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Assays of methylenetetrahydrofolate reductase and methionine synthase activities by monitoring 5-methyltetrahydrofolate and tetrahydrofolate using high-performance liquid chromatography with fluorescence detection.

Authors: Huang, L  Zhang, J  Hayakawa, T  Tsuge, H 
Citation: Huang L, etal., Anal Biochem. 2001 Dec 15;299(2):253-9.
Pubmed: (View Article at PubMed) PMID:11730351
DOI: Full-text: DOI:10.1006/abio.2001.5421

We developed a method for assays of methylenetetrahydrofolate reductase and methionine synthase activities by monitoring their products of 5-methyltetrahydrofolate (5-CH(3)-H(4)folate) and tetrahydrofolate (H(4)folate) directly, using high-performance liquid chromatography with fluorescence detection. Folate derivatives and enzymes were stable in the assay process. No reagents in the assay mixture were found to disturb the separation and detection of both H(4)folate and 5-CH(3)-H(4)folate in our assay system. The detection limit of this method was less than 20 nM H(4)folate or 5-CH(3)-H(4)folate in the enzyme assay system. This analytical method, therefore, has a sensitivity high enough to obtain accurate parameters of Michaelis-Menten kinetics and for assays of crude extracts from various biological samples. In addition, the analytical procedure is very simple and economical; it may be a useful tool for studying methylenetetrahydrofolate reductase and methionine synthase activities.

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CRRD Object Information
CRRD ID: 1601423
Created: 2007-04-19
Species: All species
Last Modified: 2007-04-19
Status: ACTIVE



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RGD is funded by grant HL64541 from the National Heart, Lung, and Blood Institute on behalf of the NIH.