[2,3-13C]-labeling of Aromatic Residues-Getting a Head Start in the Magic-Angle-Spinning NMR Assignment of Membrane Proteins
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- 作者：Matthias Hiller ; Victoria A. Higman ; Stefan Jehle ; Barth-Jan van Rossum ; Werner Kü ; hlbrandt ; Hartmut Oschkinat
- 刊名：Journal of the American Chemical Society
- 出版年：2008
- 出版时间：January 16, 2008
- 年：2008
- 卷：130
- 期：2
- 页码：408 - 409
- 全文大小：359K
- 年卷期：v.130,no.2(January 16, 2008)
- ISSN：1520-5126
文摘
The assignment of magic-angle-spinning NMR spectra of large membrane proteins such as the 281-residue outer membrane protein G (OmpG) is difficult because of substantial signal overlap and line-broadening effects. To obtain sequence specific assignments, a new labeling strategy was applied to OmpG in which alanine and glycine were uniformly [15N,13C]-labeled and phenylalanine and tyrosine were [15N, C
,C
-13C]-labeled. This labeling pattern resulted in many long-range inter-residue cross-peaks being observed in spectra recorded with long mixing times. In addition, a reduction in the number of C' labels resulted in NCO-type spectra with little overlap and provided the basis for unambiguous sequential assignments. In conjunction with spectra from [1,3-13C]- and [2-13C]-glycerol samples, a total of 45 reliable sequence specific assignments could be made for OmpG on the basis of this sample.
,C-13C]-labeled. This labeling pattern resulted in many long-range inter-residue cross-peaks being observed in spectra recorded with long mixing times. In addition, a reduction in the number of C' labels resulted in NCO-type spectra with little overlap and provided the basis for unambiguous sequential assignments. In conjunction with spectra from [1,3-13C]- and [2-13C]-glycerol samples, a total of 45 reliable sequence specific assignments could be made for OmpG on the basis of this sample.