AbstractIn the chemotaxis of Escherichia coli, the cell’s behavioral switch involves binding of the phosphorylated form of the chemotaxis signaling protein CheY (CheYp) to the flagellar motor protein FliM, which induces the motor to rotate clockwise; otherwise, the motor rotates counterclockwise. To investigate high-pressure effects on CheYp–FliM binding at atomic resolution, we conduct molecular dynamics simulations of monomeric CheYp, the N-terminal fragment of the FliM (FliMN) that binds to CheYp, and the complex that forms between those proteins at pressures ranging from 0.1 to 100 MPa. The results show that the active form of monomeric CheYp is maintained even at 100 MPa but high pressure increases the water density in the first hydration shell and can cause conformational change of the C-terminal helix. The dissociation process of the complex is investigated by parallel cascade selection molecular dynamics (PaCS-MD), revealing that high pressure considerably induces water penetration into the complex interface. Pressure dependence of standard binding free energy calculated by the Markov state model indicates that the increase of pressure from 0.1 to 100 MPa weakens the binding by ∼ 10 kcal/mol. Using high-pressure microscopy, we observed that high hydrostatic pressure reversibly fixes the motor rotation in the counter-clockwise orientation, which supports the notion that high pressure inhibits the binding of CheYp to FliM. We conclude that high pressure induces water penetration into the complex interface, which interferes with CheYp–FliM binding and prevents motor reversal.
High pressure inhibits signaling protein binding to the flagellar motor and bacterial chemotaxis through enhanced hydration