Several studies have suggested that disrupting interactions of the G protein βγ subunits with downstream binding partners might be a valuable study for pharmaceutical development. Recently, small molecules have been found which bind to Gβγ with high apparent affinity in an enzyme-linked immunosorbent assay (ELISA), have demonstrated selective inhibition of interactions of Gβγ with downstream signaling partners, and have been shown to increase antinociceptive effects of morphine and inhibit inflammation in vivo. In this paper we examine several docking and scoring protocols for estimating binding affinities for a set of 830 ligands from the NCI diversity set to the Gβ1γ2 subunit and compared these with IC50s measured in a competition ELISA with a high-affinity peptidic ligand. The best-performing docking protocol used a consensus score and ensemble docking and resulted in a 6-fold enrichment of high-affinity compounds in the top-ranked 5% of the ligand data set.

Designing an optimal HIV-1 vaccine faces the challenge of identifying antigens that induce a broad immune capacity. One factor to control the breadth of T cell responses is the surface morphology of a peptide-MHC complex. Here, we present an in silico protocol for predicting peptide-MHC structure. A robust signature of a conformational transition was identified during all-atom molecular dynamics, which results in a model with high accuracy. A large test set was used in constructing our protocol and we went another step further using a blind test with a wild-type peptide and two highly immunogenic mutants, which predicted substantial conformational changes in both mutants. The center residues at position five of the analogs were configured to be accessible to solvent, forming a prominent surface, while the residue of the wild-type peptide was to point laterally towards the side of the binding cleft. We then experimentally determined the structures of the blind test set, using high resolution of X-ray crystallography, which verified predicted conformational changes. Our observation strongly supports a positive association of the surface morphology of a peptide-MHC complex to its immunogenicity. Our study offers the prospect of enhancing immunogenicity of vaccines by identifying MHC binding immunogens.

Previous NMR experiments on unbound G protein βγ heterodimer suggested that particular residues in the binding interface are mobile on the nanosecond timescale. In this work we performed nanosecond-timescale molecular dynamics simulations to investigate conformational changes and dynamics of Gβγ in the presence of several binding partners: a high-affinity peptide (SIGK), phosducin, and the GDP-bound α subunit. In these simulations, the high mobility of GβW99 was reduced by SIGK, and it appeared that a tyrosine might stabilize GβW99 by hydrophobic or aromatic stacking interactions in addition to hydrogen bonds. Simulations of the phosducin-Gβγ complex showed that the mobility of GβW99 was restricted, consistent with inferences from NMR. However, large-scale conformational changes of Gβγ due to binding, which were hypothesized in the NMR study, were not observed in the simulations, most likely due to their short (nanosecond) duration. A pocket consisting of hydrophobic amino acids on Gα appears to restrict GβW99 mobility in the crystal structure of the Gαβγ heterotrimer. The simulation trajectories are consistent with this idea. However, local conformational changes of residues GβW63, GβW211, GβW297, GβW332 and GβW339 were detected during the MD simulations. As expected, the magnitude of atomic fluctuations observed in simulations was greater for α than for the βγ subunits, suggesting that α has greater flexibility. These observations support the notion that to maintain the high mobility of GβW99 observed by solution NMR requires that the Gβ−α interface must open up on time scale longer than can be observed in nanosecond scale simulations.