In situ investigation of Heterotrimeric G protein βγ subunit binding and orientation on membrane bilayers

This paper investigates the binding and orientation of an important signal transduction membrane protein, G beta Y-1(2), in a membrane bilayer. This is the first time that sum frequency generation (SFG) vibrational spectroscopy has been used to deduce the orientation of a peripheral membrane protein in the membrane environment. Wild-type. and soluble G beta Y-1(2) subunits were studied and the results are compared to evaluate the anchoring role of the geranylgeranyl group. SFG studies show that without the geranylgeranyl group, G beta Y-1(2) adsorbs onto the surface with the beta-propeller facing the membrane surface. At this orientation the helical domains orient more or less parallel to the surface. In contrast, wild-type G beta Y-1(2) is anchored to the membrane via the geranylgeranyl group with the P propeller more or less perpendicular to the surface. Under this circumstance, the helical domains are no longer parallel to the surface and hence contribute the dominant spectral features. From the measured SFG ppp and ssp intensity ratio, the orientation of the G beta(1) Y-2 is found to be about -35 degrees around the y -axis. SFG results also indicate that lipid compositions can modulate either the overall G beta Y-1(2) molecular orientation or the tertiary structure of G beta Y-1(2).