Dataset Information

Betaine-homocysteine methyltransferase: human liver genotype-phenotype correlation.

ABSTRACT: Betaine-homocysteine methyltransferase (BHMT) catalyzes the remethylation of homocysteine. BHMT is highly expressed in the human liver. In the liver, BHMT catalyzes up to 50% of homocysteine metabolism. Understanding the relationship between BHMT genetic polymorphisms and function might increase our understanding of the role of this reaction in homocysteine remethylation and in S-adenosylmethionine-dependent methylation. To help achieve those goals, we measured levels of BHMT enzyme activity and immunoreactive protein in 268 human hepatic surgical biopsy samples from adult subjects as well as 73 fetal hepatic tissue samples obtained at different gestational ages. BHMT protein levels were correlated significantly (p<0.001) with levels of enzyme activity in both fetal and adult tissues, but both were decreased in fetal tissue when compared with levels in the adult hepatic biopsies. To determine possible genotype-phenotype correlations, 12 tag SNPs for BHMT and the closely related BHMT2 gene were selected from SNPs observed during our own gene resequencing studies as well as from HapMap. These SNPs data were used to genotype DNA from the adult hepatic surgical biopsy samples, and genotype-phenotype association analysis was performed. Three SNPs (rs41272270, rs16876512, and rs6875201), located 28kb upstream, in the 5'-UTR and in intron 1 of BHMT, respectively, were significantly correlated with both BHMT activity (p=3.41E-8, 2.55E-9 and 2.46E-10, respectively) and protein levels (p=5.78E-5, 1.08E-5 and 6.92E-6, respectively). We also imputed 230 additional SNPs across the BHMT and BHMT2 genes, identifying an additional imputed SNP, rs7700790, that was also highly associated with hepatic BHMT enzyme activity and protein. However, none of the 3 genotyped or one imputed SNPs displayed a "shift" during electrophoretic mobility shift assays. These observations may help us to understand individual variation in the regulation of BHMT in the human liver and its possible relationship to variation in methylation.

SUBMITTER: Feng Q

PROVIDER: S-EPMC3053054 | biostudies-literature | 2011 Feb

REPOSITORIES: biostudies-literature

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Betaine-homocysteine methyltransferase: human liver genotype-phenotype correlation.

Feng Qiping Q Kalari Krishna K Fridley Brooke L BL Jenkins Gregory G Ji Yuan Y Abo Ryan R Hebbring Scott S Zhang Jianping J Nye Monica D MD Leeder J Steven JS Weinshilboum Richard M RM

Molecular genetics and metabolism 20101021 2

Betaine-homocysteine methyltransferase (BHMT) catalyzes the remethylation of homocysteine. BHMT is highly expressed in the human liver. In the liver, BHMT catalyzes up to 50% of homocysteine metabolism. Understanding the relationship between BHMT genetic polymorphisms and function might increase our understanding of the role of this reaction in homocysteine remethylation and in S-adenosylmethionine-dependent methylation. To help achieve those goals, we measured levels of BHMT enzyme activity and ...[more]

PMID: 21093336

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Project description:Betaine-homocysteine S-methyltransferase (BHMT) transfers a methyl group from betaine to Hcy to form DMG (dimethylglycine) and Met. The reaction is ordered Bi Bi; Hcy is the first substrate to bind and Met is the last product off. Using intrinsic tryptophan fluorescence [Castro, Gratson, Evans, Jiracek, Collinsova, Ludwig and Garrow (2004) Biochemistry 43, 5341-5351], it was shown that BHMT exists in three steady-state conformations: enzyme alone, enzyme plus occupancy at the first substrate-binding site (Hcy or Met), or enzyme plus occupancy at both substrate-binding sites (Hcy plus betaine, or Hcy plus DMG). Betaine or DMG alone do not bind to the enzyme, indicating that the conformational change associated with Hcy binding creates the betaine-binding site. CBHcy [S-(d-carboxybutyl)-D,L-homocysteine] is a bisubstrate analogue that causes BHMT to adopt the same conformation as the ternary complexes. We report that BHMT is susceptible to conformation-dependent oxidative inactivation. Two oxidants, MMTS (methyl methanethiosulphonate) and hydrogen peroxide (H2O2), cause a loss of the enzyme's catalytic Zn (Zn2+ ion) and a correlative loss of activity. Addition of 2-mercaptoethanol and exogenous Zn after MMTS treatment restores activity, but oxidation due to H2O2 is irreversible. CD and glutaraldehyde cross-linking indicate that H2O2 treatment causes small perturbations in secondary structure but no change in quaternary structure. Oxidation is attenuated when both binding sites are occupied by CBHcy, but Met alone has no effect. Partial digestion of ligand-free BHMT with trypsin produces two large peptides, excising a seven-residue peptide within loop L2. CBHcy but not Met binding slows down proteolysis by trypsin. These findings suggest that L2 is involved in the conformational change associated with occupancy at the betaine-binding site and that this conformational change and/or occupancy at both ligand-binding sites protect the enzyme from oxidative inactivation.

Dataset Information

Betaine-homocysteine methyltransferase: human liver genotype-phenotype correlation.

Publications

Betaine-homocysteine methyltransferase: human liver genotype-phenotype correlation.

Similar Datasets

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