Mechanisms of Inter- and Intramolecular Communication in GPCRs and G Proteins

Abstract

This study represents the first attempt to couple, by computational experiments, the mechanisms of intramolecular and intermolecular communication concerning a guanidine nucleotide exchange factor (GEF), the thromboxane A2 receptor (TXA2R), and the cognate G protein (Gq) in its heterotrimeric GDP-bound state. Two-way pathways mediate the communication between the receptor-G protein interface and both the agonist binding site of the receptor and the nucleotide binding site of the G protein. The increase in solvent accessibility in the neighborhoods of the highly conserved E/DRY receptor motif, in response to agonist binding, is instrumental in favoring the penetration of the C-terminus of Gqalpha in between the cytosolic ends of H3, H5, and H6. The arginine of the E/DRY motif is predicted to be an important mediator of the intramolecular and intermolecular communication involving the TXA2R. The receptor-G protein interface is predicted to involve multiple regions from the receptor and the G protein alpha-subunit. However, receptor contacts with the C-terminus of the alpha5-helix seem to be the major players in the receptor-catalyzed motion of the alpha-helical domain with respect to the Ras-like domain and in the formation of a nucleotide exit route in between the alphaF-helix and beta6/alpha5 loop of Gqalpha. The inferences from this study are of wide interest, as they are expected to apply to the whole rhodopsin family, given also the considerable G protein promiscuity.

Superactive mutants of thromboxane prostanoid receptor: functional and computational analysis of an active form alternative to constitutively active mutants

“…All the receptor domains except for the C-term were modeled (i.e., 1-321 sequence). The sequence alignment employed in this study as well as the deletions in the opsin template are the same as the ones utilized in our previous computational modeling study [28]. a-Helix restraints were applied to the 47-54, 99-106, 268-273, and 283-294 amino acid stretches of the target receptors.…”

“…The latter was achieved by comparative modeling by using the Ga i1 structure as a template (PDB code 1GP2 [33]). Different from the heterotrimeric G q employed in the previous study [28], the G q employed herein held the last ten amino acids in the same conformation as the recent X-ray structure of the transducin C-term (PDB code: 3DQB) [25]. The new model of the a-subunit was merged with the bc-subunits extracted from the Ga i1 b 1 c 2 heterotrimer (i.e., PDB code: 1GP2) [33].…”

“…The new model of the a-subunit was merged with the bc-subunits extracted from the Ga i1 b 1 c 2 heterotrimer (i.e., PDB code: 1GP2) [33]. Docking simulations and analyses followed the protocol previously described [28]. Substantially similar reliable solutions were achieved for the three receptor forms due to the very low structural deviation between them [i.e., a Ca-atom root mean square deviation (Ca-RMSD) around 0.2 Å ].…”

“…It was, indeed, found that the increase in solvent accessibility near the E/DRY motif in response to agonist binding is instrumental in favoring the penetration of the C-term of Ga q in between the cytosolic ends of H3, H5, and H6 [28]. In line with these predictions, the recent crystallographic structures of the constitutively active opsin apoprotein show that the breakage of the salt bridge interactions between R3.50 and both E3.49 and E3.60, associated with about 200 Å 2 increases in solvent accessibility in the neighborhoods of the E/DRY motif compared to dark rhodopsin, favoring the establishment of interactions between the R3.50 itself and the peptide from transducin [13,25].…”

Light on the structure of thromboxane A2 receptor heterodimers

“…The computational modeling of the TPa receptor is described in detail in a recent publication [34] and consists of comparative modeling followed by ns molecular dynamics (MD) simulations in implicit membrane [34].…”