Ward R, Zoltner M, Beer L, El Mkami H, Henderson I R, Palmer T, Norman D G
College of Life Sciences, University of Dundee, Dundee, UK.
Structure. 2009 Sep 9;17(9):1187-94. doi: 10.1016/j.str.2009.07.011.
The outer membrane beta-barrel trans-membrane proteins in gram-negative bacteria are folded into the membrane with the aid of polypeptide transport-associated (POTRA) domains. These domains occur, and probably function, as a tandem array situated on the periplasmic side of the outer membrane. Two crystal structures and one NMR study have attempted to define the structure and articulation of the POTRA domains of the Escherichia coli, prototypic Omp85 protein BamA. We have used pulsed electron paramagnetic resonance (EPR) to determine the distance and distance distribution between (1-Oxyl-2,2,5,5-tetramethylpyrroline-3-methyl) methanethiosulfonate spin labels (MTSSL), placed across the domain interface of the first two POTRA domains of BamA. Our results show tightly defined interdomain distance distributions that indicate a well-defined domain orientation. Examination of the known structures revealed that none of them fitted the EPR data. A combination of EPR and NMR data was used to generate converged structures with defined domain-domain orientation.
革兰氏阴性菌的外膜β桶跨膜蛋白借助多肽转运相关（POTRA）结构域折叠进入膜内。这些结构域以串联阵列的形式存在于外膜的周质侧，并且可能发挥相应功能。两项晶体结构研究和一项核磁共振研究试图确定大肠杆菌的原型Omp85蛋白BamA的POTRA结构域的结构和连接方式。我们利用脉冲电子顺磁共振（EPR）来确定放置在BamA前两个POTRA结构域的结构域界面上的（1-氧基-2,2,5,5-四甲基吡咯啉-3-甲基）甲硫基磺酸盐自旋标记（MTSSL）之间的距离和距离分布。我们的结果显示出紧密定义的结构域间距离分布，这表明结构域的取向明确。对已知结构的检查表明，没有一个结构符合EPR数据。结合EPR和NMR数据生成了具有明确结构域-结构域取向的收敛结构。
