title: Engineering of peptide synthetases. Key role of the thioesterase-like domain for efficient production of recombinant peptides.
abstract: Peptide synthetases are large enzymatic complexes that catalyze the synthesis of biologically active peptides in microorganisms and fungi and typically have an unusual structure and sequence. Peptide synthetases have recently been engineered to modify the substrate specificity to produce peptides of a new sequence. In this study we show that surfactin synthetase can also be modified by moving the carboxyl-terminal intrinsic thioesterase region to the end of the internal amino acid binding domains, thus generating strains that produce new truncated peptides of the predicted sequence. Omission of the thioesterase domain results in nonproducing strains, thus showing the essential role of this region and the possibility of obtaining peptides of different lengths by genetic engineering. Secretion of the peptides depends on the presence of a functional sfp gene.
authors.affiliation: Genetic Engineering and Microbiology Laboratories, Environmental Technologies, Eniricerche S.p.A., 20097 San Donato Milanese, Italy. fdeferra@enricerche.eni.it
authors.forename: F
authors.initials: F
authors.lastname: de Ferra
authors.forename: F
authors.initials: F
authors.lastname: Rodriguez
authors.forename: O
authors.initials: O
authors.lastname: Tortora
authors.forename: C
authors.initials: C
authors.lastname: Tosi
authors.forename: G
authors.initials: G
authors.lastname: Grandi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001419:Bacteria / Q000201:enzymology; D001426:Bacterial Proteins; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular / Q000379:methods; D017124:Conserved Sequence; D017931:DNA Primers; D008969:Molecular Sequence Data; D010453:Peptide Synthases / Q000096:biosynthesis* / Q000737:chemistry; D010957:Plasmids; D015202:Protein Engineering / Q000379:methods; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013869:Thiolester Hydrolases / Q000737:chemistry
chemical_list: D001426:Bacterial Proteins; D017931:DNA Primers; D011994:Recombinant Proteins; D013869:Thiolester Hydrolases; D010453:Peptide Synthases; C069542:surfactin synthetase