Determination of reserve albumin-equivalent for ligand binding, probing two distinct binding functions of the protein.
Determination of reserve albumin-equivalent for ligand binding, probing two distinct binding functions of the protein.
复制标题
确定配体结合的储备白蛋白当量，探测蛋白质的两种不同的结合功能。
DOI：
10.1016/0003-2697(82)90499-7
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发表时间：
1982
影响因子：
2.9
通讯作者：
Larsen,S
中科院分区：
文献类型：
--
作者：
Brodersen,R;Andersen,S;Jacobsen,C;Sønderskov,O;Ebbesen,F;Cashore,WJ;Larsen,S
本文报道了一种在50 μl样品体积中测定各种配体与白蛋白结合容量的方法。将少量放射性标记的测试配体添加到未稀释的样品中，并测量游离配体透析到没有添加配体的相同样品中的速率。储备白蛋白当量浓度定义为标准白蛋白制剂的浓度，其在缓冲溶液中提供相同的透析速率，因此提供相同的添加配体的游离/结合浓度比。结果表明，储备白蛋白当量浓度，因此定义，是相同的载体物种的浓度的总和，每个乘以第一化学计量结合常数的测试配体的载体，并除以其第一化学计量结合常数的标准白蛋白。该参数的测定适用于研究单个配体的化学势和转移亲和力以及测定几种结合物质之间的相互作用。已使用两种测试配体，单乙酰基二氨基二苯砜和地西泮。前者与胆红素竞争性结合，而地西泮具有另一种独立的结合功能。因此，该方法可用于单独测定白蛋白的两种不同结合功能的饱和度。可以研究几种载体蛋白与相互作用配体的复杂混合物。
A method is reported for determination of albumin binding capacity for various ligands in 50-μl sample volumes. A small amount of a radioactively labeled test ligand is added to the undiluted sample and the rate of dialysis of the free ligand into an identical sample without added ligand is measured. The reserve albumin-equivalent concentration is defined as the concentration of a standard albumin preparation which in buffered solution gives the same rate of dialysis and hence the same ratio of free/bound concentrations of the added ligand. It is shown that the reserve albumin-equivalent concentration, thus defined, is identical with the sum of concentrations of carrier species, each multiplied by the first stoichiometric binding constant of the test ligand to the carrier and divided by its first stoichiometric binding constant to the standard albumin. Determinations of this parameter are suitable for studies of the chemical potential and transfer affinities of individual ligands and for determination of interaction among several binding substances. Two test ligands have been used, monoacetyldiaminodiphenyl sulfone and diazepam. The former is bound competitively with bilirubin while diazepam engages another, independent binding function. The method can thus be used for separate determinations of the degree of saturation of two distinct binding functions of albumin. Complex mixtures of several carrier proteins with interacting ligands can be studied.