Chemotaxis is a biological process allowing bacteria to swim toward nutrients and away from repellents. Chemotaxis occurs through arrays made of core signaling units (CSU), consisting of the chemoreceptor, the kinase CheA, and the coupling protein CheW. Our goal is to determine whether CheW plays only a structural role in the arrays, or whether it is also involved in signaling from the receptor to CheA. To answer this, we utilize hydrogen deuterium exchange mass spectrometry (HDX) to monitor the global structural changes between CheW in solution and in both on and off signaling states. Experiments on CheW alone demonstrate the beta barrel core of CheW exchanges slowly, while the rest fully exchanges with deuterium in 3 minutes. Experiments on CheW in complexes show that all three protein binding surfaces in CheW have slower uptake when incorporated into complexes. Preliminary HDX on both signaling states show CheW has less uptake in the kinase-on state compared to the kinase-off state, suggesting CheW has a signaling role in complexes. Moving forward, additional replicate HDX experiments on CheW in both signaling states will establish the threshold for significant difference uptake, revealing what regions of CheW change with signaling state as a foundation for models for the role of CheW in propagating the signal. Such models will be further tested by DEPC covalent labeling to clarify whether changes between signaling states observed in HDX occur due to changes in solvent exposure or protein dynamics. Finally, HDX experiments that distinguish between CheW in CSUs vs CheW rings will provide insight into the multiple roles of CheW in the structure, signaling, and cooperativity of these remarkable signaling arrays. This research is supported by NIH R01 GM120195 and T32 GM139789.
Jankowski et al. (Sun,) studied this question.