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 signal 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, residues 124-257, that binds the response regulator CheY. Backbone atom resonance assignments were obtained by analysis of 3D HNCACB, 3D CBCA(CO)NH, and HNCO spectra, whereas side-chain assignments were obtained primarily by analysis of 3D H(CCO)NH, 3D C(CO)NH, 3D HCCH-TOCSY, and 3D 1H, 15N TOCSY-HSMQC spectra. NOE cross peak patterns and intensities as well as torsion angle restraints were used to determine the secondary structure, and a low-resolution structure was calculated by hybrid distance-geometry simulated annealing methods. The CheA124-257 fragment 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. The 15N-1H correlation spectrum of 15N-labeled CheA124-257 bound to unlabeled CheY shows specific localized changes that may correspond to a CheY-binding face on CheA.
McEvoy
M M
MM
Institute of Molecular Biology, University of Oregon, Eugene 97403, USA.
Zhou
H
H
Roth
A F
AF
Lowry
D F
DF
Morrison
T B
TB
Kay
L E
LE
Dahlquist
F W
FW
D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D002352:Carrier Proteins; D002633:Chemotaxis; D003196:Computer Graphics; D004926:Escherichia coli / Q000737:chemistry* / Q000201:enzymology; D029968:Escherichia coli Proteins; D000071677:Histidine Kinase; D006860:Hydrogen Bonding; D009682:Magnetic Resonance Spectroscopy; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism*; D000072236:Methyl-Accepting Chemotaxis Proteins; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017510:Protein Folding; D011494:Protein Kinases / Q000737:chemistry* / Q000378:metabolism; D017433:Protein Structure, Secondary
D001426:Bacterial Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D000072236:Methyl-Accepting Chemotaxis Proteins; C000607606:cheY protein, E coli; D011494:Protein Kinases; D000071677:Histidine Kinase; C000607608:cheA protein, E coli