嗜热自养甲烷杆菌DX01抗胁迫蛋白质的分离纯化与鉴定  

Clone and Expression Analysis of An Anti-stress Protein from Methanothermobacter marburgensis DX01

作　　者：丁霞[1] 闵航[2] 杨卫军[2] 

机构地区：[1]南昌大学生命科学与食品工程学院,江西南昌337000 [2]浙江大学生命科学学院,浙江杭州310058

出　　处：《江西农业大学学报》2010年第4期797-801,共5页Acta Agriculturae Universitatis Jiangxiensis

基　　金：江西省自然科学基金(2009GQN0091);江西省教育厅基金(GJJ10048)

摘　　要：从中国广东温泉分离得到一株嗜热自养甲烷杆菌DX01(Methanothermobacter marburgensis DX01)。分析嗜热自养甲烷杆菌DX01在不同生长温度的细胞全蛋白质图谱,分离得到一个抗胁迫相关蛋白质---甲基辅酶M还原酶I,它在70℃时上调表达,以此帮助宿主抵抗高温引发的胁迫影响。PCR克隆了甲基辅酶M还原酶I的基因全序列,系统发育分析表明甲基辅酶M还原酶I在不同的产甲烷菌株中保守性很高,M.marburgen-sis DX01与M.thermoautotrophicus和M.marburgensis Marburg的序列相似度分别是97.6%和99.2%。实时定量PCR分析表明甲基辅酶M还原酶I在不同培养温度下的MCR I mRNA表达量有显著的差异,MCR I mRNA表达量随着温度增加而增加。嗜热自养甲烷杆菌DX01菌株的抗胁迫相关蛋白质(甲基辅酶M还原酶I)的分离为研究极端古菌适应极端环境提供了新线索。Thermophilic methanogen,Methanothermobacter marburgensis DX01,was isolated from a hot spring in China.The optimal temperature for its growth is 65 ℃.In order to identify the gene potentially involved in the anti-stress protein in M.marburgensis DX01,prorein expression profiles in response to different growth temperature were investigated.The protein,up-regulated expression at 70 ℃ obviously,was cleaved out and sequenced by N-terminal amino acid sequences,which corresponded to Methyl-coenzyme M reductase I （MCR I） