The long term goal of this research is to characterize the metabolic regulation of pathways of folate metabolism in both normal and diseased states, and to determine reaction mechanisms for folate-dependent enzymes. We propose to continue our studies on methylenetetrahydrofolate reductase, a flavoprotein which regulates a key branch point in folate metabolism. This enzyme catalyzes the first step in the flow of one carbon unit from methylenetetrahydrofolate into the pathway for de novo biosynthesis of methyl groups. We will develop affinity chromatography procedures which are designed to facilitate purification of the enzyme to homogeneity in good yield. We will then determine the physical and catalytic properties of the enzyme, and examine further the exchange of methylene protons associated with reduction of methylenetetrahydrofolate by measuring the isotopic discrimination associated with the exchange reaction. Purification procedures will be developed for the next enzyme in the pathway of methyl group biosynthesis, vitamin B12-dependent methyltetrahydrofolate-homocysteine methyl transferase. We will examine the way in which methyltetrahydrofolate is activated for methyl transfer to the B12 prosthetic group. Particular emphasis will be placed on characterizing interactions between methylenetetrahydrofolate reductase and methyltetrahydrofolate-homocysteine methyl transferase which affect methyl group transfer.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM024908-08
Application #
3272644
Study Section
Biochemistry Study Section (BIO)
Project Start
1978-04-01
Project End
1986-03-31
Budget Start
1985-04-01
Budget End
1986-03-31
Support Year
8
Fiscal Year
1985
Total Cost
Indirect Cost
Name
University of Michigan Ann Arbor
Department
Type
Organized Research Units
DUNS #
791277940
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
Koutmos, Markos; Gherasim, Carmen; Smith, Janet L et al. (2011) Structural basis of multifunctionality in a vitamin B12-processing enzyme. J Biol Chem 286:29780-7
Matthews, Rowena G (2009) Cobalamin- and corrinoid-dependent enzymes. Met Ions Life Sci 6:53-114
Liptak, Matthew D; Fleischhacker, Angela S; Matthews, Rowena G et al. (2009) Spectroscopic and computational characterization of the base-off forms of cob(II)alamin. J Phys Chem B 113:5245-54
Matthews, Rowena G; Koutmos, Markos; Datta, Supratim (2008) Cobalamin-dependent and cobamide-dependent methyltransferases. Curr Opin Struct Biol 18:658-66
Fleischhacker, Angela S; Matthews, Rowena G (2007) Ligand trans influence governs conformation in cobalamin-dependent methionine synthase. Biochemistry 46:12382-92
Huang, Sha; Romanchuk, Gail; Pattridge, Katherine et al. (2007) Reactivation of methionine synthase from Thermotoga maritima (TM0268) requires the downstream gene product TM0269. Protein Sci 16:1588-95
Pejchal, Robert; Campbell, Elizabeth; Guenther, Brian D et al. (2006) Structural perturbations in the Ala --> Val polymorphism of methylenetetrahydrofolate reductase: how binding of folates may protect against inactivation. Biochemistry 45:4808-18
Taurog, Rebecca E; Matthews, Rowena G (2006) Activation of methyltetrahydrofolate by cobalamin-independent methionine synthase. Biochemistry 45:5092-102
Yamada, Kazuhiro; Gravel, Roy A; Toraya, Tetsuo et al. (2006) Human methionine synthase reductase is a molecular chaperone for human methionine synthase. Proc Natl Acad Sci U S A 103:9476-81
Taurog, Rebecca E; Jakubowski, Hieronim; Matthews, Rowena G (2006) Synergistic, random sequential binding of substrates in cobalamin-independent methionine synthase. Biochemistry 45:5083-91

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