influence on sulphur-containing amino acids metabolism has not been established. The aim of our research was to investigate experimentally the influence of thyroid gland functional state on the main enzymatic systems of sulphur-containing amino acids metabolism in liver and kidneys, the content of homocysteine, cysteine and H2S in blood. The rats were administered with L-thyroxine and mercazolil to simulate the states of hyper- and hypothyroidism, which were confirmed by the content of fT3, fT4 and TSH in the blood. In liver and kidneys of the animals with hypothyroidism we observed the decrease in the activity of enzymes of remethylation cycle of S-adenosylmethioninsyntase, S-adenosylhomocysteinhyhdrolase, betaine-homocysteine methyltransferase. Suppression of transsulfuration transformation of homocysteine to cysteine in hypothyroidism was mainly due to the inhibition of cystathionine synthase activity of cystathionine-β-synthase, wherein cystathionase activity of cystathionine-γ-lyase was not changed. In animals with hypothyroidism we also noticed the inhibition of cysteine desulfunation reactions: the activity of enzymes of cystathionine-β-synthase, cystathionine-γ-lyase and cysteine aminotransferase significantly decreased in liver and kidneys. Experimental hyperthyroidism was accompanied by increase in activity of remethylation cycle enzymes, increase in cystationine synthase activity of cystathionine-β-synthase in liver and activity of these enzymes in kidneys. The simulation of hyperthyroidism led to the decrease of homocysteine concentration, and of hypothyroidism - to the increase of homocysteine and cysteine concentrations and reduced H2S content in blood of the animals. Thus, the significant risk factors for the development of atherosclerosis

  16. Cobalamin-Independent Methionine Synthase (MetE): A Face-to-Face Double Barrel that Evolved by Gene Duplication

    International Nuclear Information System (INIS)

    Pejcha, Robert; Ludwig, Martha L.

    2005-01-01

    Cobalamin-independent methionine synthase (MetE) catalyzes the transfer of a methyl group from methyltetrahydrofolate to L-homocysteine (Hcy) without using an intermediate methyl carrier. Although MetE displays no detectable sequence homology with cobalamin-dependent methionine synthase (MetH), both enzymes require zinc for activation and binding of Hcy. Crystallographic analyses of MetE from T. maritima reveal an unusual dual-barrel structure in which the active site lies between the tops of the two (βα) 8 barrels. The fold of the N-terminal barrel confirms that it has evolved from the C-terminal polypeptide by gene duplication; comparisons of the barrels provide an intriguing example of homologous domain evolution in which binding sites are obliterated. The C-terminal barrel incorporates the zinc ion that binds and activates Hcy. The zinc-binding site in MetE is distinguished from the (Cys) 3 Zn site in the related enzymes, MetH and betaine-homocysteine methyltransferase, by its position in the barrel and by the metal ligands, which are histidine, cysteine, glutamate, and cysteine in the resting form of MetE. Hcy associates at the face of the metal opposite glutamate, which moves away from the zinc in the binary E · Hcy complex. The folate substrate is not intimately associated with the N-terminal barrel; instead, elements from both barrels contribute binding determinants in a binary complex in which the folate substrate is incorrectly oriented for methyl transfer. Atypical locations of the Hcy and folate sites in the C-terminal barrel presumably permit direct interaction of the substrates in a ternary complex. Structures of the binary substrate complexes imply that rearrangement of folate, perhaps accompanied by domain rearrangement, must occur before formation of a ternary complex that is competent for methyl transfer.

  17. Cobalamin-independent methionine synthase (MetE: a face-to-face double barrel that evolved by gene duplication.

    Directory of Open Access Journals (Sweden)

    Robert Pejchal

    2005-02-01

    Full Text Available Cobalamin-independent methionine synthase (MetE catalyzes the transfer of a methyl group from methyltetrahydrofolate to L-homocysteine (Hcy without using an intermediate methyl carrier. Although MetE displays no detectable sequence homology with cobalamin-dependent methionine synthase (MetH, both enzymes require zinc for activation and binding of Hcy. Crystallographic analyses of MetE from T. maritima reveal an unusual dual-barrel structure in which the active site lies between the tops of the two (betaalpha(8 barrels. The fold of the N-terminal barrel confirms that it has evolved from the C-terminal polypeptide by gene duplication; comparisons of the barrels provide an intriguing example of homologous domain evolution in which binding sites are obliterated. The C-terminal barrel incorporates the zinc ion that binds and activates Hcy. The zinc-binding site in MetE is distinguished from the (Cys(3Zn site in the related enzymes, MetH and betaine-homocysteine methyltransferase, by its position in the barrel and by the metal ligands, which are histidine, cysteine, glutamate, and cysteine in the resting form of MetE. Hcy associates at the face of the metal opposite glutamate, which moves away from the zinc in the binary E.Hcy complex. The folate substrate is not intimately associated with the N-terminal barrel; instead, elements from both barrels contribute binding determinants in a binary complex in which the folate substrate is incorrectly oriented for methyl transfer. Atypical locations of the Hcy and folate sites in the C-terminal barrel presumably permit direct interaction of the substrates in a ternary complex. Structures of the binary substrate complexes imply that rearrangement of folate, perhaps accompanied by domain rearrangement, must occur before formation of a ternary complex that is competent for methyl transfer.

  18. Sulfur amino acid metabolism in doxorubicin-resistant breast cancer cells

    International Nuclear Information System (INIS)

    Ryu, Chang Seon; Kwak, Hui Chan; Lee, Kye Sook; Kang, Keon Wook; Oh, Soo Jin; Lee, Ki Ho; Kim, Hwan Mook; Ma, Jin Yeul; Kim, Sang Kyum

    2011-01-01

    Although methionine dependency is a phenotypic characteristic of tumor cells, it remains to be determined whether changes in sulfur amino acid metabolism occur in cancer cells resistant to chemotherapeutic medications. We compared expression/activity of sulfur amino acid metabolizing enzymes and cellular levels of sulfur amino acids and their metabolites between normal MCF-7 cells and doxorubicin-resistant MCF-7 (MCF-7/Adr) cells. The S-adenosylmethionine/S-adenosylhomocysteine ratio, an index of transmethylation potential, in MCF-7/Adr cells decreased to ∼ 10% relative to that in MCF-7 cells, which may have resulted from down-regulation of S-adenosylhomocysteine hydrolase. Expression of homocysteine-clearing enzymes, such as cystathionine beta-synthase, methionine synthase/methylene tetrahydrofolate reductase, and betaine homocysteine methyltransferase, was up-regulated in MCF-7/Adr cells, suggesting that acquiring doxorubicin resistance attenuated methionine-dependence and activated transsulfuration from methionine to cysteine. Homocysteine was similar, which is associated with a balance between the increased expressions of homocysteine-clearing enzymes and decreased extracellular homocysteine. Despite an elevation in cysteine, cellular GSH decreased in MCF-7/Adr cells, which was attributed to over-efflux of GSH into the medium and down-regulation of the GSH synthesis enzyme. Consequently, MCF-7/Adr cells were more sensitive to the oxidative stress induced by bleomycin and menadione than MCF-7 cells. In conclusion, our results suggest that regulating sulfur amino acid metabolism may be a possible therapeutic target for chemoresistant cancer cells. These results warrant further investigations to determine the role of sulfur amino acid metabolism in acquiring anticancer drug resistance in cancer cells using chemical and biological regulators involved in sulfur amino acid metabolism. - Research highlights: → MCF-7/Adr cells showed decreases in cellular GSH

