The structure of bacteriorhodopsin at 3.0 Å resolution based on electron crystallography:: Implication of the charge distribution
The structure of bacteriorhodopsin at 3.0 Å resolution based on electron crystallography:: Implication of the charge distribution
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DOI：
10.1006/jmbi.1998.2529
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发表时间：
1999-02-26
影响因子：
5.6
通讯作者：
Fujiyoshi, Y
中科院分区：
文献类型：
--
作者：
Mitsuoka, K;Hirai, T;Fujiyoshi, Y
电子晶体学具有可视化原子电荷状态的潜力。这是由于在低分辨率范围内（通常小于5埃），中性和电离原子对电子的散射因子显著不同。在之前的工作中，我们在光驱动质子泵细菌视紫红质（bR）的实验（\F-o\）图中观察到酸性残基周围有两种不同类型的密度。我们认为这些可能反映了酸性残基的不同状态，即质子化（中性）和去质子化（带负电荷）状态。为了更定量地评估所观察到的电荷，我们使用我们的电子晶体学数据集在8.0和3.0埃分辨率之间改进了bR和八种周围脂质的原子模型，其中电荷效应很小。改进后的模型产生的X因子为23.7%，自由R因子为33.0%。为了评估电荷对密度图的影响，我们计算了一个差值（\F-o-\F-c\）图，其中包括分辨率低于8.0埃的数据。分辨率，其中电荷效应显着。我们在差异图谱中发现了主要在跨膜螺旋的骨架区域中的强峰。我们将这些峰解释为来自α-螺旋主链中极性基团的极化，并通过假设肽羰基的部分电荷为0.5来对此进行检查。由此产生的R和自由R因子分别从0.250和0.341下降到0.246和0.336。此外，我们还观察到一些强峰周围的一些侧链，这可以被分配给带正电荷的原子。因此，我们可以表明Asp 36和Asp 102可能与附近的阳离子相互作用。此外，在酸性残基Glu 74、Glu 194和Glu 212周围发现的峰具有不同的特征，并且可能代表极化水分子或水合氢离子上的正电荷。(C)北京：科学出版社.
Electron crystallography has the potential to visualise the charge status of atoms. This is due to the significantly different scattering factors of neutral and ionised atoms for electrons in the low-resolution range (typically less than 5 Angstrom). In previous work, we observed two different types of densities around acidic residues ill the experimental (\F-o\) map of bacteriorhodopsin (bR), a light-driven proton pump. We suggested that these might reflect different states of the acidic residues; namely, the protonated (neutral) and the deprotonated (negatively charged) state. To evaluate the observed charge more quantitatively, we refined the atomic model for bR and eight surrounding lipids using our electron crystallographic data set between 8.0 and 3.0 Angstrom resolution, where the charge effect is small. The refined model yielded an X-factor of 23.7 % and a free R-factor of 33.0%.To evaluate the effect of charges on the density map, we calculated a difference (\F-o-\F-c\) map including data of a resolution lower than 8.0 Angstrom resolution, where the charge effect is significant. We found strong peaks in the difference map mainly in the backbone region of the transmembrane helices. We interpreted these peaks to come from the polarisation of the polar groups in the main chain of the alpha-helices and we examined this by assuming a partial charge of 0.5 for the peptide carbonyl groups. The resulting R and free R-factors dropped from 0.250 and 0.341 to 0.246 and 0.336, respectively. Furthermore, we also observed some strong peaks around some side-chains, which could be assigned to positively charged atoms. Thus, we could show that Asp36 and Asp102 are likely to interact with cations nearby. In addition, peaks found around the acidic residues Glu74, Glu194 and Glu212 have different features and might represent positive charges on polarised water molecules or hydroxonium ions. (C) 1999 Academic Press.