Purification and properties of biotinyl-CoA synthetase from Mycoplana sp. No. 166.
Biotinyl-CoA synthetase, the first enzyme involved in biotin degradation, was purified from a cell-free extract of a biotin-degrading bacterium, Mycoplana sp. No. 166. The purification procedures comprised polyethyleneimine treatment, ammonium sulfate fractionation, and DEAE-Sepharose, Blue-Sepharose, Sephadex G-100, and FPLC (Mono Q HR 5/5) column chromatographies. The enzyme was found to be homogeneous on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme was a monomeric enzyme with a molecular weight and an isoelectric point of about 55,000 and 4.85, respectively. The purified enzyme catalyzed the stoichiometric conversion of biotin, ATP, and CoA into biotinyl-CoA, AMP, and PPi. Dethiobiotin and actithiazic acid, a synthetic biotin analog, were also effective as substrates. Other properties of the purified enzyme were also investigated.
D003066:Coenzyme A Ligases / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D004795:Enzyme Stability; D006088:Gram-Negative Aerobic Bacteria / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D007700:Kinetics; D008970:Molecular Weight; D013379:Substrate Specificity; D013696:Temperature
D003066:Coenzyme A Ligases; C051127:biotin-CoA ligase