Tanaka K, Yoshimura T, Kumatori A, Ichihara A, Ikai A, Nishigai M, Kameyama K, Takagi T
Institute for Enzyme Research, University of Tokushima, Japan.
J Biol Chem. 1988 Nov 5;263(31):16209-17.
Latent multicatalytic protease complexes, named proteasomes, were purified to apparent homogeneity from various eukaryotic sources, such as human, rat, and chicken liver, Xenopus laevis ovary, and yeast (Saccharomyces cerevisiae), and their functional and structural properties were compared. They showed latency in breakdown of [methyl-3H]casein, but were greatly activated in various ways, such as by addition of polylysine. They all degraded three types of fluorogenic oligopeptides at the carboxyl side of basic, neutral, and acidic amino acids, and the three cleavage reactions showed different spectra for inhibition, suggesting that they had three distinct active sites. The proteasomes all seemed to be seryl endopeptidases with similar pH optima in the weakly alkaline region. Their physiochemical properties, such as their sedimentation coefficients (19 S to 22 S), diffusion coefficients (2.0-2.6 X 10(-7) cm2 s-1), molecular masses (700-900 kDa), and circular dichroic spectra, were similar. Their amino acid compositions were also very similar. Electron microscopy showed that they had similar well-defined symmetrical morphology, appearing to be ring-shaped particles with a small hole in the center. All the proteasomes seemed to be multisubunit complexes consisting of 15-20 polypeptides with molecular masses of 22-33 kDa and isoelectric points of pH 3-10, but they showed species-specific differences in subunit multiplicity. Moreover, they differed immunologically, as shown by Ouchterlony tests and immunoblotting analyses, although cross-immunoreactivities of some subunits or domains were observed. These results indicate that the sizes and shapes of these proteasomes have been highly conserved during evolution, but that they show species-specific differences in immunoreactivities and subunit structures. Thus proteasomes with similar structure and function seem to be ubiquitously distributed in eukaryotic organisms ranging from man to yeast. This distribution implies the general importance of these proteasomes for proteolysis.
从人、大鼠和鸡的肝脏、非洲爪蟾卵巢以及酵母（酿酒酵母）等多种真核生物来源中纯化出了潜在的多催化蛋白酶复合物，即蛋白酶体，并使其达到了表观均一性，同时对它们的功能和结构特性进行了比较。它们在分解[甲基-3H]酪蛋白时表现出潜伏性，但通过多种方式可被极大激活，比如添加多聚赖氨酸。它们都能在碱性、中性和酸性氨基酸的羧基侧降解三种类型的荧光寡肽，并且这三种切割反应呈现出不同的抑制谱，这表明它们具有三个不同的活性位点。这些蛋白酶体似乎都是丝氨酸内肽酶，在弱碱性区域具有相似的最适pH值。它们的物理化学性质，如沉降系数（19 S至22 S）、扩散系数（2.0 - 2.6×10⁻⁷ cm² s⁻¹）、分子量（700 - 900 kDa）以及圆二色光谱，都很相似。它们的氨基酸组成也非常相似。电子显微镜显示它们具有相似的明确对称形态，看起来是中心有小孔的环形颗粒。所有的蛋白酶体似乎都是由15 - 20种分子量为22 - 33 kDa且等电点为pH 3 - 10的多肽组成的多亚基复合物，但它们在亚基多样性上表现出物种特异性差异。此外，如免疫双扩散试验和免疫印迹分析所示，它们在免疫学上存在差异，尽管观察到了一些亚基或结构域的交叉免疫反应性。这些结果表明，这些蛋白酶体的大小和形状在进化过程中高度保守，但它们在免疫反应性和亚基结构上表现出物种特异性差异。因此，具有相似结构和功能的蛋白酶体似乎普遍存在于从人类到酵母的真核生物中。这种分布意味着这些蛋白酶体在蛋白水解中具有普遍重要性。
