title: Structure and function of the dihydropteroate synthase from Staphylococcus aureus.
abstract: The gene encoding the dihydropteroate synthase of staphylococcus aureus has been cloned, sequenced and expressed in Escherichia coli. The protein has been purified for biochemical characterization and X-ray crystallographic studies. The enzyme is a dimer in solution, has a steady state kinetic mechanism that suggests random binding of the two substrates and half-site reactivity. The crystal structure of apo-enzyme and a binary complex with the substrate analogue hydroxymethylpterin pyrophosphate were determined at 2.2 A and 2.4 A resolution, respectively. The enzyme belongs to the group of "TIM-barrel" proteins and crystallizes as a non-crystallographic dimer. Only one molecule of the substrate analogue bound per dimer in the crystal. Sequencing of nine sulfonamide-resistant clinical isolates has shown that as many as 14 residues could be involved in resistance development. The residues are distributed over the surface of the protein, which defies a simple interpretation of their roles in resistance. Nevertheless, the three-dimensional structure of the substrate analogue binary complex could give important insight into the molecular mechanism of this enzyme.
authors.affiliation: F. Hoffmann-La Roche Ltd, Pharma Preclinical Research Department, Basel, Switzerland.
authors.forename: I C
authors.initials: IC
authors.lastname: Hampele
authors.forename: A
authors.initials: A
authors.lastname: D'Arcy
authors.forename: G E
authors.initials: GE
authors.lastname: Dale
authors.forename: D
authors.initials: D
authors.lastname: Kostrewa
authors.forename: J
authors.initials: J
authors.lastname: Nielsen
authors.forename: C
authors.initials: C
authors.lastname: Oefner
authors.forename: M G
authors.initials: MG
authors.lastname: Page
authors.forename: H J
authors.initials: HJ
authors.lastname: Schönfeld
authors.forename: D
authors.initials: D
authors.lastname: Stüber
authors.forename: R L
authors.initials: RL
authors.lastname: Then
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D004094:Dihydropteroate Synthase / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D004352:Drug Resistance, Microbial / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D007700:Kinetics; D008826:Microbial Sensitivity Tests; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013211:Staphylococcus aureus / Q000187:drug effects / Q000201:enzymology*; D013420:Sulfamethoxazole / Q000494:pharmacology; D013449:Sulfonamides / Q000494:pharmacology
chemical_list: D011994:Recombinant Proteins; D013449:Sulfonamides; D004094:Dihydropteroate Synthase; D013420:Sulfamethoxazole