AFM characterization of tilt and intrinsic flexibility of Rhodobacter sphaeroides light harvesting complex 2 (LH2).
AFM characterization of tilt and intrinsic flexibility of Rhodobacter sphaeroides light harvesting complex 2 (LH2).
复制标题
球形红细菌光捕获复合物 2 (LH2) 的倾斜和内在灵活性的 AFM 表征。
DOI：
10.1016/s0022-2836(02)01241-x
复制
发表时间：
2003
影响因子：
5.6
通讯作者：
J. Rigaud
中科院分区：
文献类型：
--
作者：
S. Scheuring;J. Seguin;S. Marco;D. Lévy;C. Breyton;B. Robert;J. Rigaud
原子力显微镜（AFM）已经发展成为一个强大的工具，在接近自然的环境中研究膜蛋白表面。在这里，我们报告的表面形貌Rhodobacter sphaeroides捕光复合物2（LH 2）重组成二维晶体。这些光合作用跨膜蛋白形成圆柱形的寡聚复合物，其倾斜地插入到脂质膜中。这种独特的包装的一个完整的膜蛋白，使我们能够确定低聚化和倾斜的LH 2复合物，但也突出的高度和内在的灵活性，其个别亚基。此外，表面轮廓的可靠性和原子力显微镜的限制可以进行研究。所检测的二维晶体的尺寸可达5μm，并且如10 μ m低温电子显微镜投影图所示，p22121晶体对称。晶胞的尺寸为a=B= 150 °，γ=90°，包含四个九聚体复合物，两个向上，两个向下。这些二维晶体的AFM形貌图具有10 μ m的横向分辨率。此外，高垂直分辨率为100 nm，允许确定圆柱形LH 2复合物在膜上的突出高度。这是最大的13.1在一边和3.8在另一边。有趣的是，突起高度在LH 2复合物上变化，显示复合物相对于膜平面以6.2°倾斜插入。对各个亚基的详细分析显示，膜突出肽段的固有柔性相等且独立于其突出高度。此外，我们对这种特殊包装中的膜蛋白的分析证实了原子力显微镜在生物样品上的高垂直分辨率，并使我们得出结论，图像采集功能对于高度高达150厘米的轮廓突起同样准确。
Atomic force microscopy (AFM) has developed into a powerful tool to investigate membrane protein surfaces in a close-to-native environment. Here we report on the surface topography of Rhodobacter sphaeroides light harvesting complex 2 (LH2) reconstituted into two-dimensional crystals. These photosynthetic trans-membrane proteins formed cylindrical oligomeric complexes, which inserted tilted into the lipid membrane. This peculiar packing of an integral membrane protein allowed us to determine oligomerization and tilt of the LH2 complexes, but also protrusion height and intrinsic flexibility of their individual subunits. Furthermore the surface contouring reliability and limits of the atomic force microscopy could be studied. The two-dimensional crystals examined had sizes of up to 5μm and, as revealed by a 10Å cryo electron microscopy projection map, p22121crystal symmetry. The unit cell had dimensions of a=b=150Å and γ=90°, and housed four nonameric complexes, two pointing up and two pointing down. AFM topographs of these 2D crystals had a lateral resolution of 10Å. Further, the high vertical resolution of ∼1Å, allowed the protrusion height of the cylindrical LH2 complexes over the membrane to be determined. This was maximally 13.1Å on one side and 3.8Å on the other. Interestingly, the protrusion height varied across the LH2 complexes, showing the complexes to be inserted with a 6.2° tilt with respect to the membrane plane. A detailed analysis of the individual subunits showed the intrinsic flexibility of the membrane protruding peptide stretches to be equal and independent of their protrusion height. Furthermore, our analysis of membrane proteins within this peculiar packing confirmed the high vertical resolution of the atomic force microscopy on biological samples, and led us to conclude that the image acquisition function was equally accurate for contouring protrusions with heights up to ∼15Å.
DOI：
10.1073/pnas.95.11.5935
发表时间：
1998-05-26
影响因子：
11.1
作者：
Hu, XC;Damjanovic, A;Schulten, K
通讯作者：
Schulten, K
影响因子：
5.6
作者：
HENDERSON, R;BALDWIN, JM;DOWNING, KH
通讯作者：
DOWNING, KH
影响因子：
5.7
作者：
Koepke, J;Hu, XC;Michel, H
通讯作者：
Michel, H