利用荧光光谱法测定谷胱甘肽与蛋白质的结合。

Determination of glutathione-binding to proteins by fluorescence spectroscopy.

机构信息

Department of Molecular Medicine, Chonnam National University, Graduate School, Gwangju, South Korea.

Department of Medical Science, Chonnam National University, Graduate School, Gwangju, South Korea.

出版信息

Biochem Biophys Res Commun. 2021 Jun 11;557:329-333. doi: 10.1016/j.bbrc.2021.04.016. Epub 2021 Apr 22.

DOI:10.1016/j.bbrc.2021.04.016

PMID:33895474

Abstract

Glutathione (GSH) is the most abundant non-protein thiol and its cellular concentration has been reported as 17 mM in Escherichia coli. This study introduces a label-free method to determine the binding affinity of GSH to proteins, utilizing the intrinsic fluorescence of proteins; the dissociation constants of GSH for d-arabinose 5-phosphate isomerase KdsD, fumarase C, malate dehydrogenase, and RNA polymerase subunit α have been determined as 96 ± 8, 246 ± 42, 292 ± 78, and 296 ± 97 μM, respectively. The dissociation constants, less than 2% of the cellular concentration of GSH, suggests that protein-GSH interactions are strong enough to make all of the GSH-binding sites occupied fully. The method described here may be applicable to other proteins.

摘要

谷胱甘肽（GSH）是最丰富的非蛋白巯基，其细胞浓度在大肠杆菌中被报道为 17mM。本研究介绍了一种利用蛋白质固有荧光的无标记方法来测定 GSH 与蛋白质的结合亲和力，已分别确定 GSH 与 D-阿拉伯糖 5-磷酸异构酶 KdsD、延胡索酸酶 C、苹果酸脱氢酶和 RNA 聚合酶亚基 α 的解离常数为 96±8、246±42、292±78 和 296±97μM。这些解离常数小于 GSH 细胞浓度的 2%，表明蛋白-GSH 相互作用足够强，足以使所有 GSH 结合位点完全被占据。此处描述的方法可能适用于其他蛋白质。