Uncovering the association mechanism between two intrinsically flexible proteins

Abstract

AbstractThe understanding of protein-protein interaction mechanisms is key to the atomistic description of cell signalling pathways and for the development of new drugs. In this context, the mechanism of intrinsically disordered proteins folding upon binding has attracted attention. The VirB9 C-terminal domain (VirB9Ct) and the VirB7 N-terminal motif (VirB7Nt) associate with VirB10 to form the outer membrane core complex of the Type IV Secretion System injectisome. Despite forming a stable and rigid complex, VirB7Ntbehaves as a random coil while VirB9Ctis intrinsically dynamic in the free state. Here we combined NMR, stopped-flow fluorescence and computer simulations using structure-based models to characterize the VirB9Ct-VirB7Ntcoupled folding and binding mechanism. Our data indicated that VirB9Ctbinds to VirB7Ntby way of a conformational selection mechanism. However, at higher temperatures energy barriers between different VirB9Ctconformations are more easily surpassed. Under these conditions the formation of non-native initial encounter complexes may not be neglected, providing alternative pathways towards the native complex conformation. These observations highlight the intimate relationship between folding and binding, calling attention to the fact that the two molecular partners must search for the most favored intramolecular and intermolecular interactions on a rugged and funnelled conformational energy landscape, along which multiple intermediates may lead to the final native state.