A role for glutamine 183 in the folate oxidative half-reaction of methylenetetrahydrofolate reductase from Escherichia coli.

The flavoprotein methylenetetrahydrofolate reductase (MTHFR) from Escherichia coli catalyzes a ping-pong reaction with NADH and 5,10-methylenetetrahydrofolate (CH-Hfolate) to produce NAD and 5-methyltetrahydrofolate (CH-Hfolate). This work focuses on the function of the invariant, active-site aminoacyl residue Gln183. X-ray structures of the enzyme complexes E(wild-type)•NADH and E(Glu28Gln)•CH-Hfolate indicate that Gln183 makes key hydrogen-bonding interactions with both NADH and folate in their respective half-reactions, suggesting roles in binding each substrate.

Mechanistic Studies of an Amine Oxidase Derived from d-Amino Acid Oxidase.

The flavoprotein d-amino acid oxidase has long served as a paradigm for understanding the mechanism of oxidation of amino acids by flavoproteins. Recently, a mutant d-amino acid oxidase (Y228L/R283G) that catalyzed the oxidation of amines rather than amino acids was described [Yasukawa, K., et al.

Methylenetetrahydrofolate reductase: biochemical characterization and medical significance.

Methylenetetrahydrofolate reductase (MTHFR) catalyzes the reduction of 5,10-methylenetetrahydofolate (CH2-H4folate) to 5-methyltetrahydrofolate (CH3-H4folate). The enzyme employs a noncovalently-bound flavin adenine dinucleotide (FAD), which accepts reducing equivalents from NAD(P)H and transfers them to CH2-H4folate. The reaction provides the sole source of CH3-H4folate, which is utilized by methionine synthase in the synthesis of methionine from homocysteine.

Functional role for the conformationally mobile phenylalanine 223 in the reaction of methylenetetrahydrofolate reductase from Escherichia coli.

The flavoprotein methylenetetrahydrofolate reductase from Escherichia coli catalyzes the reduction of 5,10-methylenetetrahydrofolate (CH(2)-H(4)folate) by NADH via a ping-pong reaction mechanism. Structures of the reduced enzyme in complex with NADH and of the oxidized Glu28Gln enzyme in complex with CH(3)-H(4)folate [Pejchal, R., Sargeant, R.

Novel syn intramolecular pathway in base-catalyzed 1,2-elimination reactions of beta-acetoxy esters.

As part of a comprehensive investigation of electronic effects on the stereochemistry of base-catalyzed 1,2-elimination reactions, we observed a new syn intramolecular pathway in the elimination of acetic acid from beta-acetoxy esters and thioesters. 1H and 2H NMR investigation of reactions using stereospecifically labeled tert-butyl (2R*,3R*)-3-acetoxy-2,3-2H2-butanoate (1) and its (2R*,3S*) diastereomer (2) shows that 23 +/- 2% syn elimination occurs. The elimination reactions were catalyzed with KOH or (CH3)4NOH in ethanol/water under rigorously non-ion-pairing conditions.

Aspartate 120 of Escherichia coli methylenetetrahydrofolate reductase: evidence for major roles in folate binding and catalysis and a minor role in flavin reactivity.

Escherichia coli methylenetetrahydrofolate reductase (MTHFR) catalyzes the NADH-linked reduction of 5,10-methylenetetrahydrofolate (CH(2)-H(4)folate) to 5-methyltetrahydrofolate (CH(3)-H(4)folate) using flavin adenine dinucleotide (FAD) as cofactor. MTHFR is unusual among flavin oxidoreductases because it contains a conserved, negatively rather than positively charged amino acid (aspartate 120) near the N1-C2=O position of the flavin. At this location, Asp 120 is expected to influence the redox properties of the enzyme-bound FAD.