Crystal Structures of Rhizopuspepsin/Inhibitor Complexes

  • Kevin D. Parris
  • Dennis J. Hoover
  • David R. Davies
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 306)


The crystal structures of the aspartic proteinases have been extensively studied over the past fifteen years (Hsu et al., 1977; Subramanian et al., 1977; reviewed by Davies, 1990). After the initial determinations of the native structures uncomplexed with inhibitors, a few complexes with pepstatin (Bott et al., 1982) and with a fragment of pepstatin (James et al., 1982) were reported. These studies showed that these inhibitors bound in the deep groove that separates the N- and C-terminal domains of the enzyme. Accompanying the binding of the inhibitor was a displacement of the “flap” region of the molecule, a hairpin loop that closes down on the inhibitor, the extent of the displacement depending on the initial location of the flap (James et al., 1982; Bott et al., 1982; Suguna et al., 1987 and Cooper et al., 1987). Since no other major conformational changes were observed, the crystals of these proteinases offered a convenient vehicle for examining a number of bound inhibitor conformations. Other factors facilitating the examination of these inhibitors were the availability of large numbers of renin inhibitors, and the fortunate ease of access to the combining site in several of the crystal forms, thus enabling the inhibitors to be soaked into the crystals.


Aspartic Proteinase Solvent Accessibility Renin Inhibitor Tetrahedral Intermediate Domain Rotation 


