OCTYL GLUCOSIDE EXTRACTS GTP-BINDING REGULATORY PROTEINS FROM RAT-BRAIN SYNAPTONEUROSOMES AS LARGE, POLYDISPERSE STRUCTURES DEVOID OF BETA-GAMMA-COMPLEXES AND SENSITIVE TO DISAGGREGATION BY GUANINE-NUCLEOTIDES

GTP-binding regulatory proteins are generally purified from cholate-extracted membranes in the form of heterotrimers (G proteins) consisting of a GTP-binding subunit (α protein) complexed with a tightly interacted heterodimer termed βγ. In this study we extracted the proteins from rat brain 'synaptoneurosomes' using the neutral detergent 1-octyl β-D-glucopyranoside (octyl glucoside). Using specific antibodies for detection by immunoblotting and sucrose gradients for analyzing hydrodynamic properties, we found that each species of α protein (α subunits of stimulatory, inhibitory, and brain GTP-binding proteins) exhibited a broad range (4 S to >12 S) of polydisperse structures with peak values (5 S to 7 S) considerably greater than that of heterotrimeric G proteins. The β subunit proteins, for example, appeared as a homogeneous peak at 4.4 S within which only a fraction of the total α proteins can be associated. Incubation of octyl glucose extracts at 30°C rapidly sedimented the α proteins but not the β proteins. Incubation at 30°C with guanosine 5'[γ-thio]triphosphate (10-100 μM) prevented rapid sedimentation. Hydrodynamic analysis revealed that all α proteins were converted to ~4 S structures by the actions of guanosine 5'-[γ-thio]triphosphate without change in the hydrodynamic properties of the β proteins. Extraction of the membranes with sodium cholate instead of octyl glucoside resulted in complete loss of the large, polydisperse structures of the α proteins; the S values were ~4 S, in the range for β proteins. These findings suggest that the transducing GTP-binding proteins in synaptoneurosomes exist as polydisperse, possibly multimer, structures of various size that are stable in octyl glucoside but destroyed by cholate. The polydisperse structures are not associated with βγ complexes and are sensitive to the disaggregating effects of guanosine 5'-[γ-thio]triphosphate.