The structure of the Q69L mutant of GDP-Ran shows a major conformational change in the switch II loop that accounts for its failure to bind nuclear transport factor 2 (NTF2)

Journal of Molecular Biology

Volumn 284, Issue 5, 1998, Pages 1517-1527

  Stewart, Murray ^a   Kent, Helen M ^a   McCoy, Airlie J ^a  

^a MRC LABORATORY OF MOLECULAR BIOLOGY   (United Kingdom)

Author keywords

Conformational change; Molecular interactions; Nucleocytoplasmic transport; Protein structure; X ray crystallography

Indexed keywords

CARRIER PROTEIN; GUANOSINE DIPHOSPHATE; GUANOSINE TRIPHOSPHATASE; MUTANT PROTEIN;

ARTICLE; CONFORMATIONAL TRANSITION; CRYSTAL STRUCTURE; PRIORITY JOURNAL; PROTEIN BINDING; PROTEIN CONFORMATION; PROTEIN PROTEIN INTERACTION; STRUCTURE ACTIVITY RELATION;

EID: 0032545173     PISSN: 00222836     EISSN: None     Source Type: Journal    
DOI: 10.1006/jmbi.1998.2204     Document Type: Article

References (62)

1. Avis, J. M. & Clarke, P. R. (1996). Ran, a small GTPase involved in nuclear processes: its regulators and effectors. J. Cell Sci. 109, 2423-2427.
2. Barbacid, M. (1987). Ras genes. Annu. Rev. Biochem. 56, 779-827.
3. Bischoff, F. R. & Ponstingl, H. (1991a). Mitotic regulator protein RCC1 is complexed with a nuclear ras-related polypeptide. Proc. Natl Acad. Sci. USA, 88, 10830-10834.
4. Bischoff, F. R. & Ponstingl, H. (1991b). Catalysis on guanine nucleotide exchange on Ran by the mitotic regulator RCC1. Nature, 354, 80-82.
5. Bischoff, F. R., Klebe, C., Kretschmer, J., Wittinghofer, A. & Ponstingl, H. (1995). Human RanGTPase activating protein RanGAP1 is a homologue of yeast Rnalp involved in mRNA processing and transport. Proc. Natl Acad. cid. USA, 92, 1749-1753.
6. Bullock, T. L., Clarkson, W. D., Kent, H. M. & Stewart, M. (1996). 1.6 Å structure of nuclear transport factor 2 (NTF2). J. Mol. Biol. 260, 422-431.
7. Clarkson, W. D., Kent, H. M. & Stewart, M. (1996). Separate binding sites on nuclear transport factor 2 (NTF2) for GDP-Ran and the phenylalanine-rich repeat regions of nucleoporins p62 and Nsp1p. J. Mol. Biol. 263, 517-524.
8. Clarkson, W. D., Corbett, A. H., Paschal, B. M., Kent, H. M., McCoy, A. J., Gerace, L., Silver, P. A. & Stewart, M. (1997). Nuclear protein import is decreased by engineered mutants of nuclear transport factor 2 (NTF2) that do not bind GDP-Ran. J. Mol. Biol. 272, 716-730.
9. Computational Project Number 4 (1994). The CCP4 suite: programmes for protein crystallography. Acta Crystallog. sect. D, 50, 760-763.
10. Corbett, A. H. & Silver, P. A. (1996). The NTF2 gene encodes an essential, highly conserved protein that functions in nuclear transport in vivo. J. Biol. Chem. 271, 18477-18484.
11. Corbett, A. H. & Silver, P. A. (1997). Nucleocytoplasmic transport of macromolecules. Microbiol. Mol. Biol. Revs. 61, 193-211.
12. Dickmanns, A., Bischoff, F. R., Marshallsay, C., Lürhmann, R., Ponstingl, H. & Fanning, E. (1996). The thermolability of nuclear protein import in tsBN2 cells is suppressed by microinjected Ran-GTP or Ran-GDP, but not by RanQ69L or RanT24N. J. Cell Sci. 109, 1449-1457.
13. Feldherr, C. M., Akin, A. & Moore, M. S. (1998). The nuclear import factor p10 regulates the functional size of the nuclear pore complex during oocgensis. J. Cell Sci. 111, 1889-1896.
14. Görlich, D. (1998). Transport into and out of the nucleus. EMBO J. 17, 2721-2727.
15. Görlich, D. & Mattaj, I. W. (1996). Nucleocytoplasmic transport. Science, 271, 1513-1518.
16. Görlich, D., Pante, N., Kutay, U., Aebi, U. & Bischoff, F. R. (1996). Identification of different roles for Ran-GDP and Ran-GTP in nuclear protein import. EMBO J. 15, 5584-5594.
17. Grote, M., Kubitscheck, U., Reichelt, R. & Peters, R. (1995). Mapping of nucleoporins to the centre of the nuclear pore complex by post-embedding immunogold electron microscopy. J. Cell Sci. 108, 2963-2972.
18. Hopper, A. K., Traglia, H. M. & Dunst, R. W. (1990). The yeast RNA1 gene product necessary for RNA processing is located in the cytosol and apparently excluded from the nucleus. J. Cell Biol. 111, 309-321.
