Synthesis of a S‐(Carboxymethyl)cysteine Analog of (L‐2‐Amino‐6‐adipyl)‐L‐cysteinyl‐D‐valine and its Cell Free Biosynthetic Conversion into 6‐[2‐(D‐2‐Amino‐2‐carboxyethyl)thio) acetamido]penicillanic Acid

A new tripeptide (dimer), bis[(L‐cysteine‐S‐acetyl)‐L‐hemicystinyl(S2 → S2)‐D‐valine] (6) was synthesized by coupling N‐(tert‐butoxycarbonyl)‐S‐carboxymethyl‐L‐cysteine benzyl ester (1) with S‐trityl‐L‐cysteinyl‐D‐valine benzyl ester and subsequent removal of the protecting groups. After reduction of the disulfide, the free tripeptide Cys (CH2CO‐Cys‐D‐Val) (Ib) was used as a substrate of isopenicillin‐N synthetase in a cell‐free conversion to 6‐[2‐((D‐2‐amino‐2‐carboxyethyl)thio)acetamido]penicillanic acid (IIa).