Acetyl CoA: deacetylcephalosporin C o-acetyltransferase(DCPC-ATF) catalyses the final step in the biosynthesis of cephalosporin C, the conversion of deacetylcephalosporin C to cephalosporin C. A cDNA encoding DCPC-ATF has been isolated from a cDNA library of a cephalosporin C producing fungus Acremonium chrysogenum using oligonucleotide probes based on N-terminal amino acid sequences of the enzyme. The cDNA contains a single large open reading frame for a putative precursor consisting of 12 amino acid(AA) leader peptide of unknown function, 274 AA large subunit and 126 AA small subunit at the carboxyl end. The cDNA was expressed in yeast exhibiting a functional DCPC-ATF activity. It was also indicated that the leader peptide was not essential for expression of the enzyme activity. The primary structure of DCPC-ATF shows significant homology with those of acetyl CoA: homoserine o-acetyltransferase in Saccharomyces cerevisiae and Ascobolas immersus.