Open AccessAssay of 4‐hydroxybutyryl‐CoA dehydratase from Clostridium aminobutyricum
Author(s) -
Willadsen Peter,
Buckel Wolfgang
Publication year - 1990
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.1990.tb13976.x
Subject(s) - dehydratase , stereospecificity , dehydration , enzyme , chemistry , coenzyme a , biochemistry , clostridium , cofactor , acetyl coa , stereochemistry , catalysis , biology , bacteria , reductase , genetics
1. Summary It has been proposed that Clostridium aminobutyricum contains an enzyme catalyzing an unusual reaction: the dehydration of 4‐hydroxybutyryl‐CoA to vinylacetyl‐CoA. 4‐Hydroxy‐[3‐ 3 H]butyric acid has been prepared which allows the activity of this enzyme to be assayed in the presence of acetyl‐CoA under anaerobic conditions by the release of tritiated water. Initial characterization of the enzyme from C. aminobutyricum has shown it to be largely membrane or particle bound in the crude lysates. It can be solubilized in detergent. It is inactivated by oxygen, but stable under anaerobic conditions. Only 49 ± 2% of the label is removed after enzyme‐catalyzed equilibration with water. This stereospecific release is consistent with the formation of vinylacetyl‐CoA and excludes a vitamin B 12 coenzyme‐dependent rearrangement to 3‐hydroxybutyryl‐CoA followed by dehydration to crotonyl‐CoA.