L-methionine decarboxylase from Dryopteris filix-mas: Purification, characterization, substrate specificity, abortive transamination of the coenzyme, and stereochemical courses of substrate decarboxylation and coenzyme transamination

Journal Article · Tue Aug 21 00:00:00 EDT 1990 · Biochemistry; (USA)

DOI:https://doi.org/10.1021/bi00485a013· OSTI ID:6098241

Stevenson, D E; Akhtar, M; Gani, D ^[1]

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1. Univ. of Southampton (England)

L-Methionine decarboxylase from the male fern Dryopteris filix-mas has been purified 256-fold from acetone powder extracts to very near homogeneity. The enzyme is membrane-associated and requires detergent for solubilization during the initial extraction. The enzyme is a homodimer of subunit M{sub r} 57,000 and shows a pH optimum at {approximately} 5.0 with 20 mM (2S)-methionine as substrate. A wide range of straight- and branched-chain (2S)-alkylamino acids are substrates for the enzyme. The values for the rate of decarboxylation, V{sub max}, and for the apparent Michaelis constant, K{sub m}, however, vary with structure and with the chirality at C-3. The pH dependence of V and V/K has been examined for three substrates: (2S)-methionine, valine, and leucine. The occurrence of the abortive reaction was confirmed by showing that ({sup 35}S)methionine is converted to labeled 3-(methylthio)propionaldehyde while (4{prime}-{sup 3}H)PLP is converted to labeled pyridoxamine 5{prime}-phosphate (PMP). The decarboxylation of (2S)-methionine gave 3(methylthio)-1-aminopropane. Preparation of the N-camphanamide derivative of the amine allowed the C-1 methylene protons to be distinguished by {sup 1}H NMR spectroscopy. Synthetic samples of the camphanamide were prepared in which each of the C-1 methylene protons was replaced by deuterium. When tritiated pyridoxal phosphate was incubated with the enzyme, tritiated pyridoxamine phosphate was formed. These results are used to construct possible mechanistic schemes for both reactions, decarboxylation and transamination. The position and possible identities of active-site proton donors are discussed.

• OSTI ID: 6098241
• Journal Information: Biochemistry; (USA), Vol. 29:33; ISSN 0006-2960
• Country of Publication: United States
• Language: English

Related Subjects

62 RADIOLOGY AND NUCLEAR MEDICINE
COENZYMES
BIOCHEMICAL REACTION KINETICS
DECARBOXYLASES
NUCLEAR MAGNETIC RESONANCE
AMINATION
DECARBOXYLATION
METHIONINE
PROTONS
PURIFICATION
STEREOCHEMISTRY
SUBSTRATES
TRITIUM COMPOUNDS
AMINO ACIDS
BARYONS
CARBON-CARBON LYASES
CARBOXY-LYASES
CARBOXYLIC ACIDS
CHEMICAL REACTIONS
DRUGS
ELEMENTARY PARTICLES
ENZYMES
FERMIONS
HADRONS
HYDROGEN COMPOUNDS
KINETICS
LIPOTROPIC FACTORS
LYASES
MAGNETIC RESONANCE
NUCLEONS
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC SULFUR COMPOUNDS
REACTION KINETICS
RESONANCE
550601* - Medicine- Unsealed Radionuclides in Diagnostics