Saccharomyces cerevisiae Hsp104 protein. Purification and characterization of ATP-induced structural changes.
Saccharomyces cerevisiae Hsp104 protein. Purification and characterization of ATP-induced structural changes.
复制标题
DOI：
10.1016/s0021-9258(17)41804-7
复制
发表时间：
1994-02
期刊：
影响因子：
--
通讯作者：
D. Parsell;A. Kowal;S. Lindquist
中科院分区：
文献类型：
--
作者：
D. Parsell;A. Kowal;S. Lindquist
热休克蛋白(HSP)以多种方式发挥作用，帮助细胞和生物体应对环境变化。HSP100蛋白是一类热休克蛋白，对于耐受各种极端压力条件(如高温或高浓度乙醇)尤为重要。为了开始研究Hsp100蛋白的作用机制，我们对酿酒酵母Hsp104蛋白进行了体外分析。在这里，我们报道了野生型蛋白和三个携带两个ATP结合位点共识元件突变的变异体的纯化和初步结构特征。如凝胶过滤层析和与戊二醛的交联研究所证明的那样，HSP104形成了一个高六聚体颗粒。在电子显微镜下，这些颗粒呈环状，使人想起HSP60和TF55/TCP家族中的蛋白质。与其他蛋白质不同的是，Hsp104形成单环，每个环只包含六个亚基。更引人注目的是，HSP104颗粒的组装和维持依赖于腺嘌呤核苷酸的存在。寡聚似乎主要依赖于蛋白质中两个ATP结合位点中的第二个。
Heat-shock proteins (hsps) function in a variety of ways to help cells and organisms cope with environmental changes. One class of hsps, the Hsp100 proteins, is especially important for tolerance to a variety of extremely stressful conditions (e.g. high temperatures or high concentrations of ethanol). To begin to characterize the mechanism of action of Hsp100 proteins, we have initiated an in vitro analysis of the Saccharomyces cerevisiae Hsp104 protein. Here, we report the purification and initial structural characterization of the wild-type protein and three variants carrying mutations in the two ATP-binding site consensus elements. As demonstrated by both gel filtration chromatography and by cross-linking studies with glutaraldehyde, Hsp104 forms a homohexameric particle. By electron microscopy, these particles are ring-shaped and reminiscent of proteins in the Hsp60 and TF55/TCP families. In contrast to these other proteins, Hsp104 forms single rings, each containing only six subunits. More strikingly, the assembly and maintenance of Hsp104 particles are dependent upon the presence of adenine nucleotides. Oligomerization appears to primarily depend upon the second of the two ATP-binding sites in the protein.