Book of Abstracts: Albany 2003

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Conversation 13
Abstract Book
June 17-21 2003

Crystal Structure of Purine Nucleoside Phosphorylase from Thermus Thermophilus

Purine nucleoside phosphorylases (PNPs) participate in the biosynthesis of purine nucleosides by salvage pathway. A proposed reversible enzymatic reaction of the glycosidic bond cleavage of purine ribo- and deoxyribonucleosides and their derivatives occurs by phosphorolyses:

purine nucleoside+phosphate·ribose-1-phosphate+purine base

PNPs belong to recently defined nucleoside phosphorylase-I (NP-I) family (1). The PNPs of NP-I family further divided into two groups on the basis of their amino acid sequence homology, tertiary structure organization and substrate specificity. The homotrimeric PNPs are found mainly, but not exclusively, in mammalian species and specific for guanine and hypoxanthine (2?-deoxy)ribonucleosides and their analogs, while the homohexameric PNPs are dominant form in bacterial species and specific for a wider range of substrates since in addition to 6-oxopurine nucleosides they can also accept 6-aminopurine nucleosides. Because of critical role of PNPs in T-cell proliferation the inhibitors of PNPs can be useful for treatment of T-cell leukemias and lymphomas and other T-cell related autoimmune diseases such as psoriasis, rheumatoid arthritis, multiple sclerosis and inflammatory bowel disorders.

Alignment of the primary structures of PNPs from Thermus thermophilus (TtPNP) and other organisms indicated that the TtPNP belongs to hexameric group of NP-I family. TtPNP shares high amino acid sequence identity of 40% with Escherichia coli PNP and Sulfolobus solfataricus 5'-deoxy-5'-methylthioadenosine phosphorylase for which the crystal structures have been reported (2-5). Also the phosphate and ribose binding residues are conserved in all these three enzymes, their key residues for purine base recognition appear to be different. To exploit the significance of the differences in and around the active site of TtPNP and other PNPs observed at primary structure level we determined the high resolution crystal structures of TtPNP in apo form, sulfate bound form and sulfate with adenosine or guanosine bound forms, and carried out a comparative analysis using the reported structures PNPs. The obtained information about the novel base recognition mode of TtPNP can be useful for the design of effective PNP inhibitors.

Tahir H. Tahirov1,*
Eiji Inagaki1
Seiki Kuramitsu2,4
Shigeyuki Yokoyama2,3,5,6
Masashi Miyano1

1Highthroughput Factory
2Structurome Research Group
3Cellular Signaling Laboratory
RIKEN Harima Institute
1-1-1 Kouto
Mikazuki-cho, Sayo-gun
Hyogo 679-5148, Japan
4Department of Biology
Graduate School of Science
Osaka University
Toyonaka, Osaka 560-0043, Japan
5RIKEN Genomic Sciences Center
1-7-22 Suehiro-cho
Tsurumi, Yokohama 230-0045, Japan
6Department of Biophysics and Biochemistry
Graduate School of Science
University of Tokyo
7-3-1 Hongo
Bunkyo-ku, Tokyo 113-0033, Japan

References and Footnotes
  1. Pugmire M. J., and Ealick S. E., Biochemistry Journal 361, 1-25 (2002).
  2. Appleby T. C., Mathews I. I., Porcelli M., Cacciapuoti G., and Ealick S. E., Journal of Biological Chemistry 276, 39232-39242 (2001).
  3. Koellner G., Bzowska A., Wielgus-Kutrowska B., Luic M., Steiner T., Saenger W., and Stepinski J., Journal of Molecular Biology 315, 351-371 (2002).
  4. Mao C., Cook W. J., Zhou M., Koszalka G. W., Krenitsky T. A., and Ealick S. E., Structure 5, 1373-1383 (1997).
  5. Koellner G., Luic M., Shugar D., Saenger W., Bzowska A. Journal of Molecular Biology 265, 202-216 (1997).