A detailed structural analysis of protein conformational states in solution is proposed. Two proteins have been selected for the study: HPr, a phosphoryl transfer protein of the phosphoenolpyruvate-dependent sugar transport system in E. coli, and calmodulin, a Ca2+ dependent regulatory protein. Both proteins undergo a conformational change as part of their mechanism of action, but in each case one conformation cannot be studied by X-ray crystallography. Two-dimensional 1H NMR (2DNMR) spectroscopy at 500MHz will be used to study the solution structure of the two proteins. Experiments have been proposed in light of the fact that both HPr and calmodulin are larger than any of the proteins which have been studied using 2DNMR to date. Tryptic fragments of calmodulin, each of which encompasses half of the protein molecule, will be studied as models for the whole protein. A number of non-conventional 2DNMR experiments such as relayed coherence transfer and multiple quantum spectroscopy will be performed on the proteins to alleviate the spectral overlap in the 2D spectra to the large number of protons present in larger proteins.
| Klevit, R E; Drobny, G P; Waygood, E B (1986) Two-dimensional 1H NMR studies of histidine-containing protein from Escherichia coli. 1. Sequential resonance assignments. Biochemistry 25:7760-9 |
| Klevit, R E; Drobny, G P (1986) Two-dimensional 1H NMR studies of histidine-containing protein from Escherichia coli. 2. Leucine resonance assignments by long-range coherence transfer. Biochemistry 25:7770-3 |
| Klevit, R E; Waygood, E B (1986) Two-dimensional 1H NMR studies of histidine-containing protein from Escherichia coli. 3. Secondary and tertiary structure as determined by NMR. Biochemistry 25:7774-81 |
| Klevit, R E; Blumenthal, D K; Wemmer, D E et al. (1985) Interaction of calmodulin and a calmodulin-binding peptide from myosin light chain kinase: major spectral changes in both occur as the result of complex formation. Biochemistry 24:8152-7 |
