Recombinant HMGB1 with cytokine-stimulating activity
Recombinant HMGB1 with cytokine-stimulating activity
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DOI：
10.1016/j.jim.2004.04.019
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发表时间：
2004-06-01
影响因子：
2.2
通讯作者：
Yang, H
中科院分区：
文献类型：
--
作者：
Li, JH;Wang, HC;Yang, H
我们描述了在细菌以及哺乳动物中国仓鼠卵巢（CHO）细胞中表达的全长高迁移率族蛋白B1（HMGB1）及其截短突变体的分离、纯化和特性鉴定方法。HMGB1是一种丰富的核蛋白和胞质蛋白，在物种间高度保守，从酵母到人类的真核细胞中广泛分布。作为一种普遍存在的核DNA结合蛋白，HMGB1结合DNA，促进基因转录，并稳定核小体结构。除了这些细胞内作用外，HMGB1可由活化的先天免疫细胞（即巨噬细胞、单核细胞）释放到细胞外环境中，并作为致死性内毒素血症和败血症的介质发挥作用。HMGB1的促炎细胞因子活性已成为一个热门研究领域，重组蛋白可作为探索HMGB1功能的有用工具。由于其偶极带电特性，通过某些方法分离的HMGB1可能会被细菌产物（如CpG DNA或脂多糖[LPS]）污染，这可能会干扰免疫学分析。在此我们报道我们新开发的从细菌或哺乳动物CHO细胞中分离和纯化具有生物活性且基本无污染物的HMGB1的方法。这一策略在方法学上提供了重要进展，以促进未来对HMGB1的研究。（C）2004爱思唯尔公司。保留所有权利。
We describe methods for the isolation, purification, and characterization of full-length high-mobility group box 1 (HMGB1) and truncated mutants expressed in bacteria and in mammalian Chinese Hamster Ovary (CHO) cells. HMGB I is an abundant nuclear and cytoplasmic protein, highly conserved across species and widely distributed in eukaryotic cells from yeast to man. As a ubiquitous nuclear DNA binding protein, HMGB1 binds DNA, facilitates gene transcription, and stabilizes nucleosome structure. In addition to these intracellular roles, HMGB1 can be released into the extracellular milieu by activated innate immune cells (i.e., macrophages, monocytes) and functions as a mediator of lethal endotoxemia and sepsis. The proinflammatory cytokine activity of HMGB I has become an intense area of research and recombinant protein can be a useful tool to probe HMGB1 functions. Due to its dipolar charged properties, HMGB1 isolated by some methods can be contaminated with bacterial products (such as CpG DNA or lipopolysaccharide [LPS]) that may interfere with immunological analyses. Here we report our newly developed methods for the isolation and purification of biologically active HMGB1 from bacteria or mammalian CHO cells that is essentially free of contaminants. This strategy provides an important advance in methodology to facilitate future HMGB1 studies. (C) 2004 Elsevier B.V. All rights reserved.