期刊
EMBO JOURNAL
卷 27, 期 5, 页码 727-735出版社
NATURE PUBLISHING GROUP
DOI: 10.1038/emboj.2008.10
关键词
fRET; microdomain; nanocluster; plasma membrane; Ras
资金
- Howard Hughes Medical Institute Funding Source: Medline
- NIGMS NIH HHS [R01 GM066717, R01 GM066717-05A1] Funding Source: Medline
The plasma membrane nanoscale distribution of H-ras is regulated by guanine nucleotide binding. To explore the structural basis of H-ras membrane organization, we combined molecular dynamic simulations and medium-throughput FRET measurements on live cells. We extracted a set of FRET values, termed a FRET vector, to describe the lateral segregation and orientation of H-ras with respect to a large set of nanodomain markers. We show that mutation of basic residues in helix alpha 4 or the hypervariable region (HVR) selectively alter the FRET vectors of GTP- or GDP-loaded H-ras, demonstrating a critical role for these residues in stabilizing GTP- or GDP-H-ras interactions with the plasma membrane. By a similar analysis, we find that the beta 2-beta 3 loop and helix alpha 5 are involved in a novel conformational switch that operates through helix alpha 4 and the HVR to reorient the H-ras G-domain with respect to the plasma membrane. Perturbation of these switch elements enhances MAPK activation by stabilizing GTP-H-ras in a more productive signalling conformation. The results illustrate how the plasma membrane spatially constrains signalling conformations by acting as a semi-neutral interaction partner.
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