鉴定血红素结合蛋白及其相互作用定量的方法。

Method for identification of heme-binding proteins and quantification of their interactions.

机构信息

Centre de Recherche des Cordeliers, INSERM, Sorbonne Université, USPC, Université Paris Descartes, Université Paris Diderot, F-75006, Paris, France.

出版信息

Anal Biochem. 2020 Oct 15;607:113865. doi: 10.1016/j.ab.2020.113865. Epub 2020 Jul 29.

DOI:10.1016/j.ab.2020.113865

PMID:32738209

Abstract

The standard assay for characterization of interaction of heme with proteins is absorbance spectroscopy. However, this approach demands relatively large quantities of proteins and it is difficult to perform in high-throughput manner. Here, we describe an immunosorbent assay based on the covalent in situ conjugation of heme to a pre-coated carrier. Advantage of this assay is that it allows both identification of heme-binding proteins and quantification of their binding avidity, using only minimal amounts of protein (1-10 μg). Importantly, the same approach can be used for covalent linkage of other natural or synthetic compounds and analyzing their interactions with proteins.

摘要

用于研究血红素与蛋白质相互作用的标准分析方法是吸收光谱法。然而，这种方法需要相对大量的蛋白质，并且很难进行高通量分析。在这里，我们描述了一种基于血红素与预先包被载体的共价原位结合的免疫吸附测定法。这种测定法的优点是，仅使用少量蛋白质（1-10μg），即可鉴定血红素结合蛋白并定量其结合亲和力。重要的是，该方法还可以用于其他天然或合成化合物的共价连接，并分析它们与蛋白质的相互作用。