The purine nucleotides play a key role in synthesis of DNA, RNA and ATP supply. Living organisms can produce purine nucleotides de novo or via the salvage pathway. Two pathways are inter-related, share alpha-D-phosphoribosyl-1-pyrophosphate (PRPP) as a common substrate and may impose regulation on each other. Many pathogenic organisms are deficient in de novo synthesis of the purine nucleotides, and therefore must rely on the salvage pathway. The key enzyme in the latter pathway is a hypoxanthine-guanine phosphoribosyltransferase (HGPRT, E.C. 22.214.171.124), which catalyzes a reversible transfer of the 5-phosphoribosyl group from PRPP to the N9 atom of either hypoxanthine or guanine to form inosine 5'-monophosphate (IMP) or guanosine 5'-monophosphate (GMP), respectively. It is of scientific and medical interest to understand the mechanism of the HGPRT-catalyzed reaction, because it will help to develop inhibition strategies of the enzyme, and hence create therapeutic agents for treatment of diseases caused by human pathogens.
We have solved 2.06 Å resolution crystal structure of HGPRT (hpt-1) from a human pathogen the Bacillus anthracis str. 'Ames Ancestor'. The hpt-1 enzyme is tetrameric in the crystal lattice. The enzyme possesses two domains (a core and a hood domain) that are common for this class of proteins. Conserved residues from both domains contribute to the formation of the active site. Two phosphate ions that are at ~9.5 Å distance away per each subunit have been modeled into the density. One of these phosphates occupies the same position as the phosphate group of IMP in the 1YFZ structure, which was used as a molecular replacement model. Molecules A and B of the hpt-1 enzyme are associated with sucrose whit its glucose part mimicking the base of IMP in 1YFZ. Crystals of hpt-1 were soaked in sucrose for cryo protection. Additional sucrose molecule is found in the asymmetric unit on the outer surface of the protein. The B. anthracis HGPRT was co-crystallized with GMP. Data collection is underway. Comparison of GMP-bound and apo-form of the hpt-1 protein may provide new insights on product release, possible feedback inhibition, substrate recognition and drug development.