Many bioactive polypeptides, such as tyrocidine, gramicidin, bacitracin, and vancomycin are synthesized by complex enzymes and not by ribosomal assembly (Konz and Marahiel, 1999; Marahiel et al., 1997; Walsh, 2003b). The basic chemistry of peptide bond formation is analogous to ribosomal assembly, but short-circuited with the avoidance of nucleic acids to specify amino acid sequences. The nonribosomal polypeptide synthetases (NRPSs) consist of enzymatic domains that catalyze each cycle of peptide bond formation. As in type I PKSs, NRPSs are organized in modules, each of which contains a phosphopanteth-einyl domain, known as PCP, and an amino acid activating domain, known as AAla, Aval, and so forth. Each module activates a cognate amino acid as a PCP-thioester by the mechanism shown in fig. 18-14. Reaction of an amino acid with MgATP catalyzed by a cognate activation domain leads to the aminoacyl-adenylate, which donates the aminoacyl group to the thiol of PCP to form the aminoacyl-PCP with elimination of AMP. The activated aminoacyl-PCPs retain free amino groups that can react with neighboring aminoacyl-PCP thioesters to form peptide bonds.
The sequence of modules defines the sequence in which the aminoacyl residues appear in the polypeptide. This arrangement would be too cumbersome for the assembly of very large polypeptides and proteins, which are more efficiently produced by ribosomal biosynthesis. However, the NRPSs lend themselves to the incorporation of many more amino acids than the standard 20 residues found in proteins, a process that is accomplished by two means. An activating domain in a module may recognize an amino acid other than one of the 20 common l-amino acids. Alternatively, a module may contain a domain that catalyzes a chemical modification of an aminoacyl group being incorporated into the polypeptide. Many polypeptides produced by NRPSs contain nonstandard amino acids such as d-amino acids, and this feature enhances the diversity of the polypeptides that can be produced by these complexes.
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