Dynamic domain interactions encode possible CheA autophosphorylation mechanisms revealed by coarse-grained simulations

Abstract Autophosphorylation of CheA is key to initiation of the phosphorylation cascade that eventually controls the direction of downstream flagellar motors for chemotaxis signaling in motile bacteria. The phospho-transfer reaction, from ATP bound in the P4 catalytic domain to a specific His residue in the P1 substrate domain in CheA, can be significantly accelerated within core signaling unit complexes containing chemoreceptors, CheA and CheW. Previous studies have proposed that CheA autophosphorylation activity is regulated by changing the dynamics of P4 and/or altering its interactions with P1 in response to signals transmitted from chemoreceptors. However, the positions of CheA P1 and P2 domains in the core signaling unit are not well-characterized because they form only transient interactions with other domains. Though previous studies have identified possible domain-domain interaction surfaces, especially for P1 and P4, a bottom-up analysis of these interactions has not been performed. Here, we employed extensive molecular simulations to analyze interactions among CheA domains using a hybrid resolution (HyRes) protein model designed for dynamic protein structures and interactions. The results revealed multiple major modes of dynamic P1/P4 interactions. In particular, P1 was found to bind dynamically near the preferential binding surfaces on P4 even in the ATP free state. ATP binding to P4 reduces the motion of P1 binding and promotes a trans- productive-like mode that brings His48 in close contact with the bound ATP for possible autophosphorylation. The predicted non-productive and productive P1/P4 interaction modes appear highly consistent with existing NMR, mutagenesis, and chemical modification data. Together, these findings provide a more complete picture of the dynamics of domain-domain interactions of CheA and new insights into the possible regulation mechanism of its autophosphorylation. Competing Interest Statement The authors have declared no competing interest.