Biophysical Society Conference | Tahoe 2022

Molecular Biophysics of Membranes

Wednesday Speaker Abstracts

NMR AND HYDROGEN EXCHANGE STUDIES OF BACTERIAL CHEMOTAXIS RECEPTOR COMPLEXES SUGGEST PROTEIN STABILIZATION IS KEY TO THE SIGNALING MECHANISM Lynmarie K Thompson 1 ; 1 University of Massachusetts Amherst, Chemistry, Amherst, MA, USA Bacterial chemotaxis proteins are both potential targets for novel antibiotics and a key model system for understanding transmembrane signaling mechanisms. A remarkable sensor array of membrane-bound chemoreceptors, CheW, and CheA detect molecules in the environment and control swimming direction. Signaling through this complex begins as a ligand-induced 2 Å displacement of a receptor alpha helix that extends from the periplasm through the membrane. The objective of this study is to compare the structure and dynamics of signaling states to discover how the signal is propagated an additional ~200 Å from the membrane to the cytoplasmic tip of the receptor and how this signal controls the activity of CheA. Our approach begins with assembly of homogeneous, native-like functional complexes of an E coli Asp receptor cytoplasmic fragment (CF) with the kinase CheA and coupling protein CheW. Hydrogen exchange mass spectrometry (HDX-MS) results indicate that the CF is partially disordered within functional complexes and has a small, well-ordered protein interaction region. Slower hydrogen exchange throughout the kinase-on state suggests signaling inputs modulate the disorder of the cytoplasmic domain to control the kinase activity. HDX-MS of CheA alone and in these functional complexes indicate the domain-domain interactions and changes in domain stability that occur during kinase activation and inhibition. NMR spectra of both the highly flexible and rigid regions of these proteins are mapping how and where dynamics change with signaling state, and should enable us to obtain structural information on the CF/CheA/CheW interactions. This study reveals the mechanistic role of protein disorder and stabilization in signaling and control of catalysis. This research supported by National Institutes of Health Grant R01-GM120195.

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