National High Magnet Field Lab , 1800 East Paul Dirac Drive , Tallahassee , Florida 32310 , United States.
Department of Chemistry and Biochemistry , Florida State University , Tallahassee , Florida 32306 , United States.
J Am Chem Soc. 2020 Feb 5;142(5):2115-2119. doi: 10.1021/jacs.9b09985. Epub 2020 Jan 28.
The integral membrane M2 protein is a 97-residue membrane protein that assembles as a tetramer to conduct protons at a slow rate (10-10/s) when activated by low pH. The proton conductance mechanism has been extensively debated in the literature, but it is accepted that the proton conductance is facilitated by hydrogen bonds involving the His37 residues. However, the hydrogen bonding partnership remains unresolved. Here, we report on the measurement of N-N -couplings of N His37-labeled full length M2 (M2FL) protein from virus embedded in synthetic liquid crystalline lipid bilayers using two-dimensional -resolved NMR spectroscopy. We experimentally observed the hydrogen-bond mediated -couplings between N and N of adjacent His37 imidazole rings, providing direct evidence for the existence of various imidazolium-imidazole hydrogen-bonding geometries in the histidine tetrad at low pH, thus validating the proton conduction mechanism in the M2FL protein by which the proton is transferred through the breaking and reforming of the hydrogen bonds between pairs of His37 residues.
整合膜 M2 蛋白是一种由 97 个氨基酸组成的膜蛋白，当被低 pH 值激活时，它会组装成四聚体，以缓慢的速度（10-10/s）传导质子。质子传导机制在文献中已经被广泛讨论，但人们普遍认为质子传导是通过涉及 His37 残基的氢键来促进的。然而，氢键的伙伴关系仍然没有解决。在这里，我们报告了使用二维分辨 NMR 光谱测量嵌入在合成液晶脂质双层中的 病毒全长 M2（M2FL）蛋白中 N-His37 标记的 N-N 耦合。我们实验观察到相邻 His37 咪唑环之间氢键介导的 N-N 耦合，为低 pH 下组氨酸四联体中存在各种咪唑鎓-咪唑氢键几何结构提供了直接证据，从而通过质子在 His37 残基对之间氢键的断裂和形成来验证 M2FL 蛋白中的质子传导机制。
