Membrane Protein Structures in Native Cellular Membranes Revealed by Solid-State NMR Spectroscopy.
Membrane Protein Structures in Native Cellular Membranes Revealed by Solid-State NMR Spectroscopy.
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固态核磁共振光谱揭示天然细胞膜中的膜蛋白结构。
DOI：
10.1021/jacsau.3c00564
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发表时间：
2023-12-25
期刊：
影响因子：
8
通讯作者：
Yang, Jun
中科院分区：
文献类型：
--
作者：
Zhang, Yan;Gan, Yuefang;Zhao, Weijing;Zhang, Xuning;Zhao, Yongxiang;Xie, Huayong;Yang, Jun
细胞膜环境中膜蛋白的结构表征对于理解其天然功能背景下的分子机制至关重要。然而，通过固态NMR对天然膜中的膜蛋白进行残基位点特异性结构分析面临着挑战，这是由于光谱灵敏度差和背景蛋白信号的严重干扰。在这项研究中，我们提出了一个新的协议，结合了各种战略的细胞膜样品的制备，使我们能够揭示机械敏感通道的二级结构的大电导乙酸甲烷八叠球菌（MaMscL）在大肠杆菌内膜。我们的研究结果表明，实现完整的共振分配的可行性和确定细胞膜内的膜蛋白的3D结构的潜力。我们发现在该方案中使用BL 21（DE3）菌株对于有效抑制背景蛋白标记而不损害靶蛋白的灵敏度是至关重要的。此外，我们的数据表明，不同蛋白质的结构表现出不同程度的敏感性膜环境。这些结果强调了在进行结构表征时研究其天然细胞膜内的膜蛋白的意义。总的来说，这项研究开辟了一条新的途径，实现其天然细胞膜内的膜蛋白的原子分辨率的结构表征，提供了宝贵的见解膜蛋白的天然性。
The structural characterization of membrane proteins within the cellular membrane environment is critical for understanding the molecular mechanism in their native functional context. However, conducting residue site-specific structural analysis of membrane proteins in native membranes by solid-state NMR faces challenges due to poor spectral sensitivity and serious interference from background protein signals. In this study, we present a new protocol that combines various strategies for cellular membrane sample preparations, enabling us to reveal the secondary structure of the mechanosensitive channel of large conductance from Methanosarcina acetivorans (MaMscL) in Escherichia coli inner membranes. Our findings demonstrate the feasibility of achieving complete resonance assignments and the potential for determining the 3D structures of membrane proteins within cellular membranes. We find that the use of the BL21(DE3) strain in this protocol is crucial for effectively suppressing background protein labeling without compromising the sensitivity of the target protein. Furthermore, our data reveal that the structures of different proteins exhibit varying degrees of sensitivity to the membrane environment. These results underscore the significance of studying membrane proteins within their native cellular membranes when performing structural characterizations. Overall, this study opens up a new avenue for achieving the atomic-resolution structural characterization of membrane proteins within their native cellular membranes, providing valuable insights into the nativeness of membrane proteins.
影响因子：
5.6
作者：
Ernst M;Robertson JL
通讯作者：
Robertson JL
影响因子：
16.6
作者：
Miao, Yimin;Qin, Huajun;Fu, Riqiang;Sharma, Mukesh;Can, Thach V.;Hung, Ivan;Luca, Sorin;Gor'kov, Peter L.;Brey, William W.;Cross, Timothy A.
通讯作者：
Cross, Timothy A.
影响因子：
15
作者：
Fu, Riqiang;Wang, Xingsheng;Li, Conggang;Santiago-Miranda, Adriana N.;Pielak, Gary J.;Tian, Fang
通讯作者：
Tian, Fang