The RalB-RLIP76 Complex Reveals a Novel Mode of Ral-effector Interaction

Overview

Journal Structure

Publisher Cell Press

Specialties Biochemistry
Biotechnology
Cell Biology
Molecular Biology

Date 2010 Aug 11

PMID 20696399

Citations 21

Authors

R Brynmor Fenwick

Louise J Campbell

Karthik Rajasekar

Sunil Prasannan

Daniel Nietlispach

Jacques Camonis

Darerca Owen

Helen R Mott

Affiliations

Soon will be listed here.

Abstract

RLIP76 (RalBP1) is a multidomain protein that interacts with multiple small G protein families: Ral via a specific binding domain, and Rho and R-Ras via a GTPase activating domain. RLIP76 interacts with endocytosis proteins and has also been shown to behave as a membrane ATPase that transports chemotherapeutic agents from the cell. We have determined the structure of the Ral-binding domain of RLIP76 and show that it comprises a coiled-coil motif. The structure of the RLIP76-RalB complex reveals a novel mode of binding compared to the structures of RalA complexed with the exocyst components Sec5 and Exo84. RLIP76 interacts with both nucleotide-sensitive regions of RalB, and key residues in the interface have been identified using affinity measurements of RalB mutants. Sec5, Exo84, and RLIP76 bind Ral proteins competitively and with similar affinities in vitro.

Citing Articles

Regulation of cargo exocytosis by a Reps1-Ralbp1-RalA module.

Wang S, Chen X, Crisman L, Dou X, Winborn C, Wan C Sci Adv. 2023; 9(8):eade2540.

PMID: 36812304 PMC: 9946360. DOI: 10.1126/sciadv.ade2540.

The RAL Enigma: Distinct Roles of RALA and RALB in Cancer.

Richardson D, Spehar J, Han D, Chakravarthy P, Sizemore S Cells. 2022; 11(10).

PMID: 35626682 PMC: 9139244. DOI: 10.3390/cells11101645.

RLIP76: A Structural and Functional Triumvirate.

Cornish J, Owen D, Mott H Cancers (Basel). 2021; 13(9).

PMID: 34064388 PMC: 8124665. DOI: 10.3390/cancers13092206.

Affinity maturation of the RLIP76 Ral binding domain to inform the design of stapled peptides targeting the Ral GTPases.

Hurd C, Brear P, Revell J, Ross S, Mott H, Owen D J Biol Chem. 2020; 296:100101.

PMID: 33214225 PMC: 7949049. DOI: 10.1074/jbc.RA120.015735.

Conformational resolution of nucleotide cycling and effector interactions for multiple small GTPases determined in parallel.

Killoran R, Smith M J Biol Chem. 2019; 294(25):9937-9948.

PMID: 31088913 PMC: 6597838. DOI: 10.1074/jbc.RA119.008653.

References

Nicely N, Kosak J, de Serrano V, Mattos C . Crystal structures of Ral-GppNHp and Ral-GDP reveal two binding sites that are also present in Ras and Rap. Structure. 2004; 12(11):2025-36. DOI: 10.1016/j.str.2004.08.011. View

Eathiraj S, Pan X, Ritacco C, Lambright D . Structural basis of family-wide Rab GTPase recognition by rabenosyn-5. Nature. 2005; 436(7049):415-9. PMC: 1360218. DOI: 10.1038/nature03798. View

Brunger A, Adams P, Clore G, DeLano W, Gros P, Grosse-Kunstleve R . Crystallography & NMR system: A new software suite for macromolecular structure determination. Acta Crystallogr D Biol Crystallogr. 1998; 54(Pt 5):905-21. DOI: 10.1107/s0907444998003254. View

Dorseuil O, Romero F, Letourneur F, Saragosti S, Berger R, Tavitian A . Bridging Ral GTPase to Rho pathways. RLIP76, a Ral effector with CDC42/Rac GTPase-activating protein activity. J Biol Chem. 1995; 270(38):22473-7. DOI: 10.1074/jbc.270.38.22473. View

Yamaguchi A, Urano T, Goi T, Feig L . An Eps homology (EH) domain protein that binds to the Ral-GTPase target, RalBP1. J Biol Chem. 1998; 272(50):31230-4. DOI: 10.1074/jbc.272.50.31230. View

Thompson G, Owen D, Chalk P, Lowe P . Delineation of the Cdc42/Rac-binding domain of p21-activated kinase. Biochemistry. 1998; 37(21):7885-91. DOI: 10.1021/bi980140+. View

Frankel P, Aronheim A, Kavanagh E, Balda M, Matter K, Bunney T . RalA interacts with ZONAB in a cell density-dependent manner and regulates its transcriptional activity. EMBO J. 2004; 24(1):54-62. PMC: 544910. DOI: 10.1038/sj.emboj.7600497. View

Bauer B, Mirey G, Vetter I, Garcia-Ranea J, Valencia A, Wittinghofer A . Effector recognition by the small GTP-binding proteins Ras and Ral. J Biol Chem. 1999; 274(25):17763-70. DOI: 10.1074/jbc.274.25.17763. View

Gildea J, Harding M, Seraj M, Gulding K, Theodorescu D . The role of Ral A in epidermal growth factor receptor-regulated cell motility. Cancer Res. 2002; 62(4):982-5. View

10.

Fenwick R, Prasannan S, Campbell L, Nietlispach D, Evetts K, Camonis J . Solution structure and dynamics of the small GTPase RalB in its active conformation: significance for effector protein binding. Biochemistry. 2009; 48(10):2192-206. DOI: 10.1021/bi802129d. View

11.

Vranken W, Boucher W, Stevens T, Fogh R, Pajon A, Llinas M . The CCPN data model for NMR spectroscopy: development of a software pipeline. Proteins. 2005; 59(4):687-96. DOI: 10.1002/prot.20449. View

12.

Fenwick R, Prasannan S, Campbell L, Evetts K, Nietlispach D, Owen D . Resonance assignments for the RLIP76 Ral binding domain in its free form and in complex with the small G protein RalB. Biomol NMR Assign. 2009; 2(2):191-4. DOI: 10.1007/s12104-008-9118-4. View

13.

Jiang H, Luo J, Urano T, Frankel P, Lu Z, Foster D . Involvement of Ral GTPase in v-Src-induced phospholipase D activation. Nature. 1995; 378(6555):409-12. DOI: 10.1038/378409a0. View

14.

Fenwick R, Prasannan S, Campbell L, Evetts K, Nietlispach D, Owen D . 1H, 13C and 15N resonance assignments for the active conformation of the small G protein RalB in complex with its effector RLIP76. Biomol NMR Assign. 2009; 2(2):179-82. DOI: 10.1007/s12104-008-9115-7. View

15.

Linge J, ODonoghue S, Nilges M . Automated assignment of ambiguous nuclear overhauser effects with ARIA. Methods Enzymol. 2001; 339:71-90. DOI: 10.1016/s0076-6879(01)39310-2. View

16.

Maesaki R, Ihara K, Shimizu T, Kuroda S, Kaibuchi K, Hakoshima T . The structural basis of Rho effector recognition revealed by the crystal structure of human RhoA complexed with the effector domain of PKN/PRK1. Mol Cell. 2000; 4(5):793-803. DOI: 10.1016/s1097-2765(00)80389-5. View

17.

Jin R, Junutula J, Matern H, Ervin K, Scheller R, Brunger A . Exo84 and Sec5 are competitive regulatory Sec6/8 effectors to the RalA GTPase. EMBO J. 2005; 24(12):2064-74. PMC: 1150893. DOI: 10.1038/sj.emboj.7600699. View

18.

Cornilescu G, Delaglio F, Bax A . Protein backbone angle restraints from searching a database for chemical shift and sequence homology. J Biomol NMR. 1999; 13(3):289-302. DOI: 10.1023/a:1008392405740. View

19.

Ye M, Shima F, Muraoka S, Liao J, Okamoto H, Yamamoto M . Crystal structure of M-Ras reveals a GTP-bound "off" state conformation of Ras family small GTPases. J Biol Chem. 2005; 280(35):31267-75. DOI: 10.1074/jbc.M505503200. View

20.

Park S, Weinberg R . A putative effector of Ral has homology to Rho/Rac GTPase activating proteins. Oncogene. 1995; 11(11):2349-55. View