The overall objective of this project is to determine the high-resolution crystal structure of human erythrocytic purine nucleoside phosphorylase (PNP) and of various nucleoside analog and inhibitor complexes of the enzyme. The crystallographic results will then be used to aid in the design of novel inhibitors of PNP. These inhibitors should prove of practical value, since PNP contributes to the degradation of nucleoside analogs in transit through the bloodstream, and the enzyme is required for T-cell, but not B-cell, activity. The 6 Angstrom structure of the enzyme has now been determined and potential heavy-atom derivatives that can be used in the high-resolution analysis have been identified. In addition, the active site of the enzyme has been characterized at 6 Angstrom resolution by use of PNP complexes with formycin B, 5'-iodoformycin B, 5'-iodo-9-deazainosine and 8-iodoguanine, all of which appear to be suitable for high-resolution studies. Preliminary 3.0 Angstrom data sets have been collected by oscillation photography and synchrotron radiation. Intensity data for the heavy-atom derivatives and analog complexes that were used for the 6 Angstrom structure will be collected to 2.8 Angstrom resolution by use of oscillation photography and synchrotron radiation. These data sets will be used to calculate a 2.8 Angstrom map of the enzyme, and to further characterize the active site of the enzyme. In addition, various nucleoside analog and inhibitor complexes of the enzyme will be studied at 2.8 Angstrom resolution in order to obtain structural information about those features of the active site that are important for the biological mechanisms of the enzyme. This information will be used to guide the synthesis of new types of inhibitors of the enzyme.
| Ealick, S E; Rule, S A; Carter, D C et al. (1990) Three-dimensional structure of human erythrocytic purine nucleoside phosphorylase at 3.2 A resolution. J Biol Chem 265:1812-20 |
| Vijay-Kumar, S; Senadhi, S E; Ealick, S E et al. (1987) Crystallization and preliminary X-ray investigation of a recombinant form of human gamma-interferon. J Biol Chem 262:4804-5 |
| Stoeckler, J D; Ealick, S E; Bugg, C E et al. (1986) Design of purine nucleoside phosphorylase inhibitors. Fed Proc 45:2773-8 |
| Cook, W J; Ealick, S E; Krenitsky, T A et al. (1985) Crystallization and preliminary x-ray investigation of purine-nucleoside phosphorylase from Escherichia coli. J Biol Chem 260:12968-9 |
