Analysis of a mutant amino acid-activating domain of surfactin synthetase bearing a serine-to-alanine substitution at the site of carboxylthioester formation.

The reactive serine of the TGGHSL thioester binding motif of the first amino acid-activating domain of surfactin synthetase was replaced by alanine using site-directed mutagenesis. The multienzyme from cells of the resulting mutant lost its ability for thioester formation with L-Glu and was therefore inactive in surfactin production. The thiolation reactions catalyzed by the other amino acid-activating domains of surfactin synthetase were not affected by the mutation. The results show that L-Glu is activated at the first domain of surfactin synthetase, and give further evidence that a serine residue is essential for substrate amino acid activation at the reaction centers of peptide synthetases.