Papavoine C H, Konings R N, Hilbers C W, van de Ven F J
Laboratory of Biophysical Chemistry, NSR Center, University of Nijmegen, The Netherlands.
Biochemistry. 1994 Nov 8;33(44):12990-7. doi: 10.1021/bi00248a007.
The major coat protein (gVIIIp) of bacteriophage M13 solubilized in sodium dodecyl sulfate (SDS) detergent micelles was used as a model system to study this protein in the lipid-bound form. In order to probe the position of gVIIIp relative to the SDS micelles, stearate was added, spin-labeled at the 5- or 16-position with a doxyl group containing a stable nitroxide radical. The average position of the spin-labels in the micelles was derived from the line broadening of the resonances in the 13C spectrum of SDS. Subsequently, we derived a model of the relative position of gVIIIp in the SDS micelle from the effect of the spin-labels on the gVIIIp resonances, monitored via 1H-15N HSQC and TOCSY experiments. The results are consistent with the structure of gVIIIp having two helical strands. One strand is a long hydrophobic helix that spans the micelle, and the other is a shorter amphipathic helix on the surface of the micelle. These results are in good agreement with the structure of gVIIIp in membranes proposed by McDonnell et al. on the basis of solid state NMR data [McDonnell, P. A., Shon, K., Kim, Y., & Opella, S. J. (1993) J. Mol. Biol. 233, 447-463]. This study indicates that high-resolution NMR on this membrane protein, solubilized in detergent micelles, is a very suitable technique for mimicking these proteins in their natural environment. Furthermore, the data indicate that the structure of the micelle near the C-terminus of the major coat protein is distorted.(ABSTRACT TRUNCATED AT 250 WORDS)
噬菌体M13的主要外壳蛋白（gVIIIp）溶解于十二烷基硫酸钠（SDS）去污剂胶束中，被用作研究脂质结合形式下该蛋白的模型系统。为了探究gVIIIp相对于SDS胶束的位置，添加了在5位或16位用含有稳定氮氧自由基的羟胺基团自旋标记的硬脂酸盐。自旋标记物在胶束中的平均位置源自SDS的13C谱中共振峰的线宽。随后，我们通过1H-15N HSQC和TOCSY实验监测自旋标记物对gVIIIp共振峰的影响，从而得出gVIIIp在SDS胶束中相对位置的模型。结果与gVIIIp具有两条螺旋链的结构一致。一条链是跨越胶束的长疏水螺旋，另一条是胶束表面较短的两亲螺旋。这些结果与McDonnell等人基于固态NMR数据提出的膜中gVIIIp的结构[McDonnell, P. A., Shon, K., Kim, Y., & Opella, S. J. (1993) J. Mol. Biol. 233, 447 - 463]高度吻合。这项研究表明，对溶解于去污剂胶束中的这种膜蛋白进行高分辨率NMR是在其天然环境中模拟这些蛋白的非常合适的技术。此外，数据表明主要外壳蛋白C末端附近的胶束结构发生了扭曲。（摘要截短于250字）
