Publications

• Regulation of the enzymes involved in the biosynthesis of 2,3-dihydroxybenzoic acid in Aerobacter aerogenes and Escherichia coli.

  Young I.G., Gibson F.

  Biochim Biophys Acta 177:401-411(1969) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.

• Subcloning of the enterobactin biosynthetic gene entB: expression, purification, characterization, and substrate specificity of isochorismatase.

  Rusnak F., Liu J., Quinn N., Berchtold G.A., Walsh C.T.

  The Escherichia coli entB gene, coding for the enterobactin biosynthetic enzyme isochorismatase, has been subcloned into the multicopy plasmid pKK223-3 under the control of the tac promoter. The resulting recombinant plasmid pFR1 expresses isochorismatase amounting to over 50% of the total cellula ... >> More

  The Escherichia coli entB gene, coding for the enterobactin biosynthetic enzyme isochorismatase, has been subcloned into the multicopy plasmid pKK223-3 under the control of the tac promoter. The resulting recombinant plasmid pFR1 expresses isochorismatase amounting to over 50% of the total cellular protein. The enzyme has been purified to homogeneity and a convenient assay developed. The enzyme has a Km for isochorismate of 14.7 microM and a turnover number of 600 min-1. By use of 1H NMR spectroscopy, the progress of the reaction was followed with the expected formation of 2,3-dihydro-2,3-dihydroxybenzoate product. Several substrate analogues were also utilized by the enzyme including chorismic acid, the immediate precursor to isochorismic acid in the enterobactin biosynthetic pathway. << Less

  Biochemistry 29:1425-1435(1990) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Enterobactin biosynthesis in Escherichia coli: isochorismate lyase (EntB) is a bifunctional enzyme that is phosphopantetheinylated by EntD and then acylated by EntE using ATP and 2,3-dihydroxybenzoate.

  Gehring A.M., Bradley K.A., Walsh C.T.

  In Escherichia coli, the siderophore molecule enterobactin is synthesized in response to iron deprivation by formation of an amide bond between 2,3-dihydroxybenzoate (2,3-DHB) and l-serine and formation of ester linkages between three such N-acylated serine residues. We show that EntB, previously ... >> More

  In Escherichia coli, the siderophore molecule enterobactin is synthesized in response to iron deprivation by formation of an amide bond between 2,3-dihydroxybenzoate (2,3-DHB) and l-serine and formation of ester linkages between three such N-acylated serine residues. We show that EntB, previously described as the isochorismate lyase required for production of 2,3-DHB, is a bifunctional protein that also serves as an aryl carrier protein (ArCP) with a role in enterobactin assembly. EntB is phosphopantetheinylated near the C terminus in a reaction catalyzed by EntD with a kcat of 5 min-1 and a Km for apo-EntB of 6.5 microM. This holo-EntB is then acylated with 2,3-DHB in a reaction catalyzed by EntE, previously described as the 2,3-DHB-AMP ligase, with a kcat of 100 min-1 and a Km of <<1 microM for holo-EntB. The N-terminal 187 amino acids of EntB (isochorismate lyase domain) are not needed for reaction of EntB with either EntD or EntE as demonstrated by the equivalent catalytic efficiencies of the full-length EntB (residues 1-285) and the C-terminal EntB ArCP domain (residues 188-285) as substrates for both EntD and EntE. << Less

  Biochemistry 36:8495-8503(1997) [PubMed] [EuropePMC]

  This publication is cited by 6 other entries.

• Structure of the EntB multidomain nonribosomal peptide synthetase and functional analysis of its interaction with the EntE adenylation domain.

  Drake E.J., Nicolai D.A., Gulick A.M.

  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 o ... >> More

  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. << Less

  Chem. Biol. 13:409-419(2006) [PubMed] [EuropePMC]

  This publication is cited by 1 other entry.

• Unconventionally secreted effectors of two filamentous pathogens target plant salicylate biosynthesis.

  Liu T., Song T., Zhang X., Yuan H., Su L., Li W., Xu J., Liu S., Chen L., Chen T., Zhang M., Gu L., Zhang B., Dou D.

  Plant diseases caused by fungi and oomycetes pose an increasing threat to food security and ecosystem health worldwide. These filamentous pathogens, while taxonomically distinct, modulate host defense responses by secreting effectors, which are typically identified based on the presence of signal ... >> More

  Plant diseases caused by fungi and oomycetes pose an increasing threat to food security and ecosystem health worldwide. These filamentous pathogens, while taxonomically distinct, modulate host defense responses by secreting effectors, which are typically identified based on the presence of signal peptides. Here we show that Phytophthora sojae and Verticillium dahliae secrete isochorismatases (PsIsc1 and VdIsc1, respectively) that are required for full pathogenesis. PsIsc1 and VdIsc1 can suppress salicylate-mediated innate immunity in planta and hydrolyse isochorismate in vitro. A conserved triad of catalytic residues is essential for both functions. Thus, the two proteins are isochorismatase effectors that disrupt the plant salicylate metabolism pathway by suppressing its precursor. Furthermore, these proteins lack signal peptides, but exhibit characteristics that lead to unconventional secretion. Therefore, this secretion pathway is a novel mechanism for delivering effectors and might play an important role in host-pathogen interactions. << Less

  Nat. Commun. 5:4686-4686(2014) [PubMed] [EuropePMC]