THE JOURNAL OF BIOLOGICAL CHEMISTRY

CheY, the 14-kDa response regulator protein of the Escherichia coli chemotaxis pathway, is activated by phosphorylation of Asp"'. In order to probe the struc-tural changes associated with activation, an approach which combines 'F NMR, protein engineering, and the known crystal structure of one conformer has been utilized. This first of two papers examines the effects of Mg(I1) binding and phosphorylation on the confor-mation of CheY. The molecule was selectively labeled at its six phenylalanine positions by incorporation of 4-fluorophenylalanine, which yielded no significant effect on activity. One of these "F probe positions monitored the vicinity of Lys'09, which forms a salt bridge to Asp6 ' in the apoprotein and has been proposed to act as a structural "switch " in activation. "F NM