Abstract
The retinitis pigmentosa 2 (RP2) gene is responsible for a particular variant of X chromosome–linked eye disease. Previously, RP2 was shown to bind the GTP form of the small G protein Arf-like 3 (Arl3), thus qualifying as an effector. Here we present the Arl3–GppNHp–RP2 complex structure, which shows features resembling complexes with GTPase-activating proteins (GAPs). Biochemical analysis showing a 90,000-fold stimulation of the GTPase reaction together with the structure of an Arl3–GDP–AlF4−–RP2 transition state complex showed that RP2 is an efficient GAP for Arl3, with structural features similar to other GAPs. Furthermore, the effect of mutations in patients with retinitis pigmentosa correlated with their effect on catalysis, in particular the mutation of the arginine finger of RP2. The cognate G protein–GAP pair is conserved in yeast as Cin4–Cin2, and the ability of RP2 to act as a GAP can be correlated with its ability to complement a CIN2-deletion phenotype.
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Acknowledgements
We thank D. Kühlmann and C. Körner for technical assistance, M. Farkasovsky for support in yeast genetic experiments, I. Vetter and M. Weyand for crystallographic assistance and T. Barends, W. Blankenfeldt, Z. Guo and T. Meinhart for X-ray data collection. We are grateful to the staff of beamline X10SA at the Swiss Light Source for their support. We additionally thank W. Berger, Universität Zürich, for providing the RP2 clone, K. Siegers, Max-Planck-Institut für Biochemie, Martinsried, Germany, for providing the CIN2 deletion yeast strain and the CIN2 and CIN4 clones, and N. Cowan, New York University Medical Center, for providing the CoC clone. R.G. was supported by the Fonds der Chemischen Industrie.
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S.V. performed all the experiments and their analysis and contributed to their design; R.G. strongly supported crystallization and structure determination; E.E. established the yeast complementation assay and performed preliminary complementation experiments; A.W. supervised all the work, advised on the design of the experiments and wrote the manuscript with S.V.
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Veltel, S., Gasper, R., Eisenacher, E. et al. The retinitis pigmentosa 2 gene product is a GTPase-activating protein for Arf-like 3. Nat Struct Mol Biol 15, 373–380 (2008). https://doi.org/10.1038/nsmb.1396
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DOI: https://doi.org/10.1038/nsmb.1396
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