Mutations in leucine-rich-repeat kinase 2 (LRRK2) are the most frequent cause of late-onset Parkinson's disease (PD). LRRK2 belongs to the Roco family of proteins which share a conserved Ras-like G-domain (Roc) and a C-terminal of Roc (COR) domain tandem. The nucleotide state of small G-proteins is strictly controlled by guanine–nucleotide-exchange factors (GEFs) and GTPase-activating proteins (GAPs). Because of contradictory structural and biochemical data, the regulatory mechanism of the LRRK2 Roc G-domain and the RocCOR tandem is still under debate. In the present study, we solved the first nucleotide-bound Roc structure and used LRRK2 and bacterial Roco proteins to characterize the RocCOR function in more detail. Nucleotide binding induces a drastic structural change in the Roc/COR domain interface, a region strongly implicated in patients with an LRRK2 mutation. Our data confirm previous assumptions that the C-terminal subdomain of COR functions as a dimerization device. We show that the dimer formation is independent of nucleotide. The affinity for GDP/GTP is in the micromolar range, the result of which is high dissociation rates in the s−1 range. Thus Roco proteins are unlikely to need GEFs to achieve activation. Monomeric LRRK2 and Roco G-domains have a similar low GTPase activity to small G-proteins. We show that GTPase activity in bacterial Roco is stimulated by the nucleotide-dependent dimerization of the G-domain within the complex. We thus propose that the Roco proteins do not require GAPs to stimulate GTP hydrolysis but stimulate each other by one monomer completing the catalytic machinery of the other.
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January 2015
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Research Article|
December 12 2014
Revisiting the Roco G-protein cycle
Susanne Terheyden;
Susanne Terheyden
1
*Department of Cell Biochemistry, University of Groningen, Nijenborgh 7, 9747 AG Groningen, The Netherlands
†Max-Planck-Institut für Molekulare Physiologie, Otto-Hahnstrasse 11, 44227 Dortmund, Germany
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Franz Y. Ho;
Franz Y. Ho
1
‡Department of Biochemistry, University of Groningen, Nijenborgh 4, 9747 AG Groningen, The Netherlands
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Bernd K. Gilsbach;
Bernd K. Gilsbach
1
*Department of Cell Biochemistry, University of Groningen, Nijenborgh 7, 9747 AG Groningen, The Netherlands
†Max-Planck-Institut für Molekulare Physiologie, Otto-Hahnstrasse 11, 44227 Dortmund, Germany
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Alfred Wittinghofer;
Alfred Wittinghofer
†Max-Planck-Institut für Molekulare Physiologie, Otto-Hahnstrasse 11, 44227 Dortmund, Germany
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Arjan Kortholt
Arjan Kortholt
2
*Department of Cell Biochemistry, University of Groningen, Nijenborgh 7, 9747 AG Groningen, The Netherlands
2To whom correspondence should be addressed (email A.Kortholt@rug.nl).
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Publisher: Portland Press Ltd
Received:
August 26 2014
Revision Received:
October 09 2014
Accepted:
October 15 2014
Accepted Manuscript online:
October 15 2014
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2015 Biochemical Society
2015
Biochem J (2015) 465 (1): 139–147.
Article history
Received:
August 26 2014
Revision Received:
October 09 2014
Accepted:
October 15 2014
Accepted Manuscript online:
October 15 2014
Citation
Susanne Terheyden, Franz Y. Ho, Bernd K. Gilsbach, Alfred Wittinghofer, Arjan Kortholt; Revisiting the Roco G-protein cycle. Biochem J 1 January 2015; 465 (1): 139–147. doi: https://doi.org/10.1042/BJ20141095
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