Kangur Liina, Timpmann Kõu, Freiberg Arvi
Institute of Physics, University of Tartu, Tartu, Estonia.
J Phys Chem B. 2008 Jul 3;112(26):7948-55. doi: 10.1021/jp801943w. Epub 2008 Jun 7.
The bacteriochlorophyll a-containing LH2 and LH3 antenna complexes are the integral membrane proteins that catalyze the photosynthetic process in purple photosynthetic bacteria. The LH2 complex from Rhodobacter sphaeroides shows characteristic strong absorbance at 800 and 850 nm due to the pigment molecules confined in two separate areas of the protein. In the LH3 complex from Rhodopesudomonas acidophila the corresponding bands peak at 800 and 820 nm. Using the bacteriochlorophyll a cofactors as intrinsic probes to monitor local changes in the protein structure, we investigate spectral responses of the antenna complexes to very high hydrostatic pressures up to 2.5 GPa when embedded into natural membrane environment or extracted with detergent. We first demonstrate that high pressure does induce significant alterations to the tertiary structure of the proteins not only in proximity of the 800 nm-absorbing bacteriochlorophyll a molecules known previously (Gall, A.; et al. Biochemistry 2003, 42, 13019) but also of the 850 nm- and 820 nm-absorbing molecules, including breakage of the hydrogen bond they are involved in. The membrane-protected complexes appear more resilient to damaging effects of the compression compared with the complexes extracted into mixed detergent-buffer environment. Increased resistance of the isolated complexes is observed at high protein concentration resulting aggregation as well as when cosolvent (glycerol) is added into the solution. These stability variations correlate with ability of penetration of the surrounding polar solvent (water) into the hydrophobic protein interiors, being thus the principal reason of the pressure-induced denaturation of the proteins. Considerable variability of elastic properties of the isolated complexes was also observed, tentatively assigned to heterogeneous protein packing in detergent micelles. While a number of the isolated complexes release most of their bacteriochlorophyll a content under high pressure, quite some of them remain apparently intact. The pigmented photosynthetic antenna complexes thus constitute a suitable model system for studying in detail the stability of integral membrane proteins.
含细菌叶绿素a的LH2和LH3天线复合物是催化紫色光合细菌光合作用过程的整合膜蛋白。球形红杆菌的LH2复合物由于色素分子局限于蛋白质的两个不同区域，在800和850 nm处显示出特征性的强吸收。在嗜酸红假单胞菌的LH3复合物中，相应的吸收带在800和820 nm处达到峰值。我们使用细菌叶绿素a辅因子作为内在探针来监测蛋白质结构的局部变化，研究天线复合物在嵌入天然膜环境或用去污剂提取时，对高达2.5 GPa的极高静水压力的光谱响应。我们首先证明，高压不仅会对先前已知的吸收800 nm光的细菌叶绿素a分子附近的蛋白质三级结构产生显著改变（Gall, A.; 等人，《生物化学》2003年，42卷，13019页），还会对吸收850 nm和820 nm光的分子产生显著改变，包括它们所参与的氢键断裂。与提取到混合去污剂 - 缓冲液环境中的复合物相比，膜保护的复合物对压缩的破坏作用表现出更强的弹性。在高蛋白浓度导致聚集以及向溶液中添加共溶剂（甘油）时，观察到分离出的复合物的抗性增加。这些稳定性变化与周围极性溶剂（水）渗透到疏水蛋白质内部的能力相关，因此是压力诱导蛋白质变性的主要原因。还观察到分离出的复合物的弹性性质存在相当大的变异性，初步认为这是由于去污剂胶束中蛋白质堆积不均一所致。虽然许多分离出的复合物在高压下释放出大部分细菌叶绿素a含量，但相当一部分复合物显然仍保持完整。因此，色素光合天线复合物构成了一个适合详细研究整合膜蛋白稳定性的模型系统。