  19. Detecting 16S rRNA Methyltransferases in Enterobacteriaceae by Use of Arbekacin.

    Science.gov (United States)

    McGann, Patrick; Chahine, Sarah; Okafor, Darius; Ong, Ana C; Maybank, Rosslyn; Kwak, Yoon I; Wilson, Kerry; Zapor, Michael; Lesho, Emil; Hinkle, Mary

    2016-01-01

    16S rRNA methyltransferases confer resistance to most aminoglycosides, but discriminating their activity from that of aminoglycoside-modifying enzymes (AMEs) is challenging using phenotypic methods. We demonstrate that arbekacin, an aminoglycoside refractory to most AMEs, can rapidly detect 16S methyltransferase activity in Enterobacteriaceae with high specificity using the standard disk susceptibility test. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  20. The relative contribution of genes operating in the S-methylmethionine cycle to methionine metabolism in Arabidopsis seeds.

    Science.gov (United States)

    Cohen, Hagai; Salmon, Asaf; Tietel, Zipora; Hacham, Yael; Amir, Rachel

    2017-05-01

    Enzymes operating in the S -methylmethionine cycle make a differential contribution to methionine synthesis in seeds. In addition, mutual effects exist between the S -methylmethionine cycle and the aspartate family pathway in seeds. Methionine, a sulfur-containing amino acid, is a key metabolite in plant cells. The previous lines of evidence proposed that the S-methylmethionine (SMM) cycle contributes to methionine synthesis in seeds where methionine that is produced in non-seed tissues is converted to SMM and then transported via the phloem into the seeds. However, the relative regulatory roles of the S-methyltransferases operating within this cycle in seeds are yet to be fully understood. In the current study, we generated transgenic Arabidopsis seeds with altered expression of three HOMOCYSTEINE S-METHYLTRANSFERASEs (HMTs) and METHIONINE S-METHYLTRANSFERASE (MMT), and profiled them for transcript and metabolic changes. The results revealed that AtHMT1 and AtHMT3, but not AtHMT2 and AtMMT, are the predominant enzymes operating in seeds as altered expression of these two genes affected the levels of methionine and SMM in transgenic seeds. Their manipulations resulted in adapted expression level of genes participating in methionine synthesis through the SMM and aspartate family pathways. Taken together, our findings provide new insights into the regulatory roles of the SMM cycle and the mutual effects existing between the two methionine biosynthesis pathways, highlighting the complexity of the metabolism of methionine and SMM in seeds.

  1. The role of zinc in genomic stability

    International Nuclear Information System (INIS)

    Sharif, Razinah; Thomas, Philip; Zalewski, Peter; Fenech, Michael

    2012-01-01

    Zinc (Zn) is an essential trace element required for maintaining both optimal human health and genomic stability. Zn plays a critical role in the regulation of DNA repair mechanisms, cell proliferation, differentiation and apoptosis involving the action of various transcriptional factors and DNA or RNA polymerases. Zn is an essential cofactor or structural component for important antioxidant defence proteins and DNA repair enzymes such as Cu/Zn SOD, OGG1, APE and PARP and may also affect activities of enzymes such as BHMT and MTR involved in methylation reactions in the folate-methionine cycle. This review focuses on the role of Zn in the maintenance of genome integrity and the effects of deficiency or excess on genomic stability events and cell death.

  2. Preparation and reactivity studies of maleimidophenyl glycidylether with amines

    Directory of Open Access Journals (Sweden)

    B. Lakshmi

    2011-04-01

    Full Text Available The hybrid thermosetting maleimido epoxy compound 4-(N-maleimidophenyl glycidylether (N-MPGE was prepared by reacting N-(4-hydroxyphenyl maleimide (HPM with epichlorohydrin by using benzyltrimethylammonium chloride as a catalyst. The resulting compound possessed both the oxirane ring and maleimide group. The curing reaction of N-MPGE with amines as curing agents such as tetraethylenepentamine (TEPA, bishexamethylenetriamine (BHMT, 2-methyl pentamethylenediamine (2-MPMDA and 2,2,4-trimethy diamines (2,2,4-TMDA was studied. The cured samples exhibited good thermal stability and excellent chemical (acid/alkali/solvent and water absorption resistance. Morphological studies by the SEM technique further confirmed the phase homogeneity network of the cured systems.

  3. Pharmacogenetics of anti-cancer drugs: State of the art and implementation - recommendations of the French National Network of Pharmacogenetics.

    Science.gov (United States)

    Quaranta, Sylvie; Thomas, Fabienne

    2017-04-01

    Individualized treatment is of special importance in oncology because the drugs used for chemotherapy have a very narrow therapeutic index. Pharmacogenetics may contribute substantially to clinical routine for optimizing cancer treatment to limit toxic effects while maintaining efficacy. This review presents the usefulness of pharmacogenetic tests for some key applications: dihydropyrimidine dehydrogenase (DPYD) genotyping for fluoropyrimidine (5-fluorouracil, capecitabine), UDP glucuronosylstransferase (UGT1A1) for irinotecan and thiopurine S-methyltransferase (TPMT) for thiopurine drugs. Depending on the level of evidence, the French National Network of Pharmacogenetics (RNPGx) has issued three levels of recommendations for these pharmacogenetic tests: essential, advisable, and potentially useful. Other applications, for which the level of evidence is still discussed, will be evoked in the final section of this review. Copyright © 2017 Société française de pharmacologie et de thérapeutique. Published by Elsevier Masson SAS. All rights reserved.

  4. [Effects of dietary wheat gluten level on decreasing plasma homocysteine concentration in rats].

    Science.gov (United States)

    Liu, Yiqun; Han, Feng; Sun, Licui; Lu, Jiaxi; Sugiyama, Kimio; Huang, Zhenwu

    2015-05-01

    To investigate the effects of different level of casein and wheat gluten on decreasing plasma homocysteine concentration in rats. 48 rats of the Wistar were fed with different level of casein (12.5%, 25% and 50%) and wheat gluten (14.5%, 29% and 58%) diets for 14 days, and they were killed by decapitation to obtain blood and livers was subject to analysis the concentration of homocysteine, cysteine and other amino acids, as well as BHMT and CBS activities. Body weight gain in rats fed wheat gluten dietary was significantly less than casein dietary, but food intake was significantly decreased in wheat gluten group with increasing of the protein content. The plasma homocysteine concentration in rats fed wheat gluten was marketly less than casein, however plasma cysteine concentration in wheat gluten was higher than casein group. The effects of wheat gluten on plasma homocysteine concentration are mainly depends on the low contents of methionine and high cysteine content, but the low contents of lyscine and threonine are not ignored. The mainly mechanism is that the increased cysteine concentration promot enzyme activities of homocystein metabolism, and increase the consumption of homocysteine.

  5. Relación de la homocisteína y las enzimas involucradas en su metabolismo con enfermedad cardiovascular y otras patologías.

    Directory of Open Access Journals (Sweden)

    Natalia Arias Morales

    2016-06-01

    oficiales (OMS. Tras el análisis de la información recolectada y en cuanto a las patologías reportadas con mayor frecuencia relacionadas con alteraciones de la concentración de la HCY, se encontró un 46% de literatura reportada con enfermedad cardiovascular, un 15% de reportes de enfermedades de tipo neurológico, un 11% con cáncer, un 9% con patologías en neonatos, un 4% con hipertensión arterial, así como otras enfermedades reportadas en menor porcentaje como las enfermedades metabólicas (3%, mixtas (4%, autoinmunes (2%, anormalidades en la gestación (2%, patologías óseas (1%, musculares (1% y hematológicas (1% como también la infertilidad (1%. En cuanto a las enzimas reportadas que influyen en el metabolismo de la HCY como participantes en las posibles alteraciones y patologías, el mayor número de reportes está relacionado con la metiltetrahidrofolato reductasa-(MTHFR (45% seguido de cistationina beta sintasa-(CBS (13%, betaina homocisteína metiltransferasa-(BHMT (9%, metil transferasa-(MTR (2%, metionina sintetasa reductasa-(MTRR (2% y otros porcentajes que incluyen más de dos enzimas reportadas. Conclusiones: Las principales patologías relacionadas con hHCY reportadas en la literatura son ECV, neurológicas, neoplásicas, metabólicas, neonatales y la hipertensión arterial. La MTHFR, BHMT y CBS son las enzimas más asociadas al metabolismo de HCY, cuyas alteraciones derivan en el aumento de niveles séricos de este aminoácido; la MTHFR se relaciona con cuadros de estenosis coronaria, cardiopatía congénita, Síndrome de Down (SD y neurotoxicidad. La BHMT tiene asociación con el desarrollo de SD, hepatocarcinogénesis, enfermedad coronaria, infertilidad idiopática en hombres, labio leporino y paladar hendido. La CBS se ha vinculado con ECV, enfermedades cognitivas, labio leporino y/o paladar hendido y cáncer de seno. La MTHFR es la más reportada en cuanto a cambios a nivel genético en su secuencia que se asocian a las patologías en las que

  6. Blood lead levels, iron metabolism gene polymorphisms and homocysteine: a gene-environment interaction study.

    Science.gov (United States)

    Kim, Kyoung-Nam; Lee, Mee-Ri; Lim, Youn-Hee; Hong, Yun-Chul

    2017-12-01

    Homocysteine has been causally associated with various adverse health outcomes. Evidence supporting the relationship between lead and homocysteine levels has been accumulating, but most prior studies have not focused on the interaction with genetic polymorphisms. From a community-based prospective cohort, we analysed 386 participants (aged 41-71 years) with information regarding blood lead and plasma homocysteine levels. Blood lead levels were measured between 2001 and 2003, and plasma homocysteine levels were measured in 2007. Interactions of lead levels with 42 genotyped single-nucleotide polymorphisms (SNPs) in five genes ( TF , HFE , CBS , BHMT and MTR ) were assessed via a 2-degree of freedom (df) joint test and a 1-df interaction test. In secondary analyses using imputation, we further assessed 58 imputed SNPs in the TF and MTHFR genes. Blood lead concentrations were positively associated with plasma homocysteine levels (p=0.0276). Six SNPs in the TF and MTR genes were screened using the 2-df joint test, and among them, three SNPs in the TF gene showed interactions with lead with respect to homocysteine levels through the 1-df interaction test (plead levels. Blood lead levels were positively associated with plasma homocysteine levels measured 4-6 years later, and three SNPs in the TF gene modified the association. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

  7. 118 SNPs of folate-related genes and risks of spina bifida and conotruncal heart defects

    Directory of Open Access Journals (Sweden)

    Shaw Gary M

    2009-06-01

    Full Text Available Abstract Background Folic acid taken in early pregnancy reduces risks for delivering offspring with several congenital anomalies. The mechanism by which folic acid reduces risk is unknown. Investigations into genetic variation that influences transport and metabolism of folate will help fill this data gap. We focused on 118 SNPs involved in folate transport and metabolism. Methods Using data from a California population-based registry, we investigated whether risks of spina bifida or conotruncal heart defects were influenced by 118 single nucleotide polymorphisms (SNPs associated with the complex folate pathway. This case-control study included 259 infants with spina bifida and a random sample of 359 nonmalformed control infants born during 1983–86 or 1994–95. It also included 214 infants with conotruncal heart defects born during 1983–86. Infant genotyping was performed blinded to case or control status using a designed SNPlex assay. We examined single SNP effects for each of the 118 SNPs, as well as haplotypes, for each of the two outcomes. Results Few odds ratios (ORs revealed sizable departures from 1.0. With respect to spina bifida, we observed ORs with 95% confidence intervals that did not include 1.0 for the following SNPs (heterozygous or homozygous relative to the reference genotype: BHMT (rs3733890 OR = 1.8 (1.1–3.1, CBS (rs2851391 OR = 2.0 (1.2–3.1; CBS (rs234713 OR = 2.9 (1.3–6.7; MTHFD1 (rs2236224 OR = 1.7 (1.1–2.7; MTHFD1 (hcv11462908 OR = 0.2 (0–0.9; MTHFD2 (rs702465 OR = 0.6 (0.4–0.9; MTHFD2 (rs7571842 OR = 0.6 (0.4–0.9; MTHFR (rs1801133 OR = 2.0 (1.2–3.1; MTRR (rs162036 OR = 3.0 (1.5–5.9; MTRR (rs10380 OR = 3.4 (1.6–7.1; MTRR (rs1801394 OR = 0.7 (0.5–0.9; MTRR (rs9332 OR = 2.7 (1.3–5.3; TYMS (rs2847149 OR = 2.2 (1.4–3.5; TYMS (rs1001761 OR = 2.4 (1.5–3.8; and TYMS (rs502396 OR = 2.1 (1.3–3.3. However, multiple SNPs observed for a given gene showed evidence of linkage disequilibrium indicating

