Inactivation of deacetoxycephalosporin C synthase in extracts of Streptomyces clavuligerus during bioconversion of penicillin G to deacetoxycephalosporin G

By using cell-free extracts of Streptomyces clavuligerus containing deaceto xycephalosporin C synthase, we Can convert penicillin G into deacetoxycepha losporin G. The degree of bioconversion was low (below 1% based on charged substrate) and decreased as substrate concentration was increased. Formatio n of product occurred during the first 2 h and then ceased. Addition of inc reasing levels of cofactors and/or substrate at the 2-h point did not resul t in more product. Preincubation of the crude extracts in the presence of f errous ions plus either ascorbate or alpha-ketoglutarate led to complete lo ss of activity. Such inactivation could not be reversed by catalase, supero xide dismutase, mannitol, thiourea, dimethylsulfoxide, dithiothreitol, or b eta-mercaptoethanol. (C) 1999 Elsevier Science Inc. All rights reserved.