相较于水母绿色荧光蛋白（Aequorea EGFP），盘基网柄菌红色荧光蛋白（Discosoma DsRed）具有更高的稳定性。

High stability of Discosoma DsRed as compared to Aequorea EGFP.

作者信息

Verkhusha Vladislav V, Kuznetsova Irina M, Stepanenko Olesia V, Zaraisky Andrey G, Shavlovsky Michail M, Turoverov Konstantin K, Uversky Vladimir N

机构信息

Department of Pharmacology, University of Colorado Health Sciences Center, Denver, Colorado 80262, USA.

出版信息

Biochemistry. 2003 Jul 8;42(26):7879-84. doi: 10.1021/bi034555t.

DOI:10.1021/bi034555t

PMID:12834339

Abstract

Comparative analysis of conformational stabilities was performed for two widely used genetic reporters, EGFP and DsRed, proteins exhibiting similar beta-can folds, but possessing different oligomeric organization and chromophore structures. Two factors affecting protein stability in vitro, such as elevated temperatures and a chaotropic agent guanidine hydrochloride, were studied. In vivo tolerance of the fluorescence proteins to proteasomal-based degradation was studied in insect and mammalian cells, and in Xenopus embryos. The apparent rate constants of thermal and GdmCl-induced denaturation were several orders of magnitude lower for DsRed than for EGFP. DsRed lifetimes severalfold longer than those of EGFP were observed in cultured cells and in embryos. The remarkable fluorescence stability of DsRed under the all conditions that have been studied is attributed to a significant extent to its tetrameric organization. Therefore, DsRed can be used as a genetic reporter and advanced population marker with a significantly extended intracellular lifespan.

摘要

对两种广泛使用的基因报告蛋白EGFP和DsRed进行了构象稳定性的比较分析，这两种蛋白具有相似的β桶折叠结构，但寡聚体组织和发色团结构不同。研究了影响蛋白质体外稳定性的两个因素，如升高温度和添加离液剂盐酸胍。在昆虫细胞、哺乳动物细胞和非洲爪蟾胚胎中研究了荧光蛋白对基于蛋白酶体的降解的体内耐受性。DsRed的热变性和盐酸胍诱导变性的表观速率常数比EGFP低几个数量级。在培养细胞和胚胎中观察到DsRed的寿命比EGFP长几倍。在所研究的所有条件下，DsRed显著的荧光稳定性在很大程度上归因于其四级结构。因此，DsRed可作为一种基因报告蛋白和先进的群体标记物，其细胞内寿命显著延长。