title: Role of MbtH-like proteins in the adenylation of tyrosine during aminocoumarin and vancomycin biosynthesis.
abstract: MbtH-like proteins consist of ∼70 amino acids and are encoded in the biosynthetic gene clusters of non-ribosomally formed peptides and other secondary metabolites derived from amino acids. Recently, several MbtH-like proteins have been shown to be required for the adenylation of amino acid in non-ribosomal peptide synthesis. We now investigated the role of MbtH-like proteins in the biosynthesis of the aminocoumarin antibiotics novobiocin, clorobiocin, and simocyclinone D8 and of the glycopeptide antibiotic vancomycin. The tyrosine-adenylating enzymes CloH, SimH, and Pcza361.18, involved in the biosynthesis of clorobiocin, simocyclinone D8, and vancomycin, respectively, required the presence of MbtH-like proteins in a 1:1 molar ratio, forming heterotetrameric complexes. In contrast, NovH, involved in novobiocin biosynthesis, showed activity in the absence of MbtH-like proteins. Comparison of the active centers of CloH and NovH showed only one amino acid to be different, i.e. Leu-383 versus Met-383. Mutation of this amino acid in CloH (L383M) indeed led to MbtH-independent adenylating activity. All investigated tyrosine-adenylating enzymes exhibited remarkable promiscuity for MbtH-like proteins from different pathways and organisms. YbdZ, the MbtH-like protein from the expression host Escherichia coli, was found to bind to adenylating enzymes during expression and to influence their biochemical properties markedly. Therefore, the use of ybdZ-deficient expression hosts is important in biochemical studies of adenylating enzymes.
authors: Pharmazeutisches Institut, Universität Tübingen, 72076 Tübingen, Germany.
authors: Björn
authors: B
authors: Boll
authors: Tatjana
authors: T
authors: Taubitz
authors: Lutz
authors: L
authors: Heide
mesh_terms: D019943:Amino Acid Substitution; D049933:Aminocoumarins / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D009097:Multienzyme Complexes / Q000235:genetics / Q000378:metabolism; D020125:Mutation, Missense; D009713:Nucleotidyltransferases / Q000235:genetics / Q000378:metabolism*; D048372:Streptomyces coelicolor / Q000201:enzymology* / Q000235:genetics; D014443:Tyrosine / Q000235:genetics / Q000378:metabolism*; D014640:Vancomycin / Q000096:biosynthesis*
chemical_list: D049933:Aminocoumarins; D001426:Bacterial Proteins; D009097:Multienzyme Complexes; D014443:Tyrosine; D014640:Vancomycin; D009713:Nucleotidyltransferases