Orotate phosphoribosyltransferase and hypoxanthine/guanine phosphoribosyltransferase from yeast: nuclear magnetic relaxation studies of the structures of enzyme-bound phosphoribosyl 1-pyrophosphate.
Nuclear magnetic relaxation rate measurements have been performed on the protons and phosphorus atoms of phosphoribosyl 1-pyrophosphate (PRibPP) in the presence and absence of paramagnetic chromium(III), cobalt(II), and manganese(II) ions. The longitudinal relaxation rates were then used to calculate interatomic distances between the magnetic nuclei and these paramagnetic probes, from which was devised a conformation of the PRibPP-metal ion complex in solution. Thereafter, the experiments were accomplished in the presence of Mn(II) and a series of orotate phosphoribosyltransferase (OPRTase) and hypoxanthine/guanine phosphoribosyltransferase (HGPRTase) concentrations, and from these data were estimated the distances between Mn(II) and the PRibPP nuclei at the active sites of these two enzymes from yeast. Comparisons between the Mn(II)-PRibPP conformation in solution and this structure at the active sites of OPRTase and HGPRTase revealed that the metal ion remained coordinated with the pyrophosphate group of PRibPP in all instances, whereas the overall distances between the ribose ring and Mn(II) at the enzyme active sites were approximately 1 A further from the metal ion. Model building studies also revealed that the 5'-phosphate group of PRibPP is positioned directly over the ribose ring in solution and at the OPRTase and HGPRTase active sites and may protect the 1'-carbon of PRibPP against on-line displacements of pyrophosphate under these conditions, where the PRibPP-to-Mn(II) concentration ratio is greater than 2000.(ABSTRACT TRUNCATED AT 250 WORDS)
D007041:Hypoxanthine Phosphoribosyltransferase / Q000378:metabolism*; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008958:Models, Molecular; D008968:Molecular Conformation; D009962:Orotate Phosphoribosyltransferase / Q000378:metabolism*; D010428:Pentosephosphates / Q000378:metabolism*; D010430:Pentosyltransferases / Q000378:metabolism*; D010754:Phosphoribosyl Pyrophosphate / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology*
D010428:Pentosephosphates; D010754:Phosphoribosyl Pyrophosphate; D010430:Pentosyltransferases; D009962:Orotate Phosphoribosyltransferase; D007041:Hypoxanthine Phosphoribosyltransferase