Chapter 19. In vitro studies of phenol coupling enzymes involved in vancomycin biosynthesis

Methods Enzymol. 2009:458:487-509. doi: 10.1016/S0076-6879(09)04819-8.

Abstract

Oxidative phenol cross-linking reactions play a key role in the biosynthesis of glycopeptide antibiotics such as vancomycin. The vancomycin aglycone contains three cross-links between aromatic amino acid side-chains, which stabilize the folded backbone conformation required for binding to the target D-Ala-D-Ala dipeptide. At least the first cross-link is introduced into a peptide precursor whilst it is still bound as a thioester to a peptide carrier protein (PCP) domain (also called a thiolation domain) within the nonribosomal peptide synthetase. We described here methods for the solid-phase synthesis of peptides and their coupling to PCP domains, which may be useful for in vitro studies of cross-linking and related tailoring reactions during nonribosomal glycopeptide antibiotic biosynthesis.

MeSH terms

  • Computational Biology
  • Ferredoxin-NADP Reductase / metabolism
  • Ferredoxins / chemistry
  • Ferredoxins / metabolism
  • Molecular Structure
  • Peptide Biosynthesis
  • Peptide Synthases / metabolism
  • Vancomycin / biosynthesis*
  • Vancomycin / chemistry

Substances

  • Ferredoxins
  • Vancomycin
  • Ferredoxin-NADP Reductase
  • Peptide Synthases
  • non-ribosomal peptide synthase