Structure of the GDP-Pi complex of Gly203→Ala G(iα1): a mimic of the ternary product complex of Galpha-catalyzed GTP hydrolysis

Authors

Albert M. Berghuis, University of Texas at Dallas
Ethan Lee, University of Texas Southwestern Medical Center
André S. Raw, University of Texas Southwestern Medical Center
Alfred G. Gilman, University of Texas Southwestern Medical Center
Stephen R. Sprang, University of Montana

Document Type

Article

Publication Title

Structure

Publication Date

11-15-1996

Volume

4

Issue

11

Disciplines

Biology | Life Sciences

Abstract

Background: G proteins play a vital role in transmembrane signalling events. In their inactive form G proteins exist as heterotrimers consisting of an α subunit, complexed with GDP and a dimer of βγ subunits. Upon stimulation by receptors, G protein α subunits exchange GDP for GTP and dissociate from βγ. Thus activated, α subunits stimulate or inhibit downstream effectors. The duration of the activated state corresponds to the single turnover rate of GTP hydrolysis, which is typically in the range of seconds. In G(iα1), the Gly203→Ala mutation reduces the affinity of the substrate for Mg²⁺, inhibits a key conformational step that occurs upon GTP binding and consequently inhibits the release of βγ subunits from the GTP complex. The structure of the Gly203→Ala mutant of G(iα1) (G203AG(iα1)) bound to the slowly hydrolyzing analog of GTP (GTPγS) has been determined in order to elucidate the structural changes that take place during hydrolysis. Results: We have determined the three dimensional structure of a Gly203→Ala mutant of G(iα1) at 2.6 Å resolution. Although crystals were grown in the presence of GTPγS and Mg²⁺, the catalytic site contains a molecule of GDP and a phosphate ion, but no Mg²⁺. The phosphate ion is bound to a site near that occupied by the γ-phosphate of GTPγS in the activated wild-type α subunit. A region of the protein, termed the Switch II helix, twists and bends to adopt a conformation that is radically different from that observed in other G(iα1) subunit complexes. Conclusions: Under the conditions of crystallization, the Gly203→Ala mutation appears to stabilize a conformation that may be similar, although perhaps not identical, to the transient ternary product complex of G(iα1)-catalyzed GTP hydrolysis. The rearrangement of the Switch II helix avoids a potential static conflict caused by the mutation. However, it appears that dissociation of the γ-phosphate from the pentacoordinate intermediate also requires a conformational change in Switch II. Thus, a conformational rearrangement of the Switch II helix may be required in Gα-catalyzed GTP hydrolysis.

DOI

https://doi.org/10.1016/S0969-2126(96)00136-0

Rights

© Current Biology Ltd

Recommended Citation

Berghuis, Albert M.; Lee, Ethan; Raw, André S.; Gilman, Alfred G.; and Sprang, Stephen R., "Structure of the GDP-Pi complex of Gly203→Ala G(iα1): a mimic of the ternary product complex of Galpha-catalyzed GTP hydrolysis" (1996). Biological Sciences Faculty Publications. 541.
https://scholarworks.umt.edu/biosci_pubs/541