Rapid kinetics of G protein subunit association: A rate-limiting conformational change?

Abstract

G protein subunit association and dissociation are thought to play an important role in signal transduction. We measured αβγ heterocomplex formation using resonance energy transfer. Fluorescein-labelled α(F-α) emission was quenched ∼ 10% on mixing with eosin-labelled βγ(E-βγ). Unlabelled βγ did not quench F-α fluorescence. Stopped-flow kinetics showed a t 1 2 ranging from 2.5 s to 0.25 s for 50 nM to 1200 nM E-βγ. The rate saturated at high E-βγ concentrations consistent with a two-step mechanism. We report the first rapid-mix studies of G protein subunit association kinetics which suggest that α and βγ combine by a two-step process with a maximal rate of 4.1 ± 0.4 s −1.