Department of Molecular and Cell Biology, University of Connecticut, Storrs, Connecticut, USA.
Biophys J. 2011 Jun 22;100(12):2946-54. doi: 10.1016/j.bpj.2011.05.015.
Rhodopsin is a kinetically stable protein constituting >90% of rod outer segment disk membrane protein. To investigate the bilayer contribution to rhodopsin kinetic stability, disk membranes were systematically disrupted by octyl-β-D-glucopyranoside. Rhodopsin kinetic stability was examined under subsolubilizing (rhodopsin in a bilayer environment perturbed by octyl-β-D-glucopyranoside) and under fully solubilizing conditions (rhodopsin in a micelle with cosolubilized phospholipids). As determined by DSC, rhodopsin exhibited a scan-rate-dependent irreversible endothermic transition at all stages of solubilization. The transition temperature (T(m)) decreased in the subsolubilizing stage. However, once the rhodopsin was in a micelle environment there was little change of the T(m) as the phospholipid/rhodopsin ratio in the mixed micelles decreased during the fully solubilized stage. Rhodopsin thermal denaturation is consistent with the two-state irreversible model at all stages of solubilization. The activation energy of denaturation (E(act)) was calculated from the scan rate dependence of the T(m) and from the rate of rhodopsin thermal bleaching at all stages of solubilization. The E(act) as determined by both techniques decreased in the subsolubilizing stage, but remained constant once fully solubilized. These results indicate the bilayer structure increases the E(act) to rhodopsin denaturation.
视紫红质是一种动力学稳定的蛋白质，占杆状外段盘膜蛋白的>90%。为了研究双层结构对视紫红质动力学稳定性的贡献，用辛基-β-D-吡喃葡萄糖苷系统地破坏盘膜。在亚溶解（双层环境中的视紫红质受到辛基-β-D-吡喃葡萄糖苷的干扰）和完全溶解条件下（胶束中的视紫红质与共溶解的磷脂）下检查视紫红质的动力学稳定性。如 DSC 所确定的，在溶解的所有阶段，视紫红质都表现出与扫描速率相关的不可逆吸热转变。转变温度（T(m)）在亚溶解阶段降低。然而，一旦视紫红质处于胶束环境中，由于在完全溶解阶段混合胶束中磷脂/视紫红质的比例降低，T(m)几乎没有变化。视紫红质热变性在溶解的所有阶段都符合两态不可逆模型。从 T(m)的扫描速率依赖性和在溶解的所有阶段视紫红质热漂白的速率计算变性的活化能（E(act)）。通过这两种技术确定的 E(act)在亚溶解阶段降低，但一旦完全溶解则保持不变。这些结果表明双层结构增加了视紫红质变性的 E(act)。
