title: Oligomeric state of hypoxanthine-guanine phosphoribosyltransferase from Mycobacterium tuberculosis.
abstract: Sedimentation equilibrium and size-exclusion chromatography experiments on Mycobacterium tuberculosis hypoxanthine-guanine phosphoribosyltransferase (MtHGPRT) have established the existence of this enzyme as a reversibly associating mixture of dimeric and tetrameric species in 0.1 M Tris-HCl-0.012 M MgCl2, pH 7.4. Displacement of the equilibrium position towards the larger oligomer by phosphate signifies the probable existence of MtHGPRT as a tetramer in the biological environment. These data thus add credibility to the relevance of considering enzyme function in the light of a published tetrameric structure deduced from X-ray crystallography. Failure of 5-phospho-α-d-ribosyl-1-pyrophosphate (PRib-PP) to perturb the dimer-tetramer equilibrium position indicates the equivalence and independence of binding for this substrate (the first to bind in an ordered sequential mechanism) to the two oligomers. By virtue of the displacement of the equilibrium position towards dimer that is affected by removing MgCl2from the Tris-HCl buffer, it can be concluded that divalent metal ions, as well as phosphate, can affect the oligomerization. These characteristics of MtHGPRT in solution are correlated with published crystal structures of four enzyme-ligand complexes.
authors.affiliation: School of Chemistry and Molecular Biosciences, University of Queensland, Brisbane, Queensland 4072, Australia.
authors.forename: Wai Soon
authors.initials: WS
authors.lastname: Eng
authors.affiliation: School of Chemistry and Molecular Biosciences, University of Queensland, Brisbane, Queensland 4072, Australia.
authors.forename: Dianne T
authors.initials: DT
authors.lastname: Keough
authors.affiliation: Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, v.v.i. Flemingovo nam. 2, CZ-166 10 Prague 6, Czechia.
authors.forename: Dana
authors.initials: D
authors.lastname: Hockova
authors.affiliation: School of Chemistry and Molecular Biosciences, University of Queensland, Brisbane, Queensland 4072, Australia.
authors.forename: Donald J
authors.initials: DJ
authors.lastname: Winzor
authors.affiliation: School of Chemistry and Molecular Biosciences, University of Queensland, Brisbane, Queensland 4072, Australia. Electronic address: luke.guddat@uq.edu.au.
authors.forename: Luke W
authors.initials: LW
authors.lastname: Guddat
mesh_terms: D007041:Hypoxanthine Phosphoribosyltransferase / Q000737:chemistry* / Q000378:metabolism*; D015636:Magnesium Chloride / Q000494:pharmacology; D009169:Mycobacterium tuberculosis / Q000201:enzymology*; D011487:Protein Conformation / Q000187:drug effects; D055503:Protein Multimerization / Q000187:drug effects
chemical_list: D015636:Magnesium Chloride; D007041:Hypoxanthine Phosphoribosyltransferase
keywords: Enzyme activity; Hypoxanthine−guanine phosphoribosyltransferase; Mycobacterium tuberculosis; Oligomers; Structural biology; Three-dimensional structure