NUCLEAR-MAGNETIC-RESONANCE ASSIGNMENTS AND GLOBAL FOLD OF A CHEY-BINDING DOMAIN IN CHEA, THE CHEMOTAXIS-SPECIFIC KINASE OF ESCHERICHIA-COLI

CheA is the histidine autokinase in the Escherichia coli chemotaxis si gnal transduction pathway responsible for coupling of signals received by transmembrane receptors to the response regulators CheY and CheB. Here NMR spectroscopy is used to study a 14 kDa fragment of CheA, resi dues 124-257, that binds the response regulator CheY. Backbone atom re sonance assignments were obtained by analysis of 3D HNCACB, 3D CBCA(CO )NH, and HNCO spectra, whereas side-chain assignments were obtained pr imarily by analysis of 3D H(CCO)NH, 3D C(CO)NH, 3D HCCH-TOCSY, and 3D H-1,N-15 TOCSY-HSMQC spectra. NOE cross peak patterns and intensities as well as torsion angle restraints were used to determine the seconda ry structure, and a low-resolution structure was calculated by hybrid distance-geometry simulated annealing methods. The CheA124-257 fragmen t consists of four antiparallel beta strands and two helices, arranged in an ''open-faced beta-sandwich'' motif, as well as two unstructured ends that correspond to domain linkers in the full-length protein. Th e N-15-H-1 correlation spectrum of N-15-labeled CheA124-257 bound to u nlabeled CheY shows specific localized changes that may correspond to a CheY-binding face on CheA.