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Molecular characterization of methylenetetrahydrofolate reductase deficiencyGoyette, Philippe. January 1997 (has links)
No description available.
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Investigation of mutations in methylenetetrahydrofolate reductase deficiencyLow-Nang, Lawrence January 1991 (has links)
Methylenetetrahydrofolate reductase (MTHFR) reduces 5,10-methylene THF to 5-methyl THF, the carbon donor for the methylation of homocysteine to methionine. Patients with severe MTHFR deficiency (MRD) have neurologic abnormalities while a milder form (a thermolabile MTHFR variant) has been shown to be associated with coronary artery disease (CAD). Ten MRD patients, with reduced or non-detectable activity, were studied to characterize the nature of the mutation. Southern, Northern and Western analysis did not reveal any defects in the patients. These results suggest that the mutations may be minor insertions/deletions or single base substitutions that affect catalytic activity. Single strand conformation polymorphism (SSCP) analysis was used to detect base substitutions; 3 RFLPs were identified with this protocol. One was in the coding region (SphI) while the other two were in the 3$ sp prime$ untranslated region (MaeIII and MnlI). A difference in frequency of the SphI RFLP was found between control subjects and a small sample of CAD patients whose homocysteine levels were greater than the 99th percentile.
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Characterization of the 5' region of the human methylenetetrahydrofolate reductase (MTHFR) geneChan, Manuel January 1999 (has links)
Methylenetetrahydrofolate reductase (MTHFR) catalyses the reduction of 5, 10-methylenetetrahydrofolate to 5-methyltetrahydrofolate, a methyl donor for the re-methylation of homocysteine to methionine. A thermolabile variant of this enzyme, present in approximately 35% of alleles in the North American population, has been associated with cardiovascular disease, neural tube defects, and colon cancer. A cDNA of 2.2kb for human MTHFR has been expressed and results in an active enzyme, but the cDNA and genomic sequences 5' to the ATG start site have not been adequately investigated. The characterization of the 5' region of the human MTHFR gene is reported here. Four additional 5' exonic sequences were localized to a 4kb genomic fragment. The original exon 1 extends directly upstream into a 5' UTR. Three other 5' exons (two with open reading frames) are alternatively spliced into a common splice acceptor site, generating cDNAs with 4 possible 5' ends. The N-terminal peptide sequence of the porcine MTHFR has not been identified in the human sequence suggesting that the missing human coding sequence might be localized further upstream or not conserved across species. A putative chloride ion channel gene (ClC-6) was located in the opposite orientation, at 3.5kb upstream of the original ATG codon, suggesting an overlap with the MTHFR gene and potential co-localization of regulatory elements. A CpG island was identified in the region of a 5' exon (43S) suggesting that a transcription start site and a promoter might be nearby. This work is relevant in understanding the regulation of this important enzyme in folate metabolism.
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Molecular characterization of methylenetetrahydrofolate reductase deficiencyGoyette, Philippe. January 1997 (has links)
Methylenetetrahydrofolate reductase (MTHFR) catalyses the conversion of 5, 10-methylenctetrahydrofolate to 5-methyltetrahydrofolate, co-substrate of methionine synthesis. Two types of deficiency have been described for MTHFR. Severe MTHFR deficiency, associated with severe hyperhomocysteinemia and homocystinuria, shows levels of MTHFR activity below 20% of control values. This deficiency has a variable age of onset and shows a wide range of neurological and vascular defects. Mild MTHFR deficiency, with ≈50% enzyme activity and marked enzyme thermolability, has been proposed as a genetic factor in the development of mild hyperhomocysteinemia, a condition associated with neural tube defects and premature vascular disease. / The goal of this thesis was to determine the molecular basis for severe MTHFR deficiency. In order to study this, I isolated a 1.2 Kb partial cDNA encoding human MTHFR I determined that its primary amino acid sequence is homologous to the bacterial enzyme, and encodes the N-terminal catalytic domain of MTHFR. The cDNA was used to isolate a full length 2.27 Kb cDNA, to map the locus to chromosome position 1p36.3, and to isolate genomic clones for human and mouse MTHFR. I characterized the mouse cDNA sequence, as well as the gene structure for both human and mouse genes. I observed 90% identity at the amino acid level, almost identical sizes of exons and location of introns, and similar sizes of introns. The exon sizes ranged from 102bp to 432bp, and intron sizes varied from 250bp to 4.2Kb. / I identified 13 mutations in severe MTHFR deficiency: 10 missense mutations, 1 nonsense mutation, and 2 splicing defects. I determined that a previously-identified common variant (an Ala →Val mutation) was causative of thermolability in severe MTHFR deficiency. I showed a correlation between genotype, residual activity and phenotype in this disease. I also correlated the presence of another genetic defect, Factor V Leiden mutation, with the possible development of thrombo-embolic events in MTHFR deficiency patients. Finally, I analyzed 8 MTHFR mutations in a bacterial expression system. I determined that 4 of these caused significant reduction of activity (below 20% of control). / This thesis contains the first reports of genetic defects in folate metabolism, and a review of available data in severe MTHFR deficiency.
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Investigation of mutations in methylenetetrahydrofolate reductase deficiencyLow-Nang, Lawrence January 1991 (has links)
No description available.
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Characterization of the 5' region of the human methylenetetrahydrofolate reductase (MTHFR) geneChan, Manuel January 1999 (has links)
No description available.
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Evaluation of common polymorphisms in methylenetetrahydrofolate reductase (MTHFR) and betaine-homocysteine methyltransferase (BHMT)Weisberg, Ilan S. January 1999 (has links)
No description available.
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Evaluation of common polymorphisms in methylenetetrahydrofolate reductase (MTHFR) and betaine-homocysteine methyltransferase (BHMT)Weisberg, Ilan S. January 1999 (has links)
Methylenetetrahydrofolate reductase (MTHFR) catalyses the conversion of 5,10-methylenetetrahydrofolate to 5-methyltetrahydrofolate, a carbon donor for the remethylation of homocysteine to methionine. A common mutation in MTHFR, 677C → T, which converts an alanine (A) to a valine (V) residue, is associated with increased plasma homocysteine and, consequently, is a risk factor for several vasculopathies. Recently, a second common mutation in MTHFR was reported (1298A → C) which converts a glutamate (E) to an alanine (A) residue in the C-terminal regulatory region. Thus study was designed to assess the in vitro and in vivo impact of this polymorphism. Site-directed mutagenesis of the human MTHFR cDNA was performed to create all four combinations of the two common MTHFR polymorphisms. Enzyme activity was assessed in a bacterial expression system. / Since the 1298A → C polymorphism is due to a mutation in the regulatory domain, a parallel study was performed involving the complete deletion of this region. / For in vivo assessment of the 1298A → C variant, we examined this mutation in three clinical populations. / Betaine-homocysteine methyltransferase (BHMT) is a second remethylation enzyme that converts betaine and homocysteine to dimethylglycine and methionine, respectively. This work represents the first report of sequence variants in the newly-cloned BHMT gene. Using SSCP analysis, three common base changes were identified in a panel of healthy controls. (Abstract shortened by UMI.)
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No association between folate intake, pregnancy outcome and single nucleotide polymorphisms in the MTHFR gene in women with unexplained infertilityHoas, Annica January 2013 (has links)
Folate has a direct effect on DNA synthesis which is a major factor in embryo development. In 10% of all infertility couples, there is no apparent cause of infertility. One theory is that folate metabolism is affected in these couples and that it is a possible cause of infertility. This study has analyzed three single nucleotide polymorphisms in the gene MTHFR - rs1801131, rs1801133 and rs2274976. Evaluation of the relationship between folate intake, gene variations and pregnancy outcomes were also performed. For the analysis, blood samples from women diagnosed with infertility (n = 297) and a control group of fertile women (n = 193) was used. Genotyping was performed by using TaqMan SNP genotyping assays. The results showed no significant group difference in genotype or allele frequencies. In addition, there were no differences between genotypes and pregnancy outcomes, or between folate intake and pregnancy outcome. Among the group of women with diagnosed infertility who had children (through IVF), there was more women who had low folate levels (n = 39) than women with high folate levels (n = 28), but the difference was not significant. These studies suggest that disturbances in folate pathway are not an explanation for unexplained infertility.
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Isolation and partial characterization of the mouse gene for methylenetetrahydrofolate reductase (MTHFR)Pai, Aditya P. January 1995 (has links)
Methylenetetrahydrofolate reductase (MTHFR), an important enzyme in folate metabolism, mediates the conversion of 5,10-methylenetetrahydrofolate to 5-methyltetrahydrofolate which serves as the carbon donor for the conversion of homocysteine to methionine. It is also inhibited by S-adenosylmethionine which has shown to be actively demethylated to form S-adenosylhomocysteine, which is hydrolysed to homocysteine. MTHFR deficiency exhibits well-documented clinical and biochemical symptoms. The human MTHFR cDNA was isolated by Goyette et al (1994), and fifteen mutations have been identified at this locus. / An animal model would prove to be useful for designing therapeutic approaches for understanding the pathogenesis of this genetic disease at the molecular level. The mouse MTHFR gene and cDNA have been isolated and partially characterized. Four genomic clones were isolated by library screening. One of these clones (clone 3) contained the 5$ sp prime$ end of the gene and was completely characterized. The clone was shown to have no rearrangements and is to be used to design targeting vectors for 'knockout mice' and mice carrying a common mutation which has been postulated to be a genetic risk factor for cardiovascular disease. The other three clones contain the remaining 3$ sp prime$ portion of the gene. The coding portion has approximately 90% homology with the human cDNA and also shows a similar gene structure. / A 2.2 Kb mouse MTHFR cDNA was isolated by library screening and was found to contain a 320 base pair extension at the 5$ sp prime$ end which has not been found in the human cDNA. The cDNA contains exons -1 -3, but also contains two possibly unspliced introns. A portion of this cDNA can however still be used to rescreen libraries to isolate a full length cDNA. The above research is the first genetic data on the mouse MTHFR gene and provides the basis for future research involving mouse models of MTHFR deficiency.
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