A SOLID STATE NMR INVESTIGATION OF BACTERIORHODOPSIN

这项研究的目的是展示在固体中对均匀标记的膜蛋白进行化学位移和位移各向异性的方法。这是一个具有挑战性的项目，然而，还没有统一的13C，15N标记的蛋白质，即使是中等大小的固体还没有完全分配。核磁共振方法(脉冲序列、硬件)已经有了显著的改进，这表明这是一种可能性。特别是，具有更高效率的偶极重新耦合技术、先进的去耦合方法、能够进行25-30 kHzMAS和200 kHz去耦场的探针以及具有750 MHz 1H频率的宽带磁体应该能够实现膜蛋白的全自态核磁共振结构测定。考虑到侧链和视网膜之间的重要相互作用，视网膜结合袋中的化学位移将特别值得分析。这些位移将与量子化学方法一起用于详细分析质子被单轴泵浦的机制。

The objective of the proposed research is to demonstrate methods to assign chemical shifts and shift anisotrophies on uniformly labeled membrane proteins in the solid state. This is a challenging project however, no uniformly 13C, 15N labeled protein of even moderate size has yet been fully assigned in the solid state. There have been significant improvements in NMR methodology (pulse sequence, hardware) which point to this as a possibility. In particular, dipolar recoupling techniques with higher efficiencies, advanced decoupling methods, probes capable of 25-30 kHz MAS and 200 kHz decoupling fields, and widebore magnets with 1H frequencies of 750 MHz should lead to full sold-state NMR structure determinations of membrane proteins. The chemical shifts in the retinal binding pocket will be particularly interesting to analyze, given the significant interactions between the sidechains and retinal. These shifts will be used along with quantum chemical methods to analyze in detail the mechanism by which the proton is pumped uniaxially.