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  1. Abad-Zapatero, C., Rydel, T. J. & Erickson, J., 1990, Revised 2.3Å Structure of Porcine Pepsin: Evidence for a Flexible Subdomain, Proteins, Structure, Function and Genetics, 8: 62.CrossRefGoogle Scholar
  2. Agarwal, R. C., 1978, A new least squares refinement technique based on the fast Fourier transform algorithm, Acta Crystallogr., Section A, 34: 791.CrossRefGoogle Scholar
  3. Blundell, T. L., Cooper, J., Foundling, S. I., Jones, D. M., Atrash, B. & Szelke, M., 1987, On the rational design of Renin inhibitors: X-ray studies of aspartic proteinases complexed with transition-state analogues, Biochemistry, 26: 5585.PubMedCrossRefGoogle Scholar
  4. Bott, R., Subramanian, E. & Davies D. R., 1982, Three-dimensional structure of the complex of the Rhizopus chinensis carboxyl proteinase and pepstatin at 2.5Å resolution, Biochemistry, 21: 6956.PubMedCrossRefGoogle Scholar
  5. Bott, R., & Davies D. R., 1983, Pepstatin binding to Rhizopus chinensis aspartyl proteinase, in: “Peptides: Structure and Function”, V. J. Hruby and D. H. Rich, eds., Pierce Chemical Company.Google Scholar
  6. Case, D. A. & Karplus, M., 1979, Dynamics of ligand binding to heme proteins, J. Mol. Biol., 132: 343.PubMedCrossRefGoogle Scholar
  7. Connolly, M. L., 1983, Analytical molecular surface calculation. J. Appl. Crystallogr., 16: 548.CrossRefGoogle Scholar
  8. Cooper, J., Foundling, S. I., Hemmings, A., Blundell, T. L., Jones, D. M., Hallett, A. & Szelke, M., 1987, X-ray studies of aspartic proteinases-statine complexes, Eur. J. Biochemistry, 169: 215.CrossRefGoogle Scholar
  9. Davies, D. R., 1990, The structure and function of the aspartic proteinases, Ann. Rev. Biophys. Biophys. Chem., 19: 189.CrossRefGoogle Scholar
  10. Finzel, B. C., 1987, Incorporation of fast Fourier transforms to speed least squares refinement of protein structures, J. Appl. Crystallogr., 20: 53.CrossRefGoogle Scholar
  11. Fruton, J. S., 1976, Mechanism of the catalytic action of pepsin and related acid proteinases, Advances in Enzymology, 44: 1.Google Scholar
  12. Green, D. W., Aykent, S., Gierse, J. K. & Zupec, M. E., 1990, Substrate specificity of recombinant human renal renin: Effect of histidine in the P2 subsite on pH dependence, Biochemistry, 29: 3126.PubMedCrossRefGoogle Scholar
  13. Hendrickson, W. A., 1985, Stereochemically restrained refinement of macromolecular structures, in: “Methods in Enzymology,” H. W. Wyckoff, C. W. H. Hirs, S. N. Timasheff, eds., Academic Press, New York.Google Scholar
  14. Hofmann, T., Allen, B., Bendiner, M., Blum, M. & Cunningham, A., 1988, Effect of secondary substrate binding in penicillopepsin: contribution of subsites S3 and S2 to kcat, Biochemistry, 27: 1140.PubMedCrossRefGoogle Scholar
  15. Hsu, I.-N., Delbaere, L. T. J. & James, M. N. G., 1977, Penicillopepsin from P. janthinellum. Crystal structure at 2.8Å and sequence homology with porcine pepsin, Nature (London), 266: 140.CrossRefGoogle Scholar
  16. Hofmann, T., & Fink, A. L., 1984, Cryoenzymology of Penicillopepsin, Biochemistry, 23: 5247.PubMedCrossRefGoogle Scholar
  17. James, M. N. G. & Sielecki, A. R., 1985, Stereochemical analysis of peptide bond hydrolysis catalyzed by the aspartic proteinase penicillopepsin, Biochemistry, 24: 3701.PubMedCrossRefGoogle Scholar
  18. James, M. N. G., Sielecki, A., Salituro, F., Rich, D. H. & Hofmann, T., 1982, Conformational flexibility in the active sites of aspartyl proteinases revealed by a pepstatin fragment binding to penicillopepsin, Proc. Natl. Acad. Sci., U.S.A., 79: 6137.PubMedCrossRefGoogle Scholar
  19. Jones, T. A.,1978, A graphics model building and refinement system for macromolecules, J. Appl. Crystallogr, 11: 268.CrossRefGoogle Scholar
  20. Pearl, L., 1985, The extended binding cleft of aspartic proteinases and its role in peptide hydrolysis, in: “Aspartic Proteinases and their Inhibitors,” V. Kostka, ed., Walter de Gruyter, Berlin, New York.Google Scholar
  21. Pearl, L., 1987, The catalytic mechanism of aspartic proteinases, FEBS Letters, 214: 8.PubMedCrossRefGoogle Scholar
  22. Pearl, L. H. & Blundell, T. L., 1985, The active site of aspartic proteinases, FEBS Letters, 174: 96.CrossRefGoogle Scholar
  23. Pflugrath, J. W., Saper, M. A. & Quiocho, F. A., 1984, in: “Methods and Applications in Crystallographic Computing,” S. Hall and T. Ashida, eds., Clarendon, Oxford.Google Scholar
  24. Sali, A., Veerapandian, B., Cooper, J. B., Foundling, S. I., Hoover, D. J. & Blundell, T. L., 1989, High-resolution X-ray diffraction study of the complex between endothiapepsin and an oligopeptide inhibitor: the analysis of the inhibitor binding and description of the rigid body shift in the enzyme, EMBO J., 8: 2179.PubMedGoogle Scholar
  25. Satow, Y., Cohen, G. H., Padlan, E. A. & Davies, D. R., 1986, Phosphocholine binding immunoglobulin Fab McPC603: An X-ray diffraction study at 2.7Å, J. Mol. Biol., 190: 593.PubMedCrossRefGoogle Scholar
  26. Schechter, I. and Berger, A., 1967, On the size of the active site in proteases. I. Papain, Biochem. Biophys. Res. Commun., 27: 157.PubMedCrossRefGoogle Scholar
  27. Sheriff, S., Hendrickson, W. A., Stenkamp, R. E., Sieker, L. C. & Jensen, L. H., 1985, Influence of solvent accessibility and intermolecular contacts on atomic mobilities in hemerythrin, Proc. Natl. Acad. Sci. U.S.A., 82: 1104.PubMedCrossRefGoogle Scholar
  28. Sheriff, S., 1987, Addition of symmetry-related contact restraints to PROTIN and PROLSQ, J. Appl. Crystallogr., 20: 53.CrossRefGoogle Scholar
  29. Sielecki, A. R., Fedorov, A. A., Boodhoo, A., Andreeva, N. S. & James, M. N. G., 1990, Molecular and Crystal Structures of Monoclinic Porcine Pepsin refined at 1.8Å Resolution, J. Mol. Biol., 214: 143.PubMedCrossRefGoogle Scholar
  30. Subramanian, E., Swan, I. D. A., Liu, M., Davies, D. R., Jenkins, J. A., Tickle, I. J. & Blundell, T. L., 1977, Homology among acid proteases: Comparison of crystal structures at 3Å resolution of acid proteases from Rhizopus chinensis and Endothia parasitica., Proc. Natl. Acad. Sci., U.S.A., 74: 556.PubMedCrossRefGoogle Scholar
  31. Suguna, K., Padlan, E. A., Smith, C. W., Carlson, W. D. & Davies, D. R., 1987, Binding of a reduced peptide inhibitor to the aspartic proteinase from Rhizopus chinensis: implications for a mechanism of action. Proc. Natl. Acad. Sci., U.S.A., 74: 556.Google Scholar
  32. Suguna, K., Padlan, E. A., Bott, R., Boger, J. & Davies, D. R., 1991, Proteins: Structure, Function and Genetics, in press.Google Scholar

Copyright information

© Plenum Press, New York 1991

Authors and Affiliations

  • Kevin D. Parris
    • 1
  • Dennis J. Hoover
    • 2
  • David R. Davies
    • 1
  1. 1.Laboratory of Molecular BiologyNIDDK, NIHBethesdaUSA
  2. 2.Department of Medicinal Chemistry, Central Research DivisionPfizer IncGrotonUSA

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