Proteins of Functioning Flagellar Rotor Turnover but only in the Presence of Signalling Proteins

Abstract

For many bacteria, motility is achieved by means of one or more filaments, each being driven by a rotary motor embedded in the cell membrane. The bacterial flagellar rotary motor is one of nature's most intricate molecular machines and is composed of two main parts: the rotor and the stator. Whilst much is known about its static structure, there are little data on the dynamics and interactions of its constituents under natural conditions in living cells. Recent results revealed the rapid exchange undergone by the membrane spanning protein complex MotAB which localizes around the rotor and forms the torque generating units. The C-ring, also called “switch complex”, is part of the rotor and is localized to the cytoplasmic region of the motor. The response regulator CheY-P binds one of the C-ring components, FliM, causing the rotor to switch rotational direction thus making FliM the interface with the chemosensory pathway.Here, we use single-molecule fluorescence imaging in Escherichia coli cells expressing genomically-encoded YPet derivatives of FliM at physiological levels. Analysis of functional flagellar motors revealed that each contains ∼34 FliM molecules. We found that two FliM populations coexist within the same motor, one undergoing constant turnover and one remaining “fixed”. Surprisingly, exchange within the dynamic population relies on the presence of the response regulator protein linking the complex to the rest of the sensory pathway and may therefore play an active part of signal processing within the cell.These results show the strength of combining molecular genetics with in vivo imaging and in this case illustrate the highly dynamic and adaptive nature of the bacterial flagellar motor. Further data will be presented on the different FliM complexes observed in the cells and their possible interaction with CheY-P.