Solid-state nuclear magnetic resonance characterization of gramicidin channel structure.
The method of using orientational constraints derived from solid-state NMR for structural characterization of polypeptides in heterogeneous environments has now been demonstrated. A very high resolution structure has been achieved that has led to greater functional understanding of this channel. Much can be done to improve this structural technique to make it more efficient and more generally applicable. Others as well as ourselves are applying this approach to membrane proteins. Although solid-phase synthesis and specific site isotopic labeling has been essential for the development described here, one of the primary challenges is to be able to use amino acid-specific and uniform labeling of peptides and proteins by biosynthetic means for isotopic incorporation. This will allow for the study of many more proteins and significantly large proteins. Unlike solution NMR structural methods, there are no intrinsic molecular weight limitations. In fact, as the molecular weight increases the molecular motion will become less and the spectroscopic properties will improve. The major limitation will be sensitivity: as the molecular weight increases the number of moles will decrease in the samples, causing sensitivity to decrease. Advances in field strength and NMR technology help to address this problem. With larger molecules and more isotopically labeled sites resolution could also be a problem; however, the two- and three-dimensional methods demonstrated by Opella and co-workers clearly show the potential for enormous resolving power. In the 15N dimension alone it is shown that the resolution is greater than in solution NMR. Although challenges such as spectral assignments have yet to be completely solved, several approaches have been described, and the prospects are excellent for solving this and other problems facing the development of this novel approach for structural elucidation. Although there is an attempt to get away from solid-phase synthesis to solve larger molecular weight structures, peptide synthesis will continue to be important for generating single- and double-site labeled model compounds for characterizations of spin interaction tensors. Such characterizations will continue to be a very important aspect of this structural approach.
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Center for Interdisciplinary Magnetic Resonance, Florida State University, Talahassee 32306-4005, USA.
D006096:Gramicidin / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*
D006096:Gramicidin