A Multifunctional Single-Attachment-Point Reagent for Controlled Protein Biotinylation
A Multifunctional Single-Attachment-Point Reagent for Controlled Protein Biotinylation
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DOI：
10.1021/bc800392t
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发表时间：
2009-01-01
影响因子：
4.7
通讯作者：
Josephson, Lee
中科院分区：
文献类型：
--
作者：
Garanger, Elisabeth;Weissleder, Ralph;Josephson, Lee
生物素/亲和素系统是生物学中应用最广泛的亲和检测和捕获系统之一。然而，由于生物素不具有任何光吸收或发光特性，因此确定附着在底物上的生物素标签的确切数量是复杂的。在这里，我们描述了一个荧光生物素化试剂设计从一般多功能单附着点（MSAP）试剂的概念。Lys-Cys二肽支架用于显示生物素官能团和荧光素官能团以及n -羟基琥珀酰亚胺酯反应基团。得到的双功能MSAP试剂Fl-Biotin-NHS用于制备单生物素化版本的西妥昔单抗，并进一步与亲和素反应，得到可溶的基于亲和素的西妥昔单抗低聚物。MSAP肽支架方法允许荧光团、发色团或反应基团与生物素结合，并提供了一种广泛的方法来获得多功能生物素基试剂。
The biotin/avidin system is one of the most widely used affinity detection and affinity capture systems in biology. However, the determination of the exact number of biotin tags attached onto a substrate is complicated by the fact that biotin does not present any light-absorbing or -emitting properties. Here, we describe a fluorescent biotinylation reagent designed from the general multifunctional single-attachment-point (MSAP) reagent concept. A Lys-Cys dipeptide scaffold was used to display a biotin functional group and a fluorescein functional group along with an N-hydroxysuccinimide ester reactive group. The resulting bifunctional MSAP reagent, Fl-Biotin-NHS, was used to prepare a monobiotinylated version of cetuximab, which was further reacted with avidin to obtain a soluble avidin-based cetuximab oligomer. The MSAP peptide-scaffold approach allows fluorophores, chromophores, or reactive groups to be combined with biotin and provides a broad approach to obtain multifunctional biotin-based reagents.