Growth phase-dependent expression and degradation of histones in the thermophilic archaeon Thermococcus zilligii
Growth phase-dependent expression and degradation of histones in the thermophilic archaeon Thermococcus zilligii
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DOI：
10.1046/j.1365-2958.2000.01904.x
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发表时间：
2000-05-01
影响因子：
3.6
通讯作者：
Musgrave, DR
中科院分区：
文献类型：
--
作者：
Dinger, ME;Baillie, GJ;Musgrave, DR
HTz是古细菌组蛋白家族的一员，古细菌组蛋白与真核细胞组蛋白折叠结构域具有一级序列和结构相似性，被认为是真核细胞核小体核心组蛋白的原型祖先。本文研究了生长期对Thermococcus zillivae总可溶性蛋白的影响，变性聚丙烯酰胺凝胶电泳。在进入稳定期时，至少检测到11种蛋白质的水平发生了显著变化，其中一种蛋白质经Western杂交鉴定为HTz。随着细胞进入稳定期，HTz水平急剧下降，到稳定期晚期已检测不到。出乎意料的是，Western杂交检测到第二种蛋白质，其估计分子量约为14 kDa，其抑制了HTz水平的降低。天然纯化的HTz显示在生物体的生长温度下长时间孵育后保留完全活性，表明Hit的降低是特异性细胞调节过程。通过电喷雾电离质谱法分析天然纯化的HTz显示HTz 1和HTz 2的分子量分别为7204 +/- 3 Da和7016 +/- 3 Da。检测到的唯一非共价物质对应于HTz 1-HTz 2异二聚体的分子量。北方分析表明，T. zilliptin总RNA与htz 1基因探针表明，随着生长期的进展，htz 1的表达迅速下降，并在对数生长期后期完全抑制htz 1转录物的合成。通过N-末端序列分析鉴定了三种随生长阶段水平变化的蛋白质，第一种与迄今为止测序的所有古细菌中保守的假设蛋白质同源，第二种与Sac 10 b家族的古细菌DNA结合蛋白同源，第三种与亮氨酸响应调节DNA结合蛋白家族（LRP）的C-末端区域同源。
HTz is a member of the archaeal histone family, The archaeal histones have primary sequences and structural similarity to the eukaryal histone fold domain, and are thought to resemble the archetypal ancestor of the eukaryal nucleosome core histones, The effects of growth phase on the total soluble proteins from Thermococcus zilligii, isolated after various stages of growth from mid-logarithmic to late stationary phase, were examined by denaturing polyacrylamide gel electrophoresis. On entry into stationary phase, at least 11 proteins were detected that changed considerably in level, One of these proteins was identified by Western hybridization as HTz. The level of HTz decreased dramatically as cells entered stationary phase, and it could not be detected by late stationary phase. Unexpectedly, the Western hybridization detected a second protein, with an estimated molecular mass of approximately 14 kDa, which paralleled the decrease in level of HTz, Native purified HTz was shown to retain complete activity after prolonged incubation at the growth temperature of the organism, suggesting that the decrease in Hit was a specific cell-regulated process. Analysis of native purified HTz by electrospray ionization mass spectrometry revealed the molecular masses of HTz1 and HTz2 to be 7204 +/- 3 Da and 7016 +/- 3 Da respectively, The only non-covalent species that was detected corresponded to the molecular mass of an HTz1-HTz2 heterodimer. Northern analyses of T. zilligii total RNA with an htz1 gene probe indicated a rapid decrease in expression of htz1 with progression of the growth phase, and complete repression of htz1 transcript synthesis by late logarithmic phase. Three proteins that changed in level with growth phase were identified by N-terminal sequence analysis, The first was homologous to a hypothetical protein conserved in all Archaea sequenced to date, the second to the Sac10b family of archaeal DNA-binding proteins and the third to the C-terminal region of the leucine-responsive regulatory family of DNA-binding proteins (LRPs).