Multidimensional heteronuclear NMR techniques were applied to study the phosphotransfer domain, residues 1-134, of the histidine kinase CheA, from Escherichia coli, which contains the site of autophosphorylation, His48. Assignments of the backbone amide groups and side chain protons are nearly complete. Our studies show that this protein fragment consists of five alpha-helices (A-E) connected by turns. Analysis of NOE distance restraints provided by two-dimensional (2D) 1H-1H and three-dimensional (3D) 15N-edited NOESY spectra using model building and structure calculations indicates that the five helices form an antiparallel helix bundle with near-neighbor connectivity. The amino-terminal four helices are proposed to be arranged in a right-handed manner with helix E packing against helices C and D. From ideal hydrophobic helical packing and structure calculations, the site of autophosphorylation, His48, is nearly fully exposed to the solvent. We measured the NMR relaxation properties of the backbone 15N nuclei using inverse detected two-dimensional NMR spectroscopy. The protein backbone dynamics studies show that CheA1-134 is formed into a tight and compact structure with very limited flexibilities both in helices and turns. Structural implications of titration and phosphorylation experiments are briefly discussed.