A concerted mechanism for opening the GDP binding pocket and release of the nucleotide in hetero-trimeric G-proteins.
ABSTRACT G-protein hetero-trimers play a fundamental role in cell function. Their dynamic behavior at the atomic level remains to be understood. We have studied the Gi hetero-trimer through a combination of molecular dynamics simulations and normal mode analyses. We showed that these big proteins could undergo large-amplitude conformational changes, without any energy penalty and with an intrinsic dynamics centered on their GDP binding pocket. Among the computed collective motions, one of the modes (mode 17) was particularly able to significantly open both the base and the phosphate sides of the GDP binding pocket. This mode describing mainly a motion between the Ras-like and the helical domains of G(α) was in close agreement with some available X-ray data and with many other biochemical/biophysical observations including the kink of helix α5. The use of a new protocol, which allows extraction of the GDP ligand along the computed normal modes, supported that the exit of GDP was largely coupled to an opening motion along mode 17. We propose for the first time a "concerted mechanism" model in which the opening of the GDP pocket and the kink of the α5 helix occur concomitantly and favor GDP release from G(αβγ) complexes. This model is discussed in the context of the G-protein-coupled receptor/G-protein interaction close to the cell membrane.
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ABSTRACT: After extra-cellular stimulation of G-Protein Coupled Receptors (GPCRs), GDP/GTP exchange appears as the key, rate limiting step of the intracellular activation cycle of heterotrimeric G-proteins. Despite the availability of a large number of X-ray structures, the mechanism of GDP release out of heterotrimeric G-proteins still remains unknown at the molecular level. Starting from the available X-ray structure, extensive unconstrained/constrained molecular dynamics simulations were performed on the complete membrane-anchored Gi heterotrimer complexed to GDP, for a total simulation time overcoming 500 ns. By combining Targeted Molecular Dynamics (TMD) and free energy profiles reconstruction by umbrella sampling, our data suggest that the release of GDP was much more favored on its phosphate side. Interestingly, upon the forced extraction of GDP on this side, the whole protein encountered large, collective motions in perfect agreement with those we described previously including a domain to domain motion between the two ras-like and helical sub-domains of G(α).PLoS Computational Biology 07/2012; 8(7):e1002595. · 5.22 Impact Factor