Cysteine-specific radioiodination of proteins with fluorescein maleimide

被引：9
作者：
Palmer, M
Buchkremer, M
Valeva, A
Bhakdi, S
机构：
[1] Institute of Medical Microbiology, University of Mainz, Augustusplatz
来源：
ANALYTICAL BIOCHEMISTRY | 1997年 / 253卷 / 02期
关键词：
cysteine; sulfhydryl group; fluorescein; maleimides; I-125; radioactive labeling;
D O I：
10.1006/abio.1997.2364
中图分类号：
Q5 [生物化学];
学科分类号：
071010 ; 081704 ;
摘要：
A protocol is described for coupling of carrier-free iodine to protein sulfhydryl groups via fluorescein maleimide. I-125 is first coupled to fluorescein maleimide in the presence of chloramine T. Iodination is stopped with sodium thiosulfate, and the iodine-substituted fluorescein maleimide is reacted with free cysteines of the protein. Excess label is then removed by gel-permeation chromatography. The procedure avoids exposition of the protein to oxidative conditions and does not require purification of the labeled carrier reagent. Suitability of the method for a given protein can be evaluated spectrophotometrically without employing radioactivity. It can be applied under denaturing conditions and may be particularly useful with mutant proteins carrying engineered single cysteine residues at sites that are not functionally critical. (C) 1997 Academic Press.
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收藏
页码：175 / 179
页数：5
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