Solid-State REDOR NMR Distance Measurements at the Ligand Site of a Bacterial Chemotaxis Membrane Receptor

Abstract

The Escherichia coli serine receptor senses serine levels in the environment and transmits this information across the bacterial inner membrane to modulate a protein phosphorylation cascade which controls swimming behavior. Solid-state nuclear magnetic resonance (NMR) has been used to characterize specific structural features of the ligand binding site interactions in the intact, membrane-bound Ser receptor. Rotational-echo double-resonance (REDOR) experiments on [¹⁵N]Ser bound to a [1-¹³C]Phe-receptor preparation are used to measure distances between the ligand amino group and the carbonyls of two phenylalanine residues in the ligand binding pocket. The results indicate two 4.0 ± 0.2 Å distances, in excellent agreement with the X-ray crystal structure of a soluble fragment of the homologous aspartate receptor [Milburn et al. (1991) Science 254, 1342−1347]. These results confirm the similarity of the binding sites of the Asp and Ser receptors, and demonstrate the feasibility of using solid-state NMR measurements to obtain specific structural information on the 120 kDa intact receptor for probing transmembrane signaling mechanisms.