Chemoenzymatic synthesis of peptide beta-lactones and beta-hydroxy acids

A method of producing a peptide beta-lactone, the method comprising:

• forming a reaction mixture comprising:
• a beta-hydroxy-alpha-amino acid selected from the group consisting of beta-OH-p-NO2-homoPhe and beta-OH-homoPhe;
• a benzoic acid derivative consisting of 2,3-dihydroxoybenzoic acid;
• ATP;
• an aryl carrier protein comprising the amino acid sequence of SEQ ID NO:1;
• and
• a non-ribosomal protein synthetase comprising the amino acid sequence of any one of SEQ ID NOS:2, 25, and 26, the non-ribosomal protein synthetase further comprising a condensation domain (C), a first adenylation domain (A1), a peptidyl carrier domain (PCP), a thioesterase domain (TE), and a second adenylation domain (A2);
• contacting the beta-hydroxy-alpha-amino acid with the non-ribosomal protein synthetase at the peptidyl carrier domain (PCP) with ATP activation by the first adenylation domain (A1) to form a PCP-beta-hydroxy-alpha-amino acid thioester comprising an alpha-amino moiety;
• ATP-activating the benzoic acid derivative at the second adenylation domain (A2) and contacting the ATP-activated benzoic acid derivative with the aryl carrier protein to form a benzoic acid derivative-aryl carrier protein thioester comprising a carbonyl moiety;
• contacting the benzoic acid derivative-aryl carrier protein thioester with the condensation domain (C) to catalyze an amide bond between the alpha-amino moiety and the carbonyl moiety to form a PCP-peptide beta-lactone precursor thioester;
• contacting the PCP-peptide beta-lactone precursor thioester with the thioesterase domain (TE) to form a transthioesterified peptide beta-lactone precursor;
• and
• releasing the transthioesterified peptide beta-lactone precursor as a peptide beta-lactone from the non-ribosomal protein synthetase.