Gene cloning and expression of cephalosporin-C deacetylase from Bacillus sp KCCM10143

Cephalosporin-C deacetylase (CAH) catalyzes the deacetylation of cephalosporin derivatives. A novel gene encoding the CAH from Bacillus sp. KCCM10143 was cloned and sequenced. The nucleotide sequence contained an open reading frame encoding a polypeptide consisting of 217 amino acids and a molecular weight of 24 kDa which was in good agreement with the value obtained by sodium dodecylsulfate-polyacrylamide gel electrophoresis. An expression plasmid was constructed by inserting the CAH gene into the region of the pTrc99A expression vector. An active form of the CAH protein was expressed in the soluble fraction obtained after cell disruption. In fermentation using a 5-1 jar fermenter, the transformant E. coli JM109 (pDST654) produced 4.12 U of CAH per ml of culture during 16 h of incubation.