Abstract

The combination of magic-angle spinning and dipolar decoupling for resolution enchancement, with cross- and double-cross polarization for sensitivity and selectivity enhancement, has made practical the characterization of the structure and dynamics of single sites of proteins in biological solids. We propose to develop three additional new solids NMR experiments to help characterize protein binding sites: (1) rotational-echo double resonance, (2) DANTE-differenced 13C-13C spin diffusion, and (3) combined DANTE- and double-cross polarization spin transfer. These experiments will measure quantitatively dipolar coupling between 13C, 15N, 31P and 17O in both covalent and ionic bonds. The latter will extend solids NMR techniques to protein binding sites in stable charge-neutralized complexes. We propose to apply solids NMR experiments to the identification and characterization of stable-isotope single-, double-, and triple- labeled binding sites of immobilized proteins, protein adducts, and the peptide stems of peptidoglycans in intact bacterial cell walls.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM040634-03
Application #
3298387
Study Section
Biophysical Chemistry Study Section (BBCB)
Project Start
1988-07-01
Project End
1991-06-30
Budget Start
1990-07-01
Budget End
1991-06-30
Support Year
3
Fiscal Year
1990
Total Cost
Indirect Cost

  Institution

Name
Washington University
Department
Type
Schools of Arts and Sciences
DUNS #
062761671
City
Saint Louis
State
MO
Country
United States
Zip Code
63130

  Publications

Mehta, Anil K; Schaefer, Jacob (2003) Rotational-echo double resonance of uniformly labeled 13C clusters. J Magn Reson 163:188-91
Studelska, Daniel R; McDowell, Lynda M; Adler, Marc et al. (2003) Conformation of a bound inhibitor of blood coagulant factor Xa. Biochemistry 42:7942-9
Mehta, Anil K; Studelska, Daniel R; Fischer, Markus et al. (2002) Investigation of the binding of epimer A of the covalent hydrate of 6,7-bis(trifluoromethyl)-8-D-ribityllumazine to a recombinant F22W Bacillus subtilis lumazine synthase mutant by (15)N[(19)F] REDOR NMR. J Org Chem 67:2087-92
Li, Y; Poliks, B; Cegelski, L et al. (2000) Conformation of microtubule-bound paclitaxel determined by fluorescence spectroscopy and REDOR NMR. Biochemistry 39:281-91
McDowell, L M; McCarrick, M A; Studelska, D R et al. (1999) Conformations of trypsin-bound amidine inhibitors of blood coagulant factor Xa by double REDOR NMR and MD simulations. J Med Chem 42:3910-8
Schaefer, J (1999) REDOR-determined distances from heterospins to clusters of 13C labels. J Magn Reson 137:272-5
Studelska, D R; McDowell, L M; Espe, M P et al. (1997) Slowed enzymatic turnover allows characterization of intermediates by solid-state NMR. Biochemistry 36:15555-60
McDowell, L M; Lee, M; McKay, R A et al. (1996) Intersubunit communication in tryptophan synthase by carbon-13 and fluorine-19 REDOR NMR. Biochemistry 35:3328-34
McDowell, L M; Schaefer, J (1996) High-resolution NMR of biological solids. Curr Opin Struct Biol 6:624-9
McDowell, L M; Klug, C A; Beusen, D D et al. (1996) Ligand geometry of the ternary complex of 5-enolpyruvylshikimate-3-phosphate synthase from rotational-echo double-resonance NMR. Biochemistry 35:5395-403

Showing the most recent 10 out of 18 publications