Download MendeleyA Transient Interaction between the Phosphate Binding Loop and Switch I Contributes to the Allosteric Network between Receptor and Nucleotide in G alpha i1.
Thaker, T.M., Sarwar, M., Preininger, A.M., Hamm, H.E., Iverson, T.M.(2014) J Biol Chem 289: 11331-11341
- PubMed: 24596087
- DOI: https://doi.org/10.1074/jbc.M113.539064
- Primary Citation of Related Structures:
4N0D, 4N0E - PubMed Abstract:
Receptor-mediated activation of the G¦Á subunit of heterotrimeric G proteins requires allosteric communication between the receptor binding site and the guanine nucleotide binding site, which are separated by >30 ?. Structural changes in the allosteric network connecting these sites are predicted to be transient in the wild-type G¦Á 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 G¦Á subunit. The observed differences in relative receptor affinity for intact G¦Ái1 subunits versus C-terminal G¦Ái1 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 G¦Ái1 variant has reduced stability in both the GDP-bound and nucleotide-free states as compared with wild type but similar stability in the GTP¦ÃS-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 G¦Ái1 subunit.
Organizational Affiliation:
Department of Biochemistry and Vanderbilt University Medical Center, Nashville, Tennessee 37232.