19. Izaurralde, E., Kutay, U., von Kobbe, C, Mattaj, I. & Görlich, D. (1997). The asymmetric distribution of the constituents of the Ran system is essential for transport into and out of the nucleus. EMBO J. 16, 6535-6547.
20. Jones, T. A., Zou, J.-Y., Cowan, S. W. & Kjeldgaard, M. (1991). Improved methods for building protein models in electron density maps and for locating errors in these models. Acta Crystallog. sect. A, 47, 110-119.
21. Kent, H. M., Clarkson, W. D., Bullock, T. L. & Stewart, M. (1996). Crystallisation and preliminary X-ray diffraction analysis of nuclear transport factor (NTF2). J. Struct. Biol. 116, 325-328.
22. Klebe, C., Prinz, H., Wittinghofer, A. & Goody, R. S. (1995). Interaction of the nuclear GTP-binding protein Ran with its regulatory proteins RCC1 and RanGAP1. Biochemistry, 34, 639-647.
23. Koepp, D. M. & Silver, P. A. (1996). A GTPase controlling nuclear trafficking: running the right way or walking randomly. Cell, 87, 1-4.
24. Kraulis, P. J. (1991). MOLSCRIPT: a program to produce both detailed and schematic plots of protein structures. J. Appl. Crystallog. 24, 946-950.
25. Krengel, U., Schlichting, I., Scherer, A., Schumann, R., Frech, M., John, J., Kabsch, W., Pai, E. & Wittinghofer, A. (1990). Three-dimensional structures of H-ras p21 mutants: molecular basis for their inability to function as signal switch molecules. Cell, 62, 539-548.
26. Kutay, U., Bischoff, F. R., Kostlen, S., Kraft, R. & Görlich, D. (1997). Export of importin α from the nucleus is mediated by a specific nuclear export receptor. Cell, 90, 1061-1071.
27. Loundsbury, K. M., Beddow, A. L. & Macara, I. G. (1994). A family of proteins that stabilize the Ran/TC4 GTPase in its GTP-bound conformation. J. Biol. Chem. 269, 11285-11290.
28. Loundsbury, K. M., Richards, S. A., Perlungberger, R. R. & Macara, I. G. (1996). Ran binding domains promote the interaction of ran with p97/α-karyopherin, linking the docking and translocation steps of nuclear import. J. Biol. Chem. 271, 2357-2360.
29. Mahajan, R., Delphin, C., Guan, T., Gerace, L. & Melchior, F. (1997). A small ubiquitin-related polypeptide involved in targeting RanGAP1 to nuclear pore complex protein RanBP2. Cell, 88, 97-107.
30. Matunis, M. J., Coutavas, E. & Blobel, G. (1996). A novel ubiquitin-like modification modulates the partitioning of the Ran-GTPase-activating protein RanGAP1 between the cytosol and the nuclear pore complex. J. Cell Biol. 135, 1457-1470.
31. Melchior, F. & Gerace, L. (1998). Two-way trafficking with Ran. Trends Cell Biol. 8, 175-179.
32. Melchior, F., Paschal, B. M., Evans, J. & Gerace, L. (1993a). Inhibition of nuclear protein import by non-hydrolyzable analogues of GTP and identification of the small GTPase Ran/TC4 as an essential transport factor. J. Cell Biol. 123, 1649-1659.
33. Melchior, F., Weber, K. & Gerke, V. (1993b). A functional homologue of the RNA1 gene produce in Schizosaccharomyces pombe: purification, biochemical characterization and identification of a leucine-rich repeat motif. Mol. Cell. Biol. 4, 569-581.
34. Melchior, F., Guan, T., Yokoyama, N., Nishimoto, T. & Gerace, L. (1995). GTP hydrolysis by Ran occurs at the nuclear pore complex in an early step of protein import. J. Cell Biol. 131, 571-581.
35. Moore, M. S. & Blobel, G. (1993). The GTP-binding protein Ran/TC4 is required for protein import into the nucleus. Nature, 365, 661-663.
36. Moore, M. S. & Blobel, G. (1994). Purification of a Ran-interacting protein that is required for protein import into the nucleus. Proc. Natl Acad. Sci. USA, 91, 10212-10216.
37. Moroianu, J., Blobel, G. & Radu, A. (1996). Nuclear protein import: Ran-GTP dissociates the karyopherin αβ heterodimer by displacing β from an overlapping binding site on β. Proc. Natl Acad. Sci. USA, 93, 7059-7062.
38. Nehrbass, U. & Blobel, G. (1996). Role of nuclear transport factor p10 in nuclear import. Science, 272, 120-122.
39. Newmeyer, D. D. & Wilson, K. L. (1994). Egg extracts for nuclear import and nuclear assembly reactions. Methods Cell Biol. 36, 607-631.
40. Nigg, E. A. (1997). Nucleocytoplasmic transport: signals mechanism and regulation. Nature, 386, 779-787.
41. Nishimoto, T., Ellen, E. & Basilico, C. (1978). Premature chromosome condensation in a ts DNA-mutant of BHK cells. Cell, 15, 475-483.
42. Nishitani, H., Ohtsubo, M., Yamashita, K., Iida, H., Pines, J., Yasudo, H., Shibata, Y., Hunter, T. & Nishimoto, T. (1991). Loss of RCC1, a nuclear DNA-binding protein, uncouples the completion of DNA replication from the activation of cdc2 protein kinase and mitosis. EMBO J. 10, 1555-1564.
43. Ohtsubo, M., Okazaki, H. & Nishimoto, T. (1989). The RCC1 protein, a regulator for the onset of chromosome condensation, locates in the nucleus and binds to DNA. J. Cell Biol. 109, 1389-1397.
44. Paschal, B. M. & Gerace, L. (1995). Identification of NTF2, a cytosolic factor for nuclear import that interacts with nuclear pore complex protein p62. J. Cell Biol. 129, 925-937.
45. Paschal, B. M., Delphin, C. & Gerace, L. (1996). Nucleotide-specific interaction of Ran/TC4 with nuclear transport factors NTF2 and p97. Proc. Natl Acad. Sci. USA, 93, 7679-7683.
46. Pascal, B. M., Fritz, C., Guan, T. & Gerace, L. (1997). High levels of the GTPase Ran/TC4 relieve the requirement for nuclear protein transport factor 2. J. Biol. Chem. 272, 21534-21539.
47. Pennisi, E. (1988). The nucleus's revolving door. Science, 279, 1129-1131.
48. Ren, M., Drivas, G., D'Eustachino, P. & Rush, M. G. (1993). Ran/TC4: a small nuclear GTP-binding protein that regulates DNA synthesis. J. Cell Biol. 120, 313-323.
49. Ren, M., Villamarin, A., Shih, A., Coutavas, E., Moore, M. S., LoCurcio, M., Clarke, V., Oppenheim, J. D., D'Eustachio, P. & Rush, M. G. (1995). Separate domains of the Ran GTPase interact with different factors to regulate nuclear protein import and RNA processing. Mol. Cell. Biol. 15, 2117-2124.
50. Rexach, M. & Blobel, G. (1995). Protein import into nuclei: association and dissociation reactions involving transport substrate, transport factors and nucleoporins. Cell, 83, 683-692.
51. Rittinger, K., Walker, A. P., Eccleston, J. F., Smerdon, S. K. & Gamblin, S. J. (1997). Structure at 1.65 Å of RhoA and its GTPase-activating protein in complex with a transition-state analogue. Nature, 389, 758-762.
52. Rush, M. G., Drivas, G. & D'Eustachio, P. (1996). The small nuclear GTPase Ran: how much does it run? Bioessays, 18, 103-112.
53. Scheffzek, K., Klebe, C., Fritz-Wolf, K., Kabsch, W. & Wittinghofer, A. (1995). Crystal structure of the nuclear Ras-related protein Ran in its GDP-bound form. Nature, 374, 378-381.
54. Scheffzek, K., Ahmadian, M. R., Kabsch, W., Weismüller, L., Lautwein, A., Schmitz, F. & Wittinghofer, A. (1997). The Ras-RasGAP complex: structural basis for GTPase activation and its loss in oncogenic ras mutants. Science, 277, 333-338.
55. + RNA in the cytoplasm. Proc. Natl Acad. Sci. USA, 92, 225-229.
56. Stewart, M., Kent, H. M. & McCoy, A. J. (1998). Structural basis for molecular recognition between nuclear transport factor 2 (NTF2) and the GDP-bound form of the Ras-family GTPase Ran. J. Mol. Biol. 277, 635-646.
57. Sweet, D. J. & Gerace, L. (1996). A GTPase distinct from Ran is involved in nuclear protein import. J. Cell Biol. 133, 971-983.
58. Tachibana, T., Imamoto, N., Seino, H., Nishimoto, T. & Yoneda, Y. (1994). Loss of RCC1 leads to suppression of nuclear protein import in living cells. J. Biol. Chem. 269, 24542-24545.
59. Weis, K. (1998). Importins and exportins: how to get in and out of the nucleus. Trends Biochem. Sci. 23, 185-189.
60. Weis, K., Dingwall, C. & Lamond, A. (1996). Characterization of the nuclear protein import mechanism using Ran mutants with altered nucleotide binding specificities. EMBO J. 15, 7120-7128.
61. Wittinghofer, A. & Pai, E. (1991). The role of Ras protein: a model for a universal molecular switch. Trends Biochem. Sci. 16, 383-387.
62. Wong, D. H., Corbett, A. H., Kent, H. M., Stewart, M. & Silver, P. A. (1997). Interaction between the small GTPase Ran/Gsplp and Ntf2p is required for nuclear transport. Mol. Cell Biol. 17, 3755-3766.