CRYSTAL-STRUCTURE OF OROTATE PHOSPHORIBOSYLTRANSFERASE

Phosphoribosyltransferases (PRTases) are enzymes involved in the synth esis of purine, pyrimidine, and pyridine nucleotides. They utilize alp ha-D-5-phosphoribosyl-1-pyrophosphate (PRPP) and a nitrogenous base to form a beta-N-riboside monophosphate and pyrophosphate (PPi), and the ir functional significance in nucleotide homeostasis is evidenced by t he devastating effects of inherited diseases associated with the decre ased activity and/or stability of these enzymes. The 2.6-Angstrom stru cture of the Salmonella typhimurium orotate phosphoribosyltransferase (OPRTase) complexed with its product orotidine monophosphate (OMP) pro vides the first detailed image of a member of this group of enzymes. T he OPRTase three-dimensional structure was solved using multiple isomo rphous replacement methods and reveals two major features: a core five -stranded alpha/beta twisted sheet and an N-terminal region that parti ally covers the C-terminal portion of the core. PRTases show a very hi gh degree of base specificity. In OPRTase, this is determined by steri c constraints and the position of hydrogen bond donors/accepters of a solvent-inaccessible crevice where the orotate ring of bound OMP resid es. Crystalline OPRTase is a dimer, with catalytically important resid ues from each subunit available to the neighboring subunit, suggesting that oligomerization is necessary for its activity. On the basis of t he presence of a common PRPP binding motif among PRTases and the simil ar chemistry these enzymes perform, we propose that the alpha/beta cor e found in OPRTase will represent a common feature for PRTases. This g enerality is demonstrated by construction of a model of the human hypo xanthine-guanine phosphoribosyltransferase (HGPRTase) from secondary s tructure predictions for HGPRTase and the three-dimensional structure of OPRTase.