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The retinitis pigmentosa 2 gene product is a GTPase-activating protein for Arf-like 3

Nature Structural & Molecular Biology volume 15, pages 373–380 (2008)Cite this article

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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Figure 1: Structural and biochemical analysis of the Arl3–RP2 interaction.
Figure 2: Involvement of the N terminus of RP2 in Arl3 binding.
Figure 3: Structural view of RP2 as a GAP for Arl3.
Figure 4: Biochemical analysis of RP2 GAP activity toward Arl3.
Figure 5: Influence of RP2 mutations on GTP hydrolysis.
Figure 6: Conservation of the RP2–Arl3 function by yeast Cin2–Cin4.

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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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  1. Abteilung Strukturelle Biologie, Max-Planck-Institut für molekulare Physiologie, Otto-Hahn-Str. 11, Dortmund, 44227, Germany

    Stefan Veltel, Raphael Gasper, Elke Eisenacher & Alfred Wittinghofer

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  1. Stefan Veltel
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  2. Raphael Gasper
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Contributions

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.

Note: Supplementary information is available on the Nature Structural & Molecular Biology website.

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Correspondence to Alfred Wittinghofer.

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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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