Dual interaction of ADP ribosylation factor 1 with Sec7 domain and with lipid membranes during catalysis of guanine nucleotide exchange

Sec7 domains catalyze the replacement of GDP by GTP on the G protein ADP-ri bosylation factor 1 (myrARF1) by interacting with its switch I and II regio ns and by destabilizing, through a glutamic finger, the P-phosphate of the bound G;DP, The myristoylated N-terminal helix that allows myrARF1 to inter act with membrane lipids in a GTP-dependent manner is located some distance from the Sec7 domain-binding region. However, these two regions are connec ted. Measuring the binding to liposomes of functional or abortive complexes between myrARF1 and the Sec7 domain of ARNO demonstrates that myrARF1, in complex with the Sec7 domain, adopts a high affinity state for membrane lip ids, similar to that of the free G;TP-bound form. This tight membrane attac hment does not depend on the release of GDP induced by the Sec7 domain but is partially inhibited by the uncompetitive inhibitor brefeldin A. These re sults suggest that the conformational switch of the N-terminal helix of myr ARF1 to the membrane-bound form is an early event in the nucleotide exchang e pathway and is a prerequisite for a structural rearrangement at the myrAR F1-GDP/Sec7 domain interface that allows the glutamic finger to expel GDP f rom myrARF1.