Leckband D E, Kuhl T, Wang H K, Herron J, Müller W, Ringsdorf H
Department of Chemical Engineering, University of Illinois at Urbana, Champaign 61801, USA.
Biochemistry. 1995 Sep 12;34(36):11467-78. doi: 10.1021/bi00036a020.
The surface forces apparatus was used to identify the molecular forces that control the interactions of monoclonal 4-4-20 antifluorescyl IgG Fab' fragments with fluorescein-presenting supported planar bilayers. At long range, the electrostatic force between oriented Fab' and fluorescein monolayers was controlled by the composition of the protein exterior surrounding the antigen-combining site rather than by the overall protein charge. The measured positive electrostatic potential of the Fab' monolayer at pH > pI(Fab') was consistent with the structure of the exposed Fab' surface in which a ring of positive charge at the mouth of the antigen-combining site dominates the local electrostatic surface properties. Substantial differences in the electrostatic forces measured with denatured Fab' further demonstrated that the measured electrostatic surface properties and the consequent long-range interaction forces are controlled by the protein surface composition. At short range, the strength of the Fab'-mediated adhesion was modulated not only by the length of the fluorescein tether but also by membrane hydration. Steric hydration barriers at the membrane surface reduced the adhesion strength in proportion to their range of influence. These results provide direct evidence that long-range protein interactions with immobilized ligands are controlled by both the protein and the membrane surface compositions, while short-range, specific binding is modulated by both the protein structure and the membrane interfacial properties.
表面力仪被用于识别控制单克隆4-4-20抗荧光素IgG Fab'片段与呈现荧光素的支撑平面双层相互作用的分子力。在远距离时，定向的Fab'和荧光素单层之间的静电力由围绕抗原结合位点的蛋白质外部组成控制，而非由蛋白质的整体电荷控制。在pH > pI(Fab')时测得的Fab'单层的正静电势与暴露的Fab'表面结构一致，其中抗原结合位点开口处的正电荷环主导了局部静电表面性质。用变性Fab'测得的静电力存在显著差异，进一步证明测得的静电表面性质以及随之而来的长程相互作用由蛋白质表面组成控制。在近距离时，Fab'介导的黏附强度不仅受荧光素连接链长度的调节，还受膜水合作用的调节。膜表面的空间水合屏障按其影响范围成比例地降低黏附强度。这些结果提供了直接证据，表明蛋白质与固定化配体的长程相互作用由蛋白质和膜表面组成共同控制，而短程特异性结合则由蛋白质结构和膜界面性质共同调节。
