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High resolution crystal structures of human Rab5a and five mutants with substitutions in the catalytically important phosphate-binding loop.

Zhu G, Liu J, Terzyan S, Zhai P, Li G, Zhang XC
J Biol Chem 278 2452-60 (2003)
Related entries: 1n6i, 1n6k, 1n6l, 1n6n, 1n6o, 1n6p, 1n6r

Cited: 42 times
EuropePMC logo PMID: 12433916

Abstract

GTPase domain crystal structures of Rab5a wild type and five variants with mutations in the phosphate-binding loop are reported here at resolutions up to 1.5 A. Of particular interest, the A30P mutant was crystallized in complexes with GDP, GDP+AlF(3), and authentic GTP, respectively. The other variant crystals were obtained in complexes with a non-hydrolyzable GTP analog, GppNHp. All structures were solved in the same crystal form, providing an unusual opportunity to compare structures of small GTPases with different catalytic rates. The A30P mutant exhibits dramatically reduced GTPase activity and forms a GTP-bound complex stable enough for crystallographic analysis. Importantly, the A30P structure with bound GDP plus AlF(3) has been solved in the absence of a GTPase-activating protein, and it may resemble that of a transition state intermediate. Conformational changes are observed between the GTP-bound form and the transition state intermediate, mainly in the switch II region containing the catalytic Gln(79) residue and independent of A30P mutation-induced local alterations in the P-loop. The structures suggest an important catalytic role for a P-loop backbone amide group, which is eliminated in the A30P mutant, and support the notion that the transition state of GTPase-mediated GTP hydrolysis is of considerable dissociative character.

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  14. Structure of the extremely slow GTPase Rab6A in the GTP bound form at 1.8A resolution. Bergbrede T, Pylypenko O, Rak A, Alexandrov K. J. Struct. Biol. 152 235-238 (2005)
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  18. Molecular mechanism of Ras-related protein Rab-5A and effect of mutations in the catalytically active phosphate-binding loop. Khan FI, Aamir M, Wei DQ, Ahmad F, Hassan MI. J. Biomol. Struct. Dyn. 35 105-118 (2017)
  19. Functional study on GTP hydrolysis by the GTP-binding protein from Sulfolobus solfataricus, a member of the HflX family. Huang B, Wu H, Hao N, Blombach F, van der Oost J, Li X, Zhang XC, Rao Z. J. Biochem. 148 103-113 (2010)
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  21. Identification of PLP2 and RAB5C as novel TPD52 binding partners through yeast two-hybrid screening. Shahheydari H, Frost S, Smith BJ, Groblewski GE, Chen Y, Byrne JA. Mol. Biol. Rep. 41 4565-4572 (2014)
  22. Purification and characterization of Ras related protein, Rab5a from Tinospora cordifolia. Amir M, Wahiduzzaman, Dar MA, Haque MA, Islam A, Ahmad F, Hassan MI. Int. J. Biol. Macromol. 82 471-479 (2016)
  23. Ehrlichia type IV secretion system effector Etf-2 binds to active RAB5 and delays endosome maturation. Yan Q, Lin M, Huang W, Teymournejad O, Johnson JM, Hays FA, Liang Z, Li G, Rikihisa Y. Proc. Natl. Acad. Sci. U.S.A. 115 E8977-E8986 (2018)
  24. Identification of the Binding Sites on Rab5 and p110beta Phosphatidylinositol 3-kinase. Whitecross DE, Anderson DH. Sci Rep 7 16194 (2017)
  25. Structural plasticity mediates distinct GAP-dependent GTP hydrolysis mechanisms in Rab33 and Rab5. Majumdar S, Acharya A, Prakash B. FEBS J. 284 4358-4375 (2017)
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  27. Poly(ADP-ribose) Polymerase 1 Mediates Rab5 Inactivation after DNA Damage. Mashimo M, Morozumi A, Nobeyama A, Kanzaki M, Negi S, Kato J, Moss J, Nomura A, Fujii T. Int J Mol Sci 23 7827 (2022)
  28. Structural and Biophysical Characterization of Rab5a from Leishmania Donovani. Maheshwari D, Yadav R, Rastogi R, Jain A, Tripathi S, Mukhopadhyay A, Arora A. Biophys. J. 115 1217-1230 (2018)


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