Purification and characterization of D(-)-β-hydroxybutyrate dehydrogenase from Azospirillum brasilense Cd

S. Tal, P. Smirnoff, Y. Okon

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Abstract

D(-)-β-hydroxybutyrate dehydrogenase (BOHB-DH) (EC 1.1.1.30) was purified 991-fold from Azospirillum brasilense Cd. Its specific activity was 5650 units (mg protein)-1 min-1. The enzyme is a tetramer, with identical subunits and a total molecular mass of 100 kDa. BOHB-DH is not a glycoprotein. It is acidic and contains six disulphide bonds without free -SH groups. Under the assay conditions used, BOHB-DH activity was maximal at pH 8.0 and at 36°C. The enzyme is an NAD+ oxidoreductase, and is inhibited by NADPH and NADH. It has high affinity for β-hydroxybutyrate: the K(m) value for the β-hydroxybutyrate substrate is 1 mM. Adenosine phosphates, pyruvate, acetyl-coenzyme A, oxaloacetate and 2-oxoglutarate inhibited purified BOHB-DH.

Original languageEnglish
Pages (from-to)645-649
Number of pages5
JournalJournal of General Microbiology
Volume136
Issue number4
DOIs
StatePublished - 1990

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