Studies on the biosynthesis of isopenicillin N with a cell-free preparation of Penicillium chrysogenum.

When delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine was added to a cell-free system prepared by lysis of Penicillium chrysogenum protoplasts, "compounds X and Y" were detected after analysis on a cation-exchange column. The chromatographic position as well as results of experiments with double labelled tripeptides showed "compound X" to be the penicilloic acid of isopenicillin N. LLD-Tripeptide labelled with tritium at carbon-2 of th valine part was incorporated into isopenicillin N with retention of label. "Compound Y" retained all hydrogens on the valine part of the peptide, but lost half of the tritium on carbon-3 of the cysteine part. The results are consistent with the hypothesis that the LLD-tripeptide is converted into isopenicillin N via a monocyclic beta-lactam and without a dehydrovalinyl intermediate. Extensive transacylase activity was observed between isopenicillin N and 6-aminopenicillanic acid.