SURFACE-SPECIFIC IODINATION OF MEMBRANE PROTEINS OF VIRUSES AND EUKARYOTIC CELLS USING 1,3,4,6-TETRACHLORO-3-ALPHA,6-ALPHA-DIPHENYLGLYCOLURIL
SURFACE-SPECIFIC IODINATION OF MEMBRANE PROTEINS OF VIRUSES AND EUKARYOTIC CELLS USING 1,3,4,6-TETRACHLORO-3-ALPHA,6-ALPHA-DIPHENYLGLYCOLURIL
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DOI：
10.1021/bi00615a031
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发表时间：
1978-01-01
期刊：
影响因子：
2.9
通讯作者：
FOX, CF
中科院分区：
文献类型：
--
作者：
MARKWELL, MAK;FOX, CF
碘化试剂1，3，4，6-四氯-3 α，6. α。用以下系统研究了选择性标记膜表面蛋白的二苯基甘脲（氯甘脲）：包膜病毒（仙台和纽卡斯尔病病毒）、人红细胞和在悬浮液（EL-4）和单层培养物（BHK-21）中增殖的有核细胞。描述了特异性碘化表面蛋白同时保持完全病毒完整性或细胞活力的条件。将氯甘脲法与用于标记表面膜蛋白的乳过氧化物酶和氯胺-T法进行比较，表明氯甘脲法具有许多优点：它通常产生3- 17倍的比放射性而不牺牲病毒或细胞的完整性，它在技术上更容易使用，它不需要加入外源蛋白质来引发反应，也不需要强还原剂来终止反应。氯甘脲也被证明是氯胺的有效替代品。病毒和细胞蛋白质的非载体标记中的T。膜蛋白样品在碘化之前用洗涤剂十二烷基硫酸钠溶解，或在高碘化物浓度存在下标记而无需预先溶解。使用氯甘脲产生的特定放射性等于或大于氯胺-T法产生的放射性。氯甘脲的有效性、使用简单性和多功能性推荐其作为膜系统的表面特异性和非载体标记的碘化试剂。
The use of the iodinating reagent 1,3,4,6-tetrachloro-3.alpha.,6.alpha.-diphenylglycoluril (chloroglycoluril) to selectively label membrane surface proteins was investigated with the following systems: enveloped viruses (Sendai and Newcastle disease viruses), human erythrocytes and nucleated cells propagated both in suspension (EL-4) and in monolayer culture (BHK-21). Conditions are described for specifically iodinating surface proteins while maintaining full virus integrity or cell viability. Comparison of the chloroglycoluril method with the lactoperoxidase and chloramine-T methods for labeling surface membrane proteins shows that the chloroglycoluril method has a number of advantages: it routinely produces a 3- to 17-fold greater specific radioactivity without sacrificing viral or cellular integrity, it is technically simpler to use, it does not require the addition of extraneous protein to initiate the reaction nor a strong reducing reagent to terminate it. Chloroglycoluril also proved to be an effective substitute for chloramine-T in the nonvectorial labeling of viral and cellular proteins. Membrane protein samples were solubilized with the detergent sodium dodecyl sulfate before iodination or labeled in the presence of high iodide concentrations without prior solubilization. The resulting specific radioactivities generated by the use of chloroglycoluril were equal to or greater than those generated by the chloramine-T method. The effectiveness, simplicity of use and versatility of chloroglycoluril recommend it as an iodinating reagent for both surface-specific and nonvectorial labeling of membrane systems.