  8. Comprehensive evaluation of one-carbon metabolism pathway gene variants and renal cell cancer risk.

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    Todd M Gibson

    Full Text Available Folate and one-carbon metabolism are linked to cancer risk through their integral role in DNA synthesis and methylation. Variation in one-carbon metabolism genes, particularly MTHFR, has been associated with risk of a number of cancers in epidemiologic studies, but little is known regarding renal cancer.Tag single nucleotide polymorphisms (SNPs selected to produce high genomic coverage of 13 gene regions of one-carbon metabolism (ALDH1L1, BHMT, CBS, FOLR1, MTHFR, MTR, MTRR, SHMT1, SLC19A1, TYMS and the closely associated glutathione synthesis pathway (CTH, GGH, GSS were genotyped for 777 renal cell carcinoma (RCC cases and 1,035 controls in the Central and Eastern European Renal Cancer case-control study. Associations of individual SNPs (n = 163 with RCC risk were calculated using unconditional logistic regression adjusted for age, sex and study center. Minimum p-value permutation (Min-P tests were used to identify gene regions associated with risk, and haplotypes were evaluated within these genes.The strongest associations with RCC risk were observed for SLC19A1 (P(min-P = 0.03 and MTHFR (P(min-P = 0.13. A haplotype consisting of four SNPs in SLC19A1 (rs12483553, rs2838950, rs2838951, and rs17004785 was associated with a 37% increased risk (p = 0.02, and exploratory stratified analysis suggested the association was only significant among those in the lowest tertile of vegetable intake.To our knowledge, this is the first study to comprehensively examine variation in one-carbon metabolism genes in relation to RCC risk. We identified a novel association with SLC19A1, which is important for transport of folate into cells. Replication in other populations is required to confirm these findings.

  9. Multifactorial comparative proteomic study of cytochrome P450 2E1 function in chronic alcohol administration.

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    Yuan Wang

    Full Text Available With the use of iTRAQ technique, a multifactorial comparative proteomic study can be performed. In this study, to obtain an overview of ethanol, CYP2E1 and gender effects on liver injury and gain more insight into the underlying molecular mechanism, mouse liver proteomes were quantitatively analyzed using iTRAQ under eight conditions including mice of different genders, wild type versus CYP2E1 knockout, and normal versus alcohol diet. A series of statistical and bioinformatic analyses were explored to simplify and clarify multifactorial comparative proteomic data. First, with the Principle Component analysis, six proteins, CYP2E1, FAM25, CA3, BHMT, HIBADH and ECHS1, involved in oxidation reduction, energy and lipid metabolism and amino acid metabolism, were identified as the most differentially expressed gene products across all of the experimental conditions of our chronic alcoholism model. Second, hierarchical clustering analysis showed CYP2E1 knockout played a primary role in the overall differential protein expression compared with ethanol and gender factors. Furthermore, pair-wise multiple comparisons have revealed that the only significant expression difference lied in wild-type and CYP2E1 knockout mice both treated with ethanol. Third, K-mean clustering analysis indicated that the CYP2E1 knockout had the reverse effect on ethanol induced oxidative stress and lipid oxidation. More importantly, IPA analysis of proteomic data inferred that the gene expressions of two upstream regulators, NRF2 and PPARα, regulated by chronic alcohol feeding and CYP2E1 knockout, are involved in ethanol induced oxidative stress and lipid oxidation. The present study provides an effectively comprehensive data analysis strategy to compare multiple biological factors, contributing to biochemical effects of alcohol on the liver. The mass spectrometry proteomics data have been deposited to the ProteomeXchange with data set identifier of PXD000635.

  10. Proteomic Analysis of Stationary Phase in the Marine Bacterium "Candidatus Pelagibacter ubique"

    Energy Technology Data Exchange (ETDEWEB)

    Sowell, S. M.; Norbeck, A. D.; Lipton, M. S.; Nicora, C. D.; Callister, S. J.; Smith, R. D.; Barofsky, D. F.; Giovannoni, S. J.

    2008-05-09

    The α-proteobacterium ‘Candidatus Pelagibacter ubique’ str. HTCC1062, and most other members of the SAR11 clade, lack genes for assimilatory sulfate reduction, making them dependent on organosulfur compounds that occur naturally in seawater. To investigate how these cells adapt to sulfur limitation, batch cultures were grown in defined media containing either limiting or non-limiting amounts of dimethylsulfoniopropionate (DMSP) as the sole sulfur source. Protein and mRNA expression were measured during exponential growth, immediately prior to stationary phase, and in late stationary phase. Two distinct responses were observed: one as DMSP became exhausted, and another as cells acclimated to a sulfur-limited environment. The first response was characterized by increased transcription and translation of all Ca. P. ubique genes downstream of previously confirmed S-adenosyl methionine (SAM) riboswitches: bhmT, mmuM, and metY. Proteins encoded by these genes were up to 33 times more abundant as DMSP became limiting. Their predicted function is to shunt all available sulfur to methionine. The secondary response, observed during sulfur-depleted stationary phase, was a 6-10 fold increase in transcription of the heme c shuttle ccmC and two small genes of unknown function (SAR11_1163 and SAR11_1164). This bacterium's strategy for coping with sulfur stress appears to be intracellular redistribution to support methionine biosynthesis, rather than increasing organosulfur import. Many of the genes and SAM riboswitches involved in this response are located in a hypervariable genome region (HVR). One of these HVR genes, ordL, is located downstream of a conserved motif that evidence suggests is a novel riboswitch.

  11. Genetic Variation in Choline-Metabolizing Enzymes Alters Choline Metabolism in Young Women Consuming Choline Intakes Meeting Current Recommendations

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    Ariel B. Ganz

    2017-01-01

    Full Text Available Single nucleotide polymorphisms (SNPs in choline metabolizing genes are associated with disease risk and greater susceptibility to organ dysfunction under conditions of dietary choline restriction. However, the underlying metabolic signatures of these variants are not well characterized and it is unknown whether genotypic differences persist at recommended choline intakes. Thus, we sought to determine if common genetic risk factors alter choline dynamics in pregnant, lactating, and non-pregnant women consuming choline intakes meeting and exceeding current recommendations. Women (n = 75 consumed 480 or 930 mg choline/day (22% as a metabolic tracer, choline-d9 for 10–12 weeks in a controlled feeding study. Genotyping was performed for eight variant SNPs and genetic differences in metabolic flux and partitioning of plasma choline metabolites were evaluated using stable isotope methodology. CHKA rs10791957, CHDH rs9001, CHDH rs12676, PEMT rs4646343, PEMT rs7946, FMO3 rs2266782, SLC44A1 rs7873937, and SLC44A1 rs3199966 altered the use of choline as a methyl donor; CHDH rs9001 and BHMT rs3733890 altered the partitioning of dietary choline between betaine and phosphatidylcholine synthesis via the cytidine diphosphate (CDP-choline pathway; and CHKA rs10791957, CHDH rs12676, PEMT rs4646343, PEMT rs7946 and SLC44A1 rs7873937 altered the distribution of dietary choline between the CDP-choline and phosphatidylethanolamine N-methyltransferase (PEMT denovo pathway. Such metabolic differences may contribute to disease pathogenesis and prognosis over the long-term.

  12. Novel Tetra-Primer ARMS-PCR Assays for Thiopurine Intolerance Susceptibility Mutations NUDT15 c.415C>T and TPMT c.719A>G (TPMT*3C in East Asians

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    Chi-Chun Ho

    2017-10-01

    Full Text Available Thiopurines are clinically useful in the management of diverse immunological and malignant conditions. Nevertheless, these purine analogues can cause lethal myelosuppression, which may be prevented by prospective testing for variants in the thiopurine S-methyltransferase (TPMT and, in East Asians, Nudix hydrolase 15 (NUDT15 genes. Two single-tube, tetra-primer amplification refractory mutation system polymerase chain reaction (ARMS-PCR assays were developed to genotype the common loss-of-function variants NUDT15 c.415C>T (rs116855232 and TPMT*3C c.719A>G (rs1142345. In a group of 60 unselected patients, one and seven were found to be homozygous and heterozygous, respectively, for NUDT15 c.415C>T; one was found to be heterozygous for TPMT*3C c.719A>G. There was no non-specific amplification, and the genotypes were 100% concordant with Sanger sequencing. Limit-of-detection for both assays was below 1 ng of heterozygous template per reaction. Time- and cost-effective ARMS-PCR assays, suitable for genotyping East-Asian patients for thiopurine intolerance, were successfully developed and validated.

  13. Enhanced specificity of TPMT*2 genotyping using unidirectional wild-type and mutant allele-specific scorpion primers in a single tube.

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    Dong Chen

    Full Text Available Genotyping of thiopurine S-methyltransferase (TPMT is recommended for predicting the adverse drug response of thiopurines. In the current study, a novel version of allele-specific PCR (AS-PCR, termed competitive real-time fluorescent AS-PCR (CRAS-PCR was developed to analyze the TPMT*2 genotype in ethnic Chinese. This technique simultaneously uses wild-type and mutant allele-specific scorpion primers in a single reaction. To determine the optimal conditions for both traditional AS-PCR and CRAS-PCR, we used the Taguchi method, an engineering optimization process that balances the concentrations of all components using an orthogonal array rather than a factorial array. Instead of running up to 264 experiments with the conventional factorial method, the Taguchi method achieved the same optimization using only 16 experiments. The optimized CRAS-PCR system completely avoided non-specific amplification occurring in traditional AS-PCR and could be performed at much more relaxed reaction conditions at 1% sensitivity, similar to traditional AS-PCR. TPMT*2 genotyping of 240 clinical samples was consistent with published data. In conclusion, CRAS-PCR is a novel and robust genotyping method, and the Taguchi method is an effective tool for the optimization of molecular analysis techniques.

  14. TPMT genetic variants are associated with increased rejection with azathioprine use in heart transplantation.

    Science.gov (United States)

    Liang, Jackson J; Geske, Jennifer R; Boilson, Barry A; Frantz, Robert P; Edwards, Brooks S; Kushwaha, Sudhir S; Kremers, Walter K; Weinshilboum, Richard M; Pereira, Naveen L

    2013-12-01

    Azathioprine (AZA) is an important immunosuppressant drug used in heart transplantation (HTX). Consensus guidelines recommend that patients with thiopurine S-methyltransferase (TPMT) genetic variants be started on lower AZA dose because of higher active metabolite levels and risk of adverse events. However, in-vitro lymphocyte proliferation assays performed in participants with inactive TPMT alleles have suggested that AZA use may result in decreased immunosuppressant efficacy as compared with wild-type (WT) individuals. The objective of this study was therefore to determine the effect of TPMT genetic variation on AZA efficacy or prevention of rejection in HTX recipients treated with AZA. We genotyped 93 HTX recipients treated with AZA and measured erythrocyte TPMT enzyme activity. Acute rejection was monitored by routine endomyocardial biopsies. There were 83 WT and 10 heterozygote (HZ) HTX recipients. TPMT activity level was lower in HZ compared with WT (13.1±2.8 vs. 21±4.5 U/ml red blood cell, Prejection earlier (Prejection score was higher (P=0.02) than WT. AZA was discontinued more frequently in HZ (P=0.01) because of rejection. The incidence of leukopenia was similar between the groups (40 vs. 43%, P=1.0). HTX recipients with TPMT genetic variant alleles who are treated with AZA develop acute rejection earlier, more frequently, and of greater severity. These patients, despite having lower TPMT enzymatic activity, should be monitored carefully for possible increased risk of acute rejection.

  15. Transferability of microsatellite markers located in candidate genes for wood properties between Eucalyptus species

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    Cintia V. Acuña

    2014-12-01

    Full Text Available Aim of study:  To analyze the feasibility of extrapolating conclusions on wood quality genetic control between different Eucalyptus species, particularly from species with better genomic information, to those less characterized. For this purpose, the first step is to analyze the conservation and cross-transferability of microsatellites markers (SSRs located in candidate genes.Area of study: Eucalyptus species implanted in Argentina coming from different Australian origins.Materials and methods: Twelve validated and polymorphic SSRs in candidate genes (SSR-CGs for wood quality in E. globulus were selected for cross species amplification in six species: E. grandis, E. saligna, E. dunnii, E. viminalis, E. camaldulensis and E. tereticornis.Main results: High cross-species transferability (92% to 100% was found for the 12 polymorphic SSRs detected in E. globulus. These markers revealed allelic diversity in nine important candidate genes: cinnamoyl CoA reductase (CCR, cellulose synthase 3 (CesA3, the transcription factor LIM1, homocysteine S-methyltransferase (HMT, shikimate kinase (SK, xyloglucan endotransglycosylase 2 (XTH2, glutathione S-transferase (GST, glutamate decarboxylase (GAD and peroxidase (PER.Research highlights: The markers described are potentially suitable for comparative QTL mapping, molecular marker assisted breeding (MAB and for population genetic studies across different species within the subgenus Symphyomyrtus.Keywords: validation; cross-transferability; SSR; functional markers; eucalypts; Symphyomyrtus.

  16. Structural, mechanistic and functional insight into gliotoxin bis-thiomethylation in Aspergillus fumigatus.

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    Dolan, Stephen K; Bock, Tobias; Hering, Vanessa; Owens, Rebecca A; Jones, Gary W; Blankenfeldt, Wulf; Doyle, Sean

    2017-02-01

    Gliotoxin is an epipolythiodioxopiperazine (ETP) class toxin, contains a disulfide bridge that mediates its toxic effects via redox cycling and is produced by the opportunistic fungal pathogen Aspergillus fumigatus Self-resistance against gliotoxin is effected by the gliotoxin oxidase GliT, and attenuation of gliotoxin biosynthesis is catalysed by gliotoxin S -methyltransferase GtmA. Here we describe the X-ray crystal structures of GtmA-apo (1.66 Å), GtmA complexed to S -adenosylhomocysteine (1.33 Å) and GtmA complexed to S -adenosylmethionine (2.28 Å), providing mechanistic insights into this important biotransformation. We further reveal that simultaneous elimination of the ability of A. fumigatus to dissipate highly reactive dithiol gliotoxin, via deletion of GliT and GtmA, results in the most significant hypersensitivity to exogenous gliotoxin observed to date. Indeed, quantitative proteomic analysis of Δ gliT ::Δ gtmA reveals an uncontrolled over-activation of the gli -cluster upon gliotoxin exposure. The data presented herein reveal, for the first time, the extreme risk associated with intracellular dithiol gliotoxin biosynthesis-in the absence of an efficient dismutation capacity. Significantly, a previously concealed protective role for GtmA and functionality of ETP bis -thiomethylation as an ancestral protection strategy against dithiol compounds is now evident. © 2017 The Authors.

  17. Crystal structure of the homocysteine methyltransferase MmuM from Escherichia coli.

    Science.gov (United States)

    Li, Kunhua; Li, Gengnan; Bradbury, Louis M T; Hanson, Andrew D; Bruner, Steven D

    2016-02-01

    Homocysteine S-methyltransferases (HMTs, EC 2.1.1.0) catalyse the conversion of homocysteine to methionine using S-methylmethionine or S-adenosylmethionine as the methyl donor. HMTs play an important role in methionine biosynthesis and are widely distributed among micro-organisms, plants and animals. Additionally, HMTs play a role in metabolite repair of S-adenosylmethionine by removing an inactive diastereomer from the pool. The mmuM gene product from Escherichia coli is an archetypal HMT family protein and contains a predicted zinc-binding motif in the enzyme active site. In the present study, we demonstrate X-ray structures for MmuM in oxidized, apo and metallated forms, representing the first such structures for any member of the HMT family. The structures reveal a metal/substrate-binding pocket distinct from those in related enzymes. The presented structure analysis and modelling of co-substrate interactions provide valuable insight into the function of MmuM in both methionine biosynthesis and cofactor repair. © 2016 Authors; published by Portland Press Limited.

  18. Advances and challenges in hereditary cancer pharmacogenetics.

    Science.gov (United States)

    Cascorbi, Ingolf; Werk, Anneke Nina

    2017-01-01

    Cancer pharmacogenetics usually considers tumor-specific targets. However, hereditary genetic variants may interfere with the pharmacokinetics of antimetabolites and other anti-cancer drugs, which may lead to severe adverse events. Areas covered: Here, the impact of hereditary genes considered in drug labels such as thiopurine S-methyltransferase (TPMT), UDP-glucuronosyltransferase 1A1 (UTG1A1) and dihydropyrimidine dehydrogenase (DPYD) are discussed with respect to guidelines of the Clinical Pharmacogenetics Implementation Consortium (CPIC). Moreover, the association between genetic variants of drug transporters with the clinical outcome is comprehensively discussed. Expert opinion: Precision therapy in the field of oncology is developing tremendously. There are a number of somatic tumor genetic markers that are indicative for treatment with anti-cancer drugs. By contrast, for some hereditary variants, recommendations have been developed. Although we have vast knowledge on the association between drug transporter variants and clinical outcome, the overall data is inconsistent and the predictability of the related phenotype is low. Further developments in research may lead to the discovery of rare, but functionally relevant single nucleotide polymorphisms and a better understanding of multiple genomic, epigenomic as well as phenotypic factors, contributing to drug response in malignancies.

  19. S-methylmethionine conversion to dimethylsulfoniopropionate: evidence for an unusual transamination reaction

    International Nuclear Information System (INIS)

    Rhodes, D.; Gage, D.A.; Cooper, A.J.L.; Hanson, A.D.

    1997-01-01

    Leaves of Wollastonia biflora (L.) DC. synthesize the osmoprotectant 3-dimethylsulfoniopropionate (DMSP) from methionine via S-methylmethionine (SMM) and 3-dimethylsulfoniopropionaldehyde (DMSP-ald); no other intermediates have been detected. To test whether the amino group of SMM is lost by transamination or deamination, [methyl-2H3, 15N]SMM was supplied to leaf discs, and 15N-labeling of amino acids was monitored, along with synthesis of [2H3]DMSP. After short incubations more 15N was incorporated into glutamate than into other amino acids, and the 15N abundance in glutamate exceeded that in the amide group of glutamine (Gln). This is more consistent with transamination than deamination, because deamination would be predicted to give greater labeling of Gln amide N due to reassimilation, via Gln synthetase, of the 15NH4+ released. This prediction was borne out by control experiments with 15NH4Cl. The transamination product of SMM, 4-dimethylsulfonio-2-oxobutyrate (DMSOB), is expected to be extremely unstable. This was confirmed by attempting to synthesize it enzymatically from SMM using L-amino acid oxidase or Gln transaminase K and from 4-methylthio-2-oxobutyrate using methionine S-methyltransferase. In each case, the reaction product decomposed rapidly, releasing dimethylsulfide. The conversion of SMM to DMSP-ald is therefore unlikely to involve a simple transamination that generates free DMSOB. Plausible alternatives are that DMSOB is channeled within a specialized transaminase-decarboxylase complex or that it exists only as the bound intermediate of a single enzyme catalyzing an unusual transamination-decarboxylation reaction

  20. Fine-scale genetic mapping of a hybrid sterility factor between Drosophila simulans and D. mauritiana: the varied and elusive functions of "speciation genes"

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    Lemos Bernardo

    2010-12-01

    Full Text Available Abstract Background Hybrid male sterility (HMS is a usual outcome of hybridization between closely related animal species. It arises because interactions between alleles that are functional within one species may be disrupted in hybrids. The identification of genes leading to hybrid sterility is of great interest for understanding the evolutionary process of speciation. In the current work we used marked P-element insertions as dominant markers to efficiently locate one genetic factor causing a severe reduction in fertility in hybrid males of Drosophila simulans and D. mauritiana. Results Our mapping effort identified a region of 9 kb on chromosome 3, containing three complete and one partial coding sequences. Within this region, two annotated genes are suggested as candidates for the HMS factor, based on the comparative molecular characterization and public-source information. Gene Taf1 is partially contained in the region, but yet shows high polymorphism with four fixed non-synonymous substitutions between the two species. Its molecular functions involve sequence-specific DNA binding and transcription factor activity. Gene agt is a small, intronless gene, whose molecular function is annotated as methylated-DNA-protein-cysteine S-methyltransferase activity. High polymorphism and one fixed non-synonymous substitution suggest this is a fast evolving gene. The gene trees of both genes perfectly separate D. simulans and D. mauritiana into monophyletic groups. Analysis of gene expression using microarray revealed trends that were similar to those previously found in comparisons between whole-genome hybrids and parental species. Conclusions The identification following confirmation of the HMS candidate gene will add another case study leading to understanding the evolutionary process of hybrid incompatibility.

  1. Effects of brevetoxin exposure on the immune system of loggerhead sea turtles.

    Science.gov (United States)

    Walsh, Catherine J; Leggett, Stephanie R; Carter, Barbara J; Colle, Clarence

    2010-05-10

    Blooms of the toxic dinoflagellate, Karenia brevis, occur almost annually off the Florida coast. These blooms, commonly called "red tides", produce a group of neurotoxins collectively termed brevetoxins. Many species of sealife, including sea turtles, are severely impacted by brevetoxin exposure. Effects of brevetoxins on immune cells were investigated in rescued loggerhead sea turtles, Caretta caretta, as well as through in vitro experiments using peripheral blood leukocytes (PBL) collected from captive sea turtles. In rescued animals, plasma brevetoxin concentrations were measured using a competitive ELISA. Plasma lysozyme activity was measured using a turbidity assay. Lysozyme activity correlated positively with plasma brevetoxin concentrations. Differential expression of genes affected by brevetoxin exposure was determined using two separate suppression subtractive hybridization experiments. In one experiment, genes from PBL collected from sea turtles rescued from red tide toxin exposure were compared to genes from PBL collected from healthy captive loggerhead sea turtles. In the second experiment, PBL from healthy captive loggerhead sea turtles were exposed to brevetoxin (500 ng PbTx-2/ml) in vitro for 18 h and compared to unexposed PBL. Results from the subtraction hybridization experiment conducted with red tide rescued sea turtle PBL indicated that genes involved in oxidative stress or xenobiotic metabolism were up-regulated. Using quantitative real-time PCR, a greater than 2-fold increase in superoxide dismutase and thioredoxin and greater than 10-fold increase in expression of thiopurine S-methyltransferase were observed. Results from the in vitro subtraction hybridization experiment indicated that genes coding for cytochrome c oxidases were the major up-regulated genes. Using quantitative real-time PCR, a greater than 8-fold increase in expression of beta-tubulin and greater than 3-fold increase in expression of ubiquinol were observed. Brevetoxin

  2. Comparison of Direct Sequencing, Real-Time PCR-High Resolution Melt (PCR-HRM) and PCR-Restriction Fragment Length Polymorphism (PCR-RFLP) Analysis for Genotyping of Common Thiopurine Intolerant Variant Alleles NUDT15 c.415C>T and TPMT c.719A>G (TPMT*3C).

    Science.gov (United States)

    Fong, Wai-Ying; Ho, Chi-Chun; Poon, Wing-Tat

    2017-05-12

    Thiopurine intolerance and treatment-related toxicity, such as fatal myelosuppression, is related to non-function genetic variants encoding thiopurine S-methyltransferase (TPMT) and Nudix hydrolase 15 (NUDT15). Genetic testing of the common variants NUDT15:NM_018283.2:c.415C>T (Arg139Cys, dbSNP rs116855232 T allele) and TPMT: NM_000367.4:c.719A>G (TPMT*3C, dbSNP rs1142345 G allele) in East Asians including Chinese can potentially prevent treatment-related complications. Two complementary genotyping approaches, real-time PCR-high resolution melt (PCR-HRM) and PCR-restriction fragment length morphism (PCR-RFLP) analysis were evaluated using conventional PCR and Sanger sequencing genotyping as the gold standard. Sixty patient samples were tested, revealing seven patients (11.7%) heterozygous for NUDT15 c.415C>T, one patient homozygous for the variant and one patient heterozygous for the TPMT*3C non-function allele. No patient was found to harbor both variants. In total, nine out of 60 (15%) patients tested had genotypic evidence of thiopurine intolerance, which may require dosage adjustment or alternative medication should they be started on azathioprine, mercaptopurine or thioguanine. The two newly developed assays were more efficient and showed complete concordance (60/60, 100%) compared to the Sanger sequencing results. Accurate and cost-effective genotyping assays by real-time PCR-HRM and PCR-RFLP for NUDT15 c.415C>T and TPMT*3C were successfully developed. Further studies may establish their roles in genotype-informed clinical decision-making in the prevention of morbidity and mortality due to thiopurine intolerance.

  3. Evidence for functional convergence in genes upregulated by herbivores ingesting plant secondary compounds

    Science.gov (United States)

    2014-01-01

    Background Nearly 40 years ago, Freeland and Janzen predicted that liver biotransformation enzymes dictated diet selection by herbivores. Despite decades of research on model species and humans, little is known about the biotransformation mechanisms used by mammalian herbivores to metabolize plant secondary compounds (PSCs). We investigated the independent evolution of PSC biotransformation mechanisms by capitalizing on a dramatic diet change event—the dietary inclusion of creosote bush (Larrea tridentata)—that occurred in the recent evolutionary history of two species of woodrats (Neotoma lepida and N. bryanti). Results By comparing gene expression profiles of two populations of woodrats with evolutionary experience to creosote and one population naïve to creosote, we identified genes either induced by a diet containing creosote PSCs or constitutively higher in populations with evolutionary experience of creosote. Although only one detoxification gene (an aldo-keto reductase) was induced by both experienced populations, these populations converged upon functionally equivalent strategies to biotransform the PSCs of creosote bush by constitutively expressing aldehyde and alcohol dehydrogenases, Cytochromes P450s, methyltransferases, glutathione S-transferases and sulfotransferases. The response of the naïve woodrat population to creosote bush was indicative of extreme physiological stress. Conclusions The hepatic detoxification system of mammals is notoriously complex, with hundreds of known biotransformation enzymes. The comparison herein of woodrat taxa that differ in evolutionary and ecological experience with toxins in creosote bush reveals convergence in the overall strategies used by independent species after a historical shift in diet. In addition, remarkably few genes seemed to be important in this dietary shift. The research lays the requisite groundwork for future studies of specific biotransformation pathways used by woodrats to metabolize the

  4. Assessment of mercaptopurine (6MP) metabolites and 6MP metabolic key-enzymes in childhood acute lymphoblastic leukemia.

    Science.gov (United States)

    Wojtuszkiewicz, Anna; Barcelos, Ana; Dubbelman, Boas; De Abreu, Ronney; Brouwer, Connie; Bökkerink, Jos P; de Haas, Valerie; de Groot-Kruseman, Hester; Jansen, Gerrit; Kaspers, Gertjan L; Cloos, Jacqueline; Peters, G J

    2014-01-01

    Pediatric acute lymphoblastic leukemia (ALL) is treated with combination chemotherapy including mercaptopurine (6MP) as an important component. Upon its uptake, 6MP undergoes a complex metabolism involving many enzymes and active products. The prognostic value of all the factors engaged in this pathway still remains unclear. This study attempted to determine which components of 6MP metabolism in leukemic blasts and red blood cells are important for 6MP's sensitivity and toxicity. In addition, changes in the enzymatic activities and metabolite levels during the treatment were analyzed. In a cohort (N=236) of pediatric ALL patients enrolled in the Dutch ALL-9 protocol, we studied the enzymes inosine-5'-monophosphate dehydrogenase (IMPDH), thiopurine S-methyltransferase (TPMT), hypoxanthine guanine phosphoribosyl transferase (HGPRT), and purine nucleoside phosphorylase (PNP) as well as thioguanine nucleotides (TGN) and methylthioinosine nucleotides (meTINs). Activities of selected enzymes and levels of 6MP derivatives were measured at various time points during the course of therapy. The data obtained and the toxicity related parameters available for these patients were correlated with each other. We found several interesting relations, including high concentrations of two active forms of 6MP--TGN and meTIN--showing a trend toward association with better in vitro antileukemic effect of 6MP. High concentrations of TGN and elevated activity of HGPRT were found to be significantly associated with grade III/IV leucopenia. However, a lot of data of enzymatic activities and metabolite concentrations as well as clinical toxicity were missing, thereby limiting the number of assessed relations. Therefore, although a complex study of 6MP metabolism in ALL patients is feasible, it warrants more robust and strict data collection in order to be able to draw more reliable conclusions.

  5. Fine-scale genetic mapping of a hybrid sterility factor between Drosophila simulans and D. mauritiana: the varied and elusive functions of "speciation genes".

    Science.gov (United States)

    Araripe, Luciana O; Montenegro, Horácio; Lemos, Bernardo; Hartl, Daniel L

    2010-12-14

    Hybrid male sterility (HMS) is a usual outcome of hybridization between closely related animal species. It arises because interactions between alleles that are functional within one species may be disrupted in hybrids. The identification of genes leading to hybrid sterility is of great interest for understanding the evolutionary process of speciation. In the current work we used marked P-element insertions as dominant markers to efficiently locate one genetic factor causing a severe reduction in fertility in hybrid males of Drosophila simulans and D. mauritiana. Our mapping effort identified a region of 9 kb on chromosome 3, containing three complete and one partial coding sequences. Within this region, two annotated genes are suggested as candidates for the HMS factor, based on the comparative molecular characterization and public-source information. Gene Taf1 is partially contained in the region, but yet shows high polymorphism with four fixed non-synonymous substitutions between the two species. Its molecular functions involve sequence-specific DNA binding and transcription factor activity. Gene agt is a small, intronless gene, whose molecular function is annotated as methylated-DNA-protein-cysteine S-methyltransferase activity. High polymorphism and one fixed non-synonymous substitution suggest this is a fast evolving gene. The gene trees of both genes perfectly separate D. simulans and D. mauritiana into monophyletic groups. Analysis of gene expression using microarray revealed trends that were similar to those previously found in comparisons between whole-genome hybrids and parental species. The identification following confirmation of the HMS candidate gene will add another case study leading to understanding the evolutionary process of hybrid incompatibility.

  6. Pharmacoeconomic evaluations of pharmacogenetic and genomic screening programmes: a systematic review on content and adherence to guidelines.

    Science.gov (United States)

    Vegter, Stefan; Boersma, Cornelis; Rozenbaum, Mark; Wilffert, Bob; Navis, Gerjan; Postma, Maarten J

    2008-01-01

    The fields of pharmacogenetics and pharmacogenomics have become important practical tools to progress goals in medical and pharmaceutical research and development. As more screening tests are being developed, with some already used in clinical practice, consideration of cost-effectiveness implications is important. A systematic review was performed on the content of and adherence to pharmacoeconomic guidelines of recent pharmacoeconomic analyses performed in the field of pharmacogenetics and pharmacogenomics. Economic analyses of screening strategies for genetic variations, which were evidence-based and assumed to be associated with drug efficacy or safety, were included in the review. The 20 papers included cover a variety of healthcare issues, including screening tests on several cytochrome P450 (CYP) enzyme genes, thiopurine S-methyltransferase (TMPT) and angiotensin-converting enzyme (ACE) insertion deletion (ACE I/D) polymorphisms. Most economic analyses reported that genetic screening was cost effective and often even clearly dominated existing non-screening strategies. However, we found a lack of standardization regarding aspects such as the perspective of the analysis, factors included in the sensitivity analysis and the applied discount rates. In particular, an important limitation of several studies related to the failure to provide a sufficient evidence-based rationale for an association between genotype and phenotype. Future economic analyses should be conducted utilizing correct methods, with adherence to guidelines and including extensive sensitivity analyses. Most importantly, genetic screening strategies should be based on good evidence-based rationales. For these goals, we provide a list of recommendations for good pharmacoeconomic practice deemed useful in the fields of pharmacogenetics and pharmacogenomics, regardless of country and origin of the economic analysis.