Crystallography Journals Online : IUCR Backfile Archive 1948-2001
Chemistry and Pharmacology
The calf spleen purine nucleoside phosphorylase (PNP) ternary complex with an N(7)-acycloguanosine inhibitor and a phosphate ion has been crystallized in the cubic space group P213, with unit-cell parameter a = 94.11 Å and one monomer per asymmetric unit. X-ray diffraction data were collected using synchrotron radiation (Station X31, EMBL Outstation, DESY, Hamburg). The crystal structure was refined to a resolution of 2.2 Å and R and Rfree values of 17.5 and 24.5%, respectively. The acyclonucleoside inhibitor is bound in the active site in an inverted (`upside-down') orientation of the purine base compared with natural substrates. The side chain of Asp243 forms two hydrogen bonds with the base ring: Nδ donates a hydrogen to N(3) and Oδ accepts a hydrogen from the guanine N(2)-amino group. N(1)—H of the base is hydrogen bonded to Oε of Glu201, while N(9) accepts a hydrogen bond from Thr242 Oγ. In addition, a water molecule (W417) bridges the N(2)-amino group of the base and Oε of Glu201. In the phosphate-binding site, a phosphate ion is bound to Ser33, His64, Arg84, His86, Ala116 and Ser220. The acyclic chain of the N(7)-acycloguanosine inhibitor is in a folded conformation and together with a water molecule (W388) occupies the pentose-binding site, with possible hydrogen bonds to Tyr88 Oη and His257 Nδ1. This new binding mode fully accounts for the previously observed substrate properties of 7-β-D-ribofuranosides of hypoxanthine and guanine. It also provides a new starting point for the design of inhibitors of PNP for therapeutic and other applications.
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