Ardhammar Malin, Lincoln Per, Nordén Bengt
Department of Physical Chemistry, Chalmers University of Technology, SE-412 96 Gothenburg, Sweden.
Proc Natl Acad Sci U S A. 2002 Nov 26;99(24):15313-7. doi: 10.1073/pnas.192583499. Epub 2002 Nov 6.
Valuable information on protein-membrane organization may in principle be obtained from polarized-light absorption (linear dichroism, LD) measurement on shear-aligned lipid vesicle bilayers as model membranes. However, attempts to probe LD in the UV wavelength region (<250 nm) have so far failed because of strong polarized light scattering from the vesicles. Using sucrose to match the refractive index and suppress the light scattering of phosphatidylcholine vesicles, we have been able to detect LD bands also in the peptide-absorbing region (200-230 nm). The potential of refractive index matching in vesicle LD as a general method for studying membrane protein structure was investigated for the membrane pore-forming oligopeptide gramicidin incorporated into the liposome membranes. In the presence of sucrose, the LD signals arising from oriented tryptophan side chains as well as from n-->pi* and pi-->pi* transitions of the amide chromophore of the polypeptide backbone could be studied. The observation of a strongly negative LD for the first exciton transition ( approximately 204 nm) is consistent with a membrane-spanning orientation of two intertwined parallel gramicidin helices, as predicted by coupled-oscillator theory.
原则上，通过对剪切排列的脂质囊泡双层膜（作为模型膜）进行偏振光吸收（线性二色性，LD）测量，可以获得有关蛋白质 - 膜组织的有价值信息。然而，由于囊泡强烈的偏振光散射，迄今为止在紫外波长区域（<250 nm）探测LD的尝试均告失败。我们利用蔗糖匹配折射率并抑制磷脂酰胆碱囊泡的光散射，得以在肽吸收区域（200 - 230 nm）检测到LD带。针对掺入脂质体膜中的膜孔形成寡肽短杆菌肽，研究了囊泡LD中折射率匹配作为研究膜蛋白结构的通用方法的潜力。在蔗糖存在的情况下，可以研究由取向色氨酸侧链以及多肽主链酰胺发色团的n→π和π→π跃迁产生的LD信号。如耦合振荡器理论所预测的那样，对于第一个激子跃迁（约204 nm）观察到的强负LD与两个相互缠绕的平行短杆菌肽螺旋的跨膜取向一致。
