Folate functions in multiple coenzyme forms in acceptance, redox processing and transfer of one-carbon units, including nucleotides and certain amino acids. Folate-requiring metabolic processes are influenced by folate intake, intake of other essential nutrients, including vitamins B-12 and B-6, and at least one common genetic polymorphism. Estimates of folate requirements have been based on intakes associated with maintenance of normal plasma and erythrocyte folate concentrations and functional tests that reflect abnormalities in folate-dependent reactions. Dietary Reference Intakes for folate that have been developed recently are based primarily on metabolic studies in which erythrocyte folate concentration was considered the major indicator of adequacy. For adults >/=19 y, the Recommended Dietary Allowance (RDA) is 400 microg/d of dietary folate equivalents (DFE); for lactating and pregnant women, the RDAs include an additional 100 and 200 microg of DFE/d, respectively.

Measures of B6 status are categorized as direct biomarkers and as functional biomarkers. Direct biomarkers measure B6 vitamers in plasma/serum, urine and erythrocytes, and among these plasma pyridoxal 5-phosphate (PLP) is most commonly used. Functional biomarkers include erythrocyte transaminase activities and more recently plasma levels of metabolites involved in PLP-dependent reactions, such as the kynurenine pathway, one-carbon metabolism, transsulfuration (cystathionine), and glycine decarboxylation (serine and glycine). Vitamin B6 status is best assessed by using a combination of biomarkers because of the influence of potential confounders, such as inflammation, alkaline phosphatase activity, low serum albumin, renal function and inorganic phosphate. Ratios between substrate-products pairs have recently been investigated as a strategy to attenuate such influence. These efforts have provided promising new markers such as the PAr index, the 3-hydroxykynurenine/xanthurenic acid ratio and the oxoglutarate:glutamate ratio. Targeted metabolic profiling or untargeted metabolomics based on mass spectrometry allow the simultaneous quantification of a large number of metabolites, which are currently evaluated as functional biomarkers, using data reduction statistics.

A common genetic polymorphism results from a C-->T substitution in the gene encoding methylenetetrahydrofolate reductase (MTHFR), the enzyme that produces 5-methyltetrahydrofolate (5-methyl-THF) required for the conversion of homocysteine to methionine. In individuals with the T/T genotype (T/T), functional metabolic effects include changes in one-carbon folate derivatives, elevations in plasma homocysteine and differences in response to folic acid supplementation compared with normal (C/C) or heterozygous (C/T) genotypes. The metabolic changes associated with the T/T genotype are postulated to modify risk for chronic disease (e.g., vascular disease and cancer) and neural tube defects (NTD) when accompanied by folate deficiency. The modulation of these metabolic abnormalities by increasing folate intake suggests that folate requirements may be different in affected individuals (T/T) relative to normal (C/C) or heterozygous (C/T) individuals. The complex interaction between this common genetic polymorphism of MTHFR and folate intake is the focus of intense investigation.

Typical intakes of folic acid from fortified foods are more than twice the level originally predicted. The effect of this much higher level of fortification must be carefully assessed, especially before calls for higher levels of fortification are considered.

Pyridoxal 59-phosphate (PLP) is an essential cofactor for nearly 60 Escherichia coli enzymes but is a highly reactive molecule that is toxic in its free form. How PLP levels are regulated and how PLP is delivered to target enzymes are still open questions. The COG0325 protein family belongs to the fold-type III class of PLP enzymes and binds PLP but has no known biochemical activity although it occurs in all kingdoms of life. Various pleiotropic phenotypes of the E. coli COG0325 (yggS) mutant have been reported, some of which were reproduced and extended in this study. Comparative genomic, genetic and metabolic analyses suggest that these phenotypes reflect an imbalance in PLP homeostasis. The E. coli yggS mutant accumulates the PLP precursor pyridoxine 59-phosphate (PNP) and is sensitive to an excess of pyridoxine but not of pyridoxal. The pyridoxine toxicity phenotype is complemented by the expression of eukaryotic yggS orthologs. It is also suppressed by the presence of amino acids, specifically isoleucine, threonine and leucine, suggesting the PLP-dependent enzyme transaminase B (IlvE) is affected. These genetic results lay a foundation for future biochemical studies of the role of COG0325 proteins in PLP homeostasis.

Impaired folate-mediated 1-carbon metabolism has been linked to multiple disease outcomes. A better understanding of the nutritional and genetic influences on this complex biochemical pathway is needed to comprehend their impact on human health. To this end, we created a mathematical model of folate-mediated 1-carbon metabolism. The model uses published data on folate enzyme kinetics and regulatory mechanisms to simulate the impact of genetic and nutritional variation on critical aspects of the pathway. We found that the model predictions match experimental data, while providing novel insights into pathway kinetics. Our primary observations were as follows: 1) the inverse association between folate and homocysteine is strongest at very low folate concentrations, but there is no association at high folate concentrations; 2) the DNA methylation reaction rate is relatively insensitive to changes in folate pool size; and 3) as folate concentrations become very high, enzyme velocities decrease. With regard to polymorphisms in 5,10-methylenetetrahydrofolate reductase (MTHFR), the modeling predicts that decrease MTHFR activity reduces concentrations of S-adenosylmethionine and 5-methyltetrahydrofolate, as well as DNA methylation, while modestly increasing S-adenosylhomocysteine and homocysteine concentrations and thymidine or purine synthesis. Decreased folate together with a simulated vitamin B-12 deficiency results in decreases in DNA methylation and purine and thymidine synthesis. Decreased MTHFR activity superimposed on the B-12 deficiency appears to reverse the declines in purine and thymidine synthesis. These mathematical simulations of folate-mediated 1-carbon metabolism provide a cost-efficient approach to in silico experimentation that can complement and help guide laboratory studies.

, respectively). However, the rate of overall homocysteine remethylation (ϳ8 mol⅐kg Ϫ1 ⅐h Ϫ1) was twice that of previous reports, which suggests a larger role for homocysteine remethylation in methionine metabolism than previously thought. By use of estimates of intracellular [3-13 C]serine enrichment based on a conservative correction of plasma [3-13 C]serine enrichment, serine was calculated to contribute ϳ100% of the methyl groups used for total body homocysteine remethylation under the conditions of this protocol. This contribution represented only a small fraction (ϳ2.8%) of total serine flux. Our dual-tracer procedure is well suited to measure the effects of nutrient deficiencies, genetic polymorphisms, and other metabolic perturbations on homocysteine synthesis and total and folate-dependent homocysteine remethylation. methionine; methylation cycle; cystathionine ELEVATED PLASMA HOMOCYSTEINE CONCENTRATION is considered an independent risk factor for the development of cardiovascular disease (5,30,32). Accordingly, many investigators have sought to define the genetic and environmental factors that affect plasma homocysteine concentration. Associations exist between plasma homocysteine concentrations and gene polymorphisms, as well as lifestyle and other environmental factors (5). Strong evidence implicates nutritional deficiencies of folate, vitamins B 6 and B 12 , and the methylenetetrahydrofolate reductase (MTHFR) 677C 3 T polymorphism as causes of elevated plasma homocysteine concentration (12, 25). Folate and vitamin B 6 deficiencies and the MTHFR 677C 3 T polymorphism are thought to increase circulating homocysteine concentrations by decreasing the availability of 5-methyltetrahydrofolate (5-CH 3 THF) and thereby inhibiting homocysteine remethylation (24). However, a causal relationship between reduced homocysteine remethylation and hyperhomocysteinemia under these conditions has not been confirmed in humans in vivo.Steady-state plasma homocysteine concentration is not solely a function of the rate of its removal by remethylation but is also affected by the rates of homocysteine production, catabolism through transsulfuration, and loss in renal excretion (24). Specific measurements of homocysteine metabolism through these individual pathways are needed to clarify why homocysteine concentration is elevated by particular genetic variations as well as by nutritional and other environmental conditions. To test the hypothesis that homocysteine remethylation is compromised when 5-CH 3 THF availability is reduced, homocysteine remethylation rates must be measured in individuals affected by these homocysteineelevating factors.Remethylation rates have been measured in humans in vivo by use of methionine tracers labeled with stable isotopes at both the carboxyl and methyl groups or else by measurements of separate methyl-and carboxyl-labeled methionine tracers in primed constant infusion experiments (18,26). By use of these methodologies, the effects of dietary sulfur amino acid intake, sex, age, prandial statu...

See https://pubs.acs.org/sharingguidelines for options on how to legitimately share published articles.