title: The Functional Unit of Neisseria meningitidis 3-Deoxy-ᴅ-Arabino-Heptulosonate 7-Phosphate Synthase Is Dimeric.
abstract: Neisseria meningitidis 3-deoxy-D-arabino-heptulosonate 7-phosphate synthase (NmeDAH7PS) adopts a homotetrameric structure consisting of an extensive and a less extensive interface. Perturbation of the less extensive interface through a single mutation of a salt bridge (Arg126-Glu27) formed at the tetramer interface of all chains resulted in a dimeric DAH7PS in solution, as determined by small angle X-ray scattering, analytical ultracentrifugation and analytical size-exclusion chromatography. The dimeric NmeDAH7PSR126S variant was shown to be catalytically active in the aldol-like condensation reaction between D-erythrose 4-phosphate and phosphoenolpyruvate, and allosterically inhibited by L-phenylalanine to the same extent as the wild-type enzyme. The dimeric NmeDAH7PSR126S variant exhibited a slight reduction in thermal stability by differential scanning calorimetry experiments and a slow loss of activity over time compared to the wild-type enzyme. Although NmeDAH7PSR126S crystallised as a tetramer, like the wild-type enzyme, structural asymmetry at the less extensive interface was observed consistent with its destabilisation. The tetrameric association enabled by this Arg126-Glu27 salt-bridge appears to contribute solely to the stability of the protein, ultimately revealing that the functional unit of NmeDAH7PS is dimeric.
authors.affiliation: Biomolecular Interaction Centre and Department of Chemistry, University of Canterbury, Christchurch, New Zealand.
authors.forename: Penelope J
authors.initials: PJ
authors.lastname: Cross
authors.affiliation: Biomolecular Interaction Centre and Department of Chemistry, University of Canterbury, Christchurch, New Zealand.
authors.forename: Logan C
authors.initials: LC
authors.lastname: Heyes
authors.affiliation: Biomolecular Interaction Centre and Department of Chemistry, University of Canterbury, Christchurch, New Zealand.
authors.forename: Shiwen
authors.initials: S
authors.lastname: Zhang
authors.affiliation: Biomolecular Interaction Centre and Department of Chemistry, University of Canterbury, Christchurch, New Zealand.
authors.forename: Ali Reza
authors.initials: AR
authors.lastname: Nazmi
authors.affiliation: Biomolecular Interaction Centre and Department of Chemistry, University of Canterbury, Christchurch, New Zealand.
authors.forename: Emily J
authors.initials: EJ
authors.lastname: Parker
mesh_terms: D010723:3-Deoxy-7-Phosphoheptulonate Synthase / Q000737:chemistry / Q000378:metabolism*; D055162:Biocatalysis; D002850:Chromatography, Gel; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D007700:Kinetics; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics; D009345:Neisseria meningitidis / Q000201:enzymology*; D010649:Phenylalanine / Q000494:pharmacology; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D053838:Scattering, Small Angle; D012996:Solutions; D013997:Time Factors
chemical_list: D012996:Solutions; D010649:Phenylalanine; D010723:3-Deoxy-7-Phosphoheptulonate Synthase
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