Involvement of water molecules in the association of monoclonal antibody HyHEL-5 with bobwhite quail lysozyme.
Involvement of water molecules in the association of monoclonal antibody HyHEL-5 with bobwhite quail lysozyme.
复制标题
水分子参与单克隆抗体 HyHEL-5 与短白鹌鹑溶菌酶的结合。
DOI：
10.1016/s0006-3495(97)78242-0
复制
发表时间：
1997
影响因子：
3.4
通讯作者：
Willson,RC
中科院分区：
文献类型：
--
作者：
Xavier,KA;Shick,KA;Smith-Gil,SJ;Willson,RC
采用荧光偏振光谱分析和等温滴定量热法研究了渗透压对抗鸡蛋溶菌酶(HEL)单抗HyHEL-5与日本血吸虫(BWQL)结合的影响。BWQL是一种禽类变异，在HyHEL-5表位上有Arg-->；Lys突变，在HyHEL-5结构表位之外还有其他三个突变。与HEL相比，该突变使HyHEL-5与BWQL的平衡结合常数降低1000倍以上。这个复合体的三维结构是最近获得的。荧光素标记的BWQL在pH 7.5下标记，疏水作用层析纯化后与HyHEL-5结合，平衡结合常数接近等温滴定量热法测定的未标记BWQL的平衡结合常数。荧光滴定、停流动力学和等温滴定量热法实验使用不同浓度的渗透压甘油、乙二醇和甜菜碱扰动结合，得出在形成HyHEL-5/BWQL络合物时摄取大约6-12个水分子的下限。
Fluorescence polarization spectroscopy and isothermal titration calorimetry were used to study the influence of osmolytes on the association of the anti-hen egg lysozyme (HEL) monoclonal antibody HyHEL-5 with bobwhite quail lysozyme (BWQL). BWQL is an avian species variant with an Arg-->Lys mutation in the HyHEL-5 epitope, as well as three other mutations outside the HyHEL-5 structural epitope. This mutation decreases the equilibrium association constant of HyHEL-5 for BWQL by over 1000-fold as compared to HEL. The three-dimensional structure of this complex has been obtained recently. Fluorescein-labeled BWQL, obtained by labeling at pH 7.5 and purified by hydrophobic interaction chromatograpy, bound HyHEL-5 with an equilibrium association constant close to that determined for unlabeled BWQL by isothermal titration calorimetry. Fluorescence titration, stopped-flow kinetics, and isothermal titration calorimetry experiments using various concentrations of the osmolytes glycerol, ethylene glycol, and betaine to perturb binding gave a lower limit of the uptake of approximately 6–12 water molecules upon formation of the HyHEL-5/BWQL complex.