BENZALDEHYDE LYASE, A NOVEL THIAMINE PPI-REQUIRING ENZYME, FROM PSEUDOMONAS-FLUORESCENS BIOVAR-I
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1989
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Abstract
Pseudomonas fluorescens biovar I can grow on benzoin as the sole carbon and energy source. This ability is due to benzaldehyde lyase, a new type of enzyme that irreversibly cleaves the acyloin linkage of benzoin, producing two molecules of benzaldehyde. Benzaldehyde lyase was purified 70-fold and found to require catalytic amounts of thiamine PPi (TPP) and a divalent cation as cofactors. Optimal activity was obtained with a 1.0 mM concentration of Mn2+, Mg2+, or Ca2+. Gel permeation chromatography indicated a native molecular weight of 80,000, whereas the enzyme migrated in sodium dodecyl sulfate-containing polyacrylamide gels as a single polypeptide with a molecular weight of 53,000. Benzaldehyde lyase is highly specific; of a variety of structurally related compounds tested, only benzoin and benzoin and anisoin (4,4''-dimethoxybenzoin) acted as substrates, their apparent Kms being 9.0 .times. 10-3 and 3.25 .times. 10-2 mM, respectively. A catalytic mechanism for the enzyme is proposed.