Cierpicki Tomasz, Liang Binyong, Tamm Lukas K, Bushweller John H
Department of Molecular Physiology and Biological Physics, University of Virginia, Charlottesville, VA 22908, USA.
J Am Chem Soc. 2006 May 31;128(21):6947-51. doi: 10.1021/ja0608343.
The structure determination of membrane proteins is one of the most challenging applications of solution NMR spectroscopy. The paucity of distance information available from the highly deuterated proteins employed requires new approaches in structure determination. Here we demonstrate that significant improvement in the structure accuracy of the membrane protein OmpA can be achieved by refinement with residual dipolar couplings (RDCs). The application of charged polyacrylamide gels allowed us to obtain two alignments and accurately measure numerous heteronuclear dipolar couplings. Furthermore, we have demonstrated that using a large set of RDCs in the refinement can yield a structure with 1 A rms deviation to the backbone of the high-resolution crystal structure. Our simulations with various data sets indicate that dipolar couplings will be critical for obtaining accurate structures of membrane proteins.
膜蛋白的结构测定是溶液核磁共振波谱学最具挑战性的应用之一。由于使用的高度氘代蛋白质可获得的距离信息匮乏，需要采用新的结构测定方法。在此我们证明，通过利用残余偶极耦合（RDCs）进行精修，可以显著提高膜蛋白OmpA的结构准确性。使用带电聚丙烯酰胺凝胶使我们能够获得两种排列并准确测量大量异核偶极耦合。此外，我们已经证明，在精修中使用大量的RDCs可以得到一个与高分辨率晶体结构主链的均方根偏差为1埃的结构。我们对各种数据集的模拟表明，偶极耦合对于获得膜蛋白的准确结构至关重要。
