Helix conformations in 7TM membrane proteins determined using oriented-sample solid-state NMR with multiple residue-specific 15N labeling
Helix conformations in 7TM membrane proteins determined using oriented-sample solid-state NMR with multiple residue-specific 15N labeling
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DOI：
10.1529/biophysj.107.116004
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发表时间：
2008-01-01
影响因子：
3.4
通讯作者：
Nielsen, Niels Chr.
中科院分区：
文献类型：
--
作者：
Vosegaard, Thomas;Kamihira-Ishijima, Miya;Nielsen, Niels Chr.
定向固态NMR结合多残基特异性N-15标记和广泛的数值光谱分析，提出了确定天然膜中的大膜蛋白的螺旋构象。该方法被证明对单轴定向样品的N-15-蛋氨酸，缬氨酸，甘氨酸标记的细菌跳蛋白在本地紫色膜。实验二维H-1-N-15偶极偶极耦合与N-15化学位移光谱的所有样品进行了数值分析，以建立七个跨膜螺旋相对于膜双层正常的方向上的组合约束。由于该方法不依赖于特定的共振分配，并证明对非理想性的样品比对鲁棒性，它可能是普遍可行的构象排列和功能诱导的构象变化的研究大的完整的膜蛋白。
Oriented solid-state NMR in combination with multiple-residue-specific N-15 labeling and extensive numerical spectral analysis is proposed to determine helix conformations of large membrane proteins in native membranes. The method is demonstrated on uniaxially oriented samples of N-15-methionine, -valine, and -glycine-labeled bacteriorhopsin in native purple membranes. Experimental two-dimensional H-1-N-15 dipole-dipole coupling versus N-15 chemical shift spectra for all samples are analyzed numerically to establish combined constraints on the orientation of the seven transmembrane helices relative to the membrane bilayer normal. Since the method does not depend on specific resonance assignments and proves robust toward nonidealities in the sample alignment, it may be generally feasible for the study of conformational arrangement and function-induced conformation changes of large integral membrane proteins.