Disease relevance of ECs0991

• Furthermore, an N-terminal portion of the herbicide resistance gene 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) containing a chloroplast localization signal fused to In (EPSPSn-In) was integrated into the nuclear DNA of N. tabacum by using Agrobacterium tumefaciens-mediated transformation [1].
• 5-Enolpyruvylshikimate-3-phosphate synthase was prepared from E. coli cells harboring this construct [2].
• The feasibility of using Salmonella typhimurium aroA mutant (SL3261) to deliver protein therapeutic agents was investigated in a murine model system [3].
• Structural studies of Streptococcus pneumoniae EPSP synthase in unliganded state, tetrahedral intermediate-bound state and S3P-GLP-bound state [4].
• Molecular characterization of the Aeromonas hydrophila aroA gene and potential use of an auxotrophic aroA mutant as a live attenuated vaccine [5].

High impact information on ECs0991

• The remaining EPSPS gene fragment (EPSPSc) fused to Ic (Ic-EPSPSc) was integrated into the chloroplast genome by homologous recombination [1].
• Western blot analysis of cell extracts from these plants showed a full-length EPSPS, demonstrating that the EPSPSn-In gene product migrated to the chloroplast and underwent trans-splicing [1].
• Structure and topological symmetry of the glyphosate target 5-enolpyruvylshikimate-3-phosphate synthase: a distinctive protein fold [6].
• A new view of the mechanisms of UDP-N-acetylglucosamine enolpyruvyl transferase (MurA) and 5-enolpyruvylshikimate-3-phosphate synthase (AroA) derived from X-ray structures of their tetrahedral reaction intermediate states [7].
• BALB/c mice administered orally or i.v. with S. typhimurium aroA mutants carrying the plasmid produced highly significant antibody responses against human IL-1 beta as determined by a solid-phase RIA [3].

Chemical compound and disease context of ECs0991

• Evidence for a reactive gamma-carboxyl group (Glu-418) at the herbicide glyphosate binding site of 5-enolpyruvylshikimate-3-phosphate synthase from Escherichia coli [8].
• Reaction of 5-enolpyruvylshikimate-3-phosphate synthase of Escherichia coli with the thiol reagent 5,5'-dithiobis-(2-nitrobenzoic acid) (DTNB) leads to a modification of only 2 of the 6 cysteines of the enzyme, with a significant loss of its enzymatic activity [9].
• The glycine to alanine substitution corresponding to Escherichia coli EPSPS G96A increases the Ki(app) (glyphosate) of petunia EPSPS 5000-fold while increasing the Km(app)(phosphoenolpyruvate) about 40-fold [10].
• To obtain chloroplast delivery of the 43-kDa 5-enolpyruvyl 3-phosphoshikimate (EPSP) synthase of Salmonella typhimurium, we constructed fusion proteins between the bacterial EPSP synthase and the ribulose bisphosphate carboxylase small subunit [11].
• These results clearly establish the nonessentiality of the active site-reactive cysteine of E. coli 5-enolpyruvylshikimate-3-phosphate synthase for either catalysis or substrate binding [9].

Biological context of ECs0991

• Site-directed mutagenesis of a conserved region of the 5-enolpyruvylshikimate-3-phosphate synthase active site [10].
• A single amino acid substitution in the enzyme 5-enolpyruvylshikimate-3-phosphate synthase confers resistance to the herbicide glyphosate [2].
• Sequence comparisons of the predicted EPSPS mature protein coding regions from both biotypes revealed four single-nucleotide differences, two of which result in amino acid changes [12].
• The nucleotide sequence of the A. hydrophila aroA gene encoded a protein of 440 amino acids which showed a high degree of homology to other bacterial AroA proteins [5].
• A tetrahedral intermediate in the EPSP synthase reaction observed by rapid quench kinetics [13].

Anatomical context of ECs0991

• Since the expressed EPSPS protein was found as an insoluble form in the inclusion body, it was extracted by 6 M urea after sonication, and then purified through immobilized nickel-affinity column chromatography to isolate EPSPS having a molecular mass of 57 kDa [14].

Associations of ECs0991 with chemical compounds

• Substitution of this glycine with serine, however, abolishes EPSPS activity but results in the elicitation of a novel EPSP hydrolase activity whereby EPSP is converted to shikimate 3-phosphate and pyruvate [10].
• This highly conserved region is critical for the interaction of the phosphate moiety of phosphoenolpyruvate with EPSPS [10].
• Under denaturing conditions, however, all 6 cysteines of 5-enolpyruvylshikimate-3-phosphate synthase react with DTNB, indicating the absence of disulfide bridges in the native protein [9].
• Incubation of 5-enolpyruvylshikimate-3-phosphate synthase, a target for the nonselective herbicide glyphosate (N-(phosphonomethyl)glycine), with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide in the presence of glycine ethyl ester resulted in a time-dependent loss of enzyme activity [8].
• As part of a solid-state NMR characterization of this structure, 15N labels were introduced selectively into the lysine, arginine and histidine residues of EPSP synthase and distances to a 13C label in Glp and to the 31P in S3P and Glp were measured by rotational-echo double-resonance NMR [15].

Other interactions of ECs0991

• UDP-N-acetylglucosamine enolpyruvyl transferase (MurA) and 5-enolpyruvylshikimate-3-phosphate synthase (AroA) constitute the small enzyme family of enolpyruvyl transferases, which catalyze the chemically unusual reaction of enolpyruvyl transfer [7].
• Subfragments of the Arom locus encoding EPSP synthase and 3-dehydroquinase have been expressed in appropriate E. coli aro mutants [16].

Analytical, diagnostic and therapeutic context of ECs0991

• Site-directed mutagenesis of Petunia hybrida 5-enolpyruvylshikimate-3-phosphate synthase: Lys-23 is essential for substrate binding [17].
• Crystallization of 5-enolpyruvylshikimate 3-phosphate synthase from Escherichia coli [18].
• Using this system, the rice (Oryza sativa) EPSPS gene, mutagenized by Error-Prone polymerase chain reaction, was introduced into an EPSPS-deficient Escherichia coli strain, AB2829, and transformants were selected on minimal medium by functional complementation [19].
• EPSP synthase: binding studies using isothermal titration microcalorimetry and equilibrium dialysis and their implications for ligand recognition and kinetic mechanism [20].
• Analytical ultracentrifugation data ruled out GLP-binding synergy mechanisms that derive from, or are influenced by, changes in oligomerization of S. pneumoniae EPSPS [21].

References