Mechanisms driving open-closed transitions and dimer stability of the large GTPases hGBP1 and hGBP5.

The human guanylate-binding proteins 1 and 5 (hGBP1/5) are key players in innate immunity, vital for defending against intracellular pathogens and mediating membrane-associated immune responses. These protein functions are closely linked to their structural dynamics, making a detailed understanding of the conformational behavior imperative. The closed conformation of hGBP1 is well-characterized through crystallography, but the structural basis for its transition to an active, open state remains less understood. This study uses all-atom and coarse-grained molecular dynamics simulations to investigate the stability and motions of extended monomeric and dimeric hGBP1 and hGBP5 with and without GTP bound. Results reveal that dimers exhibit greater stability than monomers, primarily due to extensive stalk interactions that facilitate a structural crossing of the protomers at the interface of the GTPase and middle domains. This arrangement aligns the middle and effector domains parallel to one another, further stabilizing the dimeric state through the formation of coiled-coil structures supported by salt bridges and hydrophobic contacts. Notably, monomers of both hGBP1 and hGBP5 can revert to a closed state stabilized by a network of salt bridges between the effector domain and the surrounding domains. In hGBP5, this transition is further facilitated by the geranylgeranyl group, which more effectively reaches and buries itself within a hydrophobic pocket of the GTPase domain compared to hGBP1. These findings highlight key factors affecting the stability of hGBP1/5 monomers and dimers, providing insights into their activation mechanisms that are relevant for their role in innate immunity.