期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2013
卷号：110
期号：42
页码：16838-16843
DOI：10.1073/pnas.1307678110
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：Knowing the topology and location of protein segments at water-membrane interfaces is critical for rationalizing their functions, but their characterization is challenging under physiological conditions. Here, we debut a unique spectroscopic approach by using the hydration dynamics gradient found across the phospholipid bilayer as an intrinsic ruler for determining the topology, immersion depth, and orientation of protein segments in lipid membranes, particularly at water-membrane interfaces. This is achieved through the site-specific quantification of translational diffusion of hydration water using an emerging tool, 1H Overhauser dynamic nuclear polarization (ODNP)-enhanced NMR relaxometry. ODNP confirms that the membrane-bound region of -synuclein (S), an amyloid protein known to insert an amphipathic -helix into negatively charged phospholipid membranes, forms an extended -helix parallel to the membrane surface. We extend the current knowledge by showing that residues 90-96 of bound S, which is a transition segment that links the -helix and the C terminus, adopt a larger loop than an idealized -helix. The unstructured C terminus gradually threads through the surface hydration layers of lipid membranes, with the beginning portion residing within 5-15 A above the phosphate level, and only the very end of C terminus surveying bulk water. Remarkably, the intrinsic hydration dynamics gradient along the bilayer normal extends to 20-30 A above the phosphate level, as demonstrated with a peripheral membrane protein, annexin B12. ODNP offers the opportunity to reveal previously unresolvable structure and location of protein segments well above the lipid phosphate, whose structure and dynamics critically contribute to the understanding of functional versatility of membrane proteins.
