title: The structure of a triple mutant of pI258 arsenate reductase from Staphylococcus aureus and its 5-thio-2-nitrobenzoic acid adduct.
abstract: Structural insights into formation of the complex between the ubiquitous thiol-disulfide oxidoreductase thioredoxin and its oxidized substrate are under-documented owing to its entropical instability. In vitro, it is possible via a reaction with 5,5'-dithiobis-(2-nitrobenzoic acid) to make a stable mixed-disulfide complex between thioredoxin from Staphylococcus aureus and one of its substrates, oxidized pI258 arsenate reductase (ArsC) from S. aureus. In the absence of the crystal structure of an ArsC-thioredoxin complex, the structures of two precursors of the complex, the ArsC triple mutant ArsC C10SC15AC82S and its 5-thio-2-nitrobenzoic acid (TNB) adduct, were determined. The ArsC triple mutant has a structure very similar to that of the reduced form of wild-type ArsC, with a folded redox helix and a buried catalytic Cys89. In the adduct form, the TNB molecule is buried in a hydrophobic pocket and the disulfide bridge between TNB and Cys89 is sterically inaccessible to thioredoxin. In order to form a mixed disulfide between ArsC and thioredoxin, a change in the orientation of the TNB-Cys89 disulfide in the structure is necessary.
authors.affiliation: Laboratorium voor Ultrastructuur, Vlaams Interuniversitair Instituut voor Biotechnologie (VIB), Vrije Universiteit Brussel, Pleinlaan 2, B-1050 Brussel, Belgium. joris.messens@vub.ac.be
authors.forename: Joris
authors.initials: J
authors.lastname: Messens
authors.forename: Inge
authors.initials: I
authors.lastname: Van Molle
authors.forename: Peter
authors.initials: P
authors.lastname: Vanhaesebrouck
authors.forename: Karolien
authors.initials: K
authors.lastname: Van Belle
authors.forename: Khadija
authors.initials: K
authors.lastname: Wahni
authors.forename: José C
authors.initials: JC
authors.lastname: Martins
authors.forename: Lode
authors.initials: L
authors.lastname: Wyns
authors.forename: Remy
authors.initials: R
authors.lastname: Loris
mesh_terms: D053501:Arsenite Transporting ATPases; D018360:Crystallography, X-Ray; D019281:Dimerization; D004220:Disulfides / Q000737:chemistry; D004583:Electrons; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D006639:Histidine / Q000737:chemistry; D016623:Ion Pumps / Q000737:chemistry*; D013058:Mass Spectrometry; D008958:Models, Molecular; D009097:Multienzyme Complexes / Q000737:chemistry*; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009579:Nitrobenzoates / Q000737:chemistry*; D010084:Oxidation-Reduction; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D013211:Staphylococcus aureus / Q000201:enzymology*; D013438:Sulfhydryl Compounds
chemical_list: D004220:Disulfides; D016623:Ion Pumps; D009097:Multienzyme Complexes; D009579:Nitrobenzoates; D013438:Sulfhydryl Compounds; C011136:thionitrobenzoic acid; C033223:polyhistidine; D006639:Histidine; D053501:Arsenite Transporting ATPases