Force measurements for membrane protein manipulation
Force measurements for membrane protein manipulation
复制标题
膜蛋白操作的力测量
DOI：
10.1016/s0927-7765(01)00230-2
复制
发表时间：
2002
期刊：
影响因子：
--
通讯作者：
T. Osada
中科院分区：
文献类型：
--
作者：
A. Ikai;R. Afrin;A. Itoh;H. Thøgersen;Y. Hayashi;T. Osada
原子力显微镜（AFM）的力曲线测量模式使我们能够测量迄今为止无法观察到的纳米尺寸生物样品的力学性能。通过应用这种模式，我们试图进行这样的膜蛋白的机械操作：（1）膜结合的受体和共价交联的配体分子之间的AFM针尖的分离力的测量;和（2）膜蛋白的提取后，利用它们的修饰与共价交联剂的尖端。由于在我们的实验中使用的共价体系的极限张力是成功操作的关键因素，我们首先基于我们先前的数据估计在单分子水平终止共价交联体系的力为1.6-1.7 nN。然后应用该方法测量将α2-巨球蛋白（α2-M）与其在细胞膜上的受体分离所需的力，使用涂覆有蛋白质的受体结合形式的AFM针尖。根据断裂力的双峰分布，我们获得了120 pN的平均值作为分离α2-M与其受体的非共价结合的力。当使用针对细胞表面上的氨基的具有共价交联剂的改性尖端时，断裂力的分布向更高的值移动，直方图中的峰值为400-500 pN。由于切断共价交联系统的力为1.6-1.7 nN，因此观察到的力归因于共价键形成后从细胞膜提取膜蛋白所需的力。
The force curve measurement mode of the atomic force microscope (AFM) enables us to measure hitherto unobservable mechanical properties of nanometer sized biological specimens. By applying this mode, we attempted to conduct such mechanical manipulations of membrane proteins as: (1) measurement of the separation force between a membrane bound receptor and a covalently cross-linked ligand molecule on the AFM tip; and (2) extraction of membrane proteins after harnessing them on a modified tip with covalent cross-linkers. Since the limiting tensile force of the covalent system used in our experiment was a crucial factor for successful manipulations, we first estimated the force to terminate the covalent cross-linking system at the single molecular level to be 1.6–1.7 nN, based on our previous data. The method was then applied to measure the force required to separate α2-macroglobulin (α2-M) from its receptor on the cell membrane using an AFM tip coated with the receptor binding form of the protein. From a bimodal distribution of rupture force, we obtained an average value of 120 pN as the force to separate a non-covalent association of α2-M with its receptor. When modified tips with covalent cross-linkers aimed at amino groups on the cell surface were used, distribution of the rupture force shifted toward higher values, with a peak in the histogram ≈400–500 pN. Since the force to sever covalent cross-linking system was 1.6–1.7 nN, the observed force was ascribed to the force required to extract membrane proteins from the cell membrane after covalent bond formation.
影响因子：
3.4
作者：
N. Tao;S. Lindsay;S. Lees
通讯作者：
N. Tao;S. Lindsay;S. Lees
影响因子：
3.4
作者：
Chen, A;Moy, VT
通讯作者：
Moy, VT