Partial 13C and 15N Chemical-Shift Assignments of the Disulfide-Bond-Forming Enzyme DsbB by 3D Magic-Angle Spinning NMR Spectroscopy

摘要

DsbB is a 20 kDa Escherichia coli inner-membrane protein that catalyzes disulfide-bond formation in periplasmic proteins.We teport highly resolved,multidimensional magic-angle spinning NMR spectra at 750 MHz H frequency,which enable partial C and N chemical-shift assignments of the signals.The narrow line widths observed indicate excellent microscopic order of the protein sample,suitable for full structure determination by solid- state NMR.Experiments were performed exclusively on uniformly 13C,15N-labeled DsbB.Chemical-shift-corretation experiments based on dipolar transfer yielded strong signals in the 3D spectra.many of which have been site specifically assigned to the four transmembrane helices of DsbB.Significant numbers of addition- al residues have been assigned to stretches of amino acids,al- though not yet placed in the amino acid sequence.We also report the temperature dependence of signal intensities from -50 deg C to 0 deg C,a range over which samples of DsbB are highly- stable.Structural and dynamic information derived from SSNMR studies can give insight into DsbB in a state that so far has not been successfully crystallized.