4N0D: Crystal Structure Of The K345l Variant Of The Gi Alpha1 Subunit Bound To Gtpgammas

Receptor-mediated activation of the Galpha subunit of heterotrimeric G proteins requires allosteric communication between the receptor binding site and the guanine nucleotide binding site, which are separated by >30 A. Structural changes in the allosteric network connecting these sites are predicted to be transient in the wild-type Galpha subunit, making studies of these connections challenging. In the current work, site-directed mutants that alter the energy barriers between the activation states are used as tools to better understand the transient features of allosteric signaling in the Galpha subunit. The observed differences in relative receptor affinity for intact Galphai1 subunits versus C-terminal Galphai1 peptides harboring the K345L mutation are consistent with this mutation modulating the allosteric network in the protein subunit. Measurement of nucleotide exchange rates, affinity for metarhodopsin II, and thermostability suggest that the K345L Galphai1 variant has reduced stability in both the GDP-bound and nucleotide-free states as compared with wild type but similar stability in the GTPgammaS-bound state. High resolution x-ray crystal structures reveal conformational changes accompanying the destabilization of the GDP-bound state. Of these, the conformation for Switch I was stabilized by an ionic interaction with the phosphate binding loop. Further site-directed mutagenesis suggests that this interaction between Switch I and the phosphate binding loop is important for receptor-mediated nucleotide exchange in the wild-type Galphai1 subunit.
PDB ID: 4N0DDownload
MMDB ID: 118267
PDB Deposition Date: 2013/10/1
Updated in MMDB: 2014/11
Experimental Method:
x-ray diffraction
Resolution: 1.55  Å
Source Organism:
Similar Structures:
Biological Unit for 4N0D: monomeric; determined by author and by software (PISA)
Molecular Components in 4N0D
Label Count Molecule
Protein (1 molecule)
Guanine Nucleotide-binding Protein G(i) Subunit Alpha-1(Gene symbol: Gnai1)
Molecule annotation
Chemicals (3 molecules)
* Click molecule labels to explore molecular sequence information.

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