Finn F M, Titus G, Montibeller J A, Hofmann K
J Biol Chem. 1980 Jun 25;255(12):5742-6.
Avidin can be labeled to high specific radioactivity by introducing 3-(p-hydroxyphenyl)-propionyl groups into the molecule (pHPP-avidin). 125I-pHPP-avidin binds avidly to rat liver plasma membranes and is not displaced by unlabeled pHPP-avidin. Nonspecific binding of 125I-pHPP-avidin can be substantially reduced by succinoylation of pHPP-avidin with succinic anhydride (SpHPP-avidin). Spectral changes ensuing when the dye 4-hydroxyazobenzene-2'-carboxylic acid binds to avidin cannot be used to assess the binding characteristics of the modified avidins since the absorption coefficients of the complexes are markedly different; however, the modified molecules bind theoretical amounts of [14C]biotin. Biotinylinsulin and biotinylinsulinSpHPP-avidin complexes compete with 125I-insulin for binding to receptor sites on rat liver plasma membranes. Biotinylinsulin complexes with unmodified avidin display anomalous binding behavior attributable to formation of membrane aggregates. In light of this finding, results obtained using unmodified avidin must be interpreted with caution. Biotinylinsulin125I-SpHPP-avidin binds specifically and saturably to rat liver plasma membranes. The biotinylhormoneSpHPP-avidin technique has potential for labeling peptide hormones and other compounds that cannot be iodinated by conventional procedures.
通过将3-(对羟基苯基)-丙酰基引入分子中（对羟基苯基丙酸-抗生物素蛋白），抗生物素蛋白可以被标记到高比放射性。125I-对羟基苯基丙酸-抗生物素蛋白能与大鼠肝细胞膜紧密结合，且不会被未标记的对羟基苯基丙酸-抗生物素蛋白所取代。用琥珀酸酐对羟基苯基丙酸-抗生物素蛋白进行琥珀酰化（琥珀酰化对羟基苯基丙酸-抗生物素蛋白），可大幅降低125I-对羟基苯基丙酸-抗生物素蛋白的非特异性结合。当染料4-羟基偶氮苯-2'-羧酸与抗生物素蛋白结合时产生的光谱变化不能用于评估修饰抗生物素蛋白的结合特性，因为复合物的吸收系数明显不同；然而，修饰后的分子能结合理论量的[14C]生物素。生物素化胰岛素和生物素化胰岛素-琥珀酰化对羟基苯基丙酸-抗生物素蛋白复合物与125I-胰岛素竞争结合大鼠肝细胞膜上的受体位点。未修饰抗生物素蛋白的生物素化胰岛素复合物表现出异常的结合行为，这归因于膜聚集体的形成。鉴于这一发现，使用未修饰抗生物素蛋白获得的结果必须谨慎解释。生物素化胰岛素-125I-琥珀酰化对羟基苯基丙酸-抗生物素蛋白能特异性且饱和地结合大鼠肝细胞膜。生物素化激素-琥珀酰化对羟基苯基丙酸-抗生物素蛋白技术有潜力标记肽类激素和其他无法通过常规程序进行碘化的化合物。
