Abstract: Deacetoxycephalosporin C synthase obtained from cell-free extracts of Streptomyces clavuligerus was purified to near homogeneity and characterized in inactive form. The amino acid sequence was determined and the synthase recombinantly reproduced in active form in E. coli. The recombinantly produced synthase is provided in about 97% pure form by a chromatographic process. The synthase lacks DAOC hydroxylase function; however, it possesses the ability to transform 3-exomethylenocephalosporin C to deacetylcephalosporin C.