We have extended our technology for studying macromolecular structure in solution under weakly aligning conditions. New developments increase both the size of systems that can be studied, and the accuracy that can be obtained. Rhodopsin-containing membranes contain sufficient residual diamagnetic susceptibility anisotropy that they align spontaneously in a sufficiently strong magnetic field. Undeca-peptide analogs of the C-terminus of the gamma-subunit of transducin are known to bind weakly but specifically to photo-activated rhodopsin. The orientation of the peptide occurring upon binding to rhodopsin, following photo-activation, was used by us to provide precise structural and orientational information on the peptide in the bound state, and thereby on the orientation of the entire G protein in the receptor-bound state. A prerequisite for such studies is a relatively low affinity of the peptide for the membrane-anchored receptor. For water-soluble proteins and nucleic acids, weak alignment allows very accurate measurement of internuclear dipolar couplings, thereby providing extremely precise information on local as well as global structure. For a small, 56-residue domain, we used this technology to study planarity of peptide groups and found that the out-of-plane position of the amide hydrogen has a root-mean-square value of less than about """"""""4"""""""", much smaller than predicted on the basis of literature theoretical calculations, but in good agreement with neutron diffraction results. The in-plane deviation from the line bisecting the C?-N-Ca angle is less than """"""""2"""""""". For CO-ligated adult human hemoglobin, analogous technology revealed that in solution, at 100 mM ionic strength, the quaternary structure is about half-way intermediate between the so-called R and R2 states. Application to nucleic acids shows that accurate measurement of the bending of the helical axis is possible with the weak alignment technology. We also have succeeded in measurement of interactions over much larger distances, up to 12 ?, than previously possible by NMR.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Intramural Research (Z01)
Project #
1Z01DK029020-19
Application #
6810190
Study Section
(LCP)
Project Start
Project End
Budget Start
Budget End
Support Year
19
Fiscal Year
2003
Total Cost
Indirect Cost
Name
U.S. National Inst Diabetes/Digst/Kidney
Department
Type
DUNS #
City
State
Country
United States
Zip Code
Lee, Jung Ho; Ying, Jinfa; Bax, Ad (2016) Quantitative evaluation of positive ? angle propensity in flexible regions of proteins from three-bond J couplings. Phys Chem Chem Phys 18:5759-70
Vogeli, Beat; Yao, Lishan; Bax, Ad (2008) Protein backbone motions viewed by intraresidue and sequential HN-Halpha residual dipolar couplings. J Biomol NMR 41:17-28
Chill, Jordan H; Louis, John M; Delaglio, Frank et al. (2007) Local and global structure of the monomeric subunit of the potassium channel KcsA probed by NMR. Biochim Biophys Acta 1768:3260-70
Ying, Jinfa; Chill, Jordan H; Louis, John M et al. (2007) Mixed-time parallel evolution in multiple quantum NMR experiments: sensitivity and resolution enhancement in heteronuclear NMR. J Biomol NMR 37:195-204
Grishaev, Alexander; Ying, Jinfa; Bax, Ad (2006) Pseudo-CSA restraints for NMR refinement of nucleic acid structure. J Am Chem Soc 128:10010-1
Ying, Jinfa; Grishaev, Alexander; Bryce, David L et al. (2006) Chemical shift tensors of protonated base carbons in helical RNA and DNA from NMR relaxation and liquid crystal measurements. J Am Chem Soc 128:11443-54
Ying, Jinfa; Bax, Ad (2006) 2'-hydroxyl proton positions in helical RNA from simultaneously measured heteronuclear scalar couplings and NOEs. J Am Chem Soc 128:8372-3
Chill, Jordan H; Louis, John M; Miller, Christopher et al. (2006) NMR study of the tetrameric KcsA potassium channel in detergent micelles. Protein Sci 15:684-98
Ying, Jinfa; Grishaev, Alexander; Bax, Ad (2006) Carbon-13 chemical shift anisotropy in DNA bases from field dependence of solution NMR relaxation rates. Magn Reson Chem 44:302-10
Dam, Julie; Baber, James; Grishaev, Alexander et al. (2006) Variable dimerization of the Ly49A natural killer cell receptor results in differential engagement of its MHC class I ligand. J Mol Biol 362:102-13

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