N-terminal binding domain of G alpha subunits: Involvement of amino acids 11-14 of G alpha(o) in membrane attachment

Heterotrimeric guanine nucleotide binding proteins (G-proteins) transmit signals from membrane receptors to a variety of intracellular effecters. G-proteins reversibly associate with components of the signal transduction system, yet remain membrane attached throughout the cycle of activation. The G alpha subunits remain attached to the plasma membrane through a combination of factors that are only partially defined. We now demonstrate that amino acids within the N-terminal domain of G alpha subunits are involved in membrane binding. We used in vitro translation, a technique widely utilized to characterize functional aspects of G-proteins, and interactions with donor-acceptor membranes to demonstrate that amino acids 11-14 of G alpha(o) contribute to membrane binding. The membrane binding of G alpha(o) lacking amino acids 11-14(D[11-14]) was significantly reduced at all membrane concentrations in comparison with wild-type G alpha(o). Several other N-terminal mutants of G alpha(o) were characterized as controls, and these results indicate that differences in myristoylation, palmitoylation and beta gamma interactions do not account for the reduced membrane binding of D[11-14]. Furthermore, when membrane attachment of G alpha(o) and mutants was characterized in transiently transfected S-35-labelled and [H-3]myristate-labelled COS cells, amino acids 11-14 contributed to membrane binding. These studies reveal that membrane binding of G alpha subunits occurs by a combination of factors that include lipids and amino acid sequences. These regions may provide novel sites for interaction with membrane components and allow additional modulation of signal transduction.