title: Structure of the EntB multidomain nonribosomal peptide synthetase and functional analysis of its interaction with the EntE adenylation domain.
abstract: Nonribosomal peptide synthetases are modular proteins that operate in an assembly line fashion to bind, modify, and link amino acids. In the E. coli enterobactin NRPS system, the EntE adenylation domain catalyzes the transfer of a molecule of 2,3-dihydroxybenzoic acid to the pantetheine cofactor of EntB. We present here the crystal structure of the EntB protein that contains an N-terminal isochorismate lyase domain that functions in the synthesis of 2,3-dihydroxybenzoate and a C-terminal carrier protein domain. Functional analysis showed that the EntB-EntE interaction was surprisingly tolerant of a number of point mutations on the surface of EntB and EntE. Mutational studies on EntE support our previous hypothesis that members of the adenylate-forming family of enzymes adopt two distinct conformations to catalyze the two-step reactions.
authors.affiliation: Hauptman-Woodward Medical Research Institute, 700 Ellicott Street, Buffalo, New York 14203, USA.
authors.forename: Eric J
authors.initials: EJ
authors.lastname: Drake
authors.forename: David A
authors.initials: DA
authors.lastname: Nicolai
authors.forename: Andrew M
authors.initials: AM
authors.lastname: Gulick
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D018360:Crystallography, X-Ray; D004269:DNA, Bacterial / Q000235:genetics; D004758:Enterobactin / Q000096:biosynthesis / Q000737:chemistry; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008025:Ligases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010453:Peptide Synthases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D055672:Static Electricity
chemical_list: D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D004758:Enterobactin; D008025:Ligases; D010453:Peptide Synthases; C104385:non-ribosomal peptide synthase; C061746:2,3-dihydroxybenzoate-AMP ligase, E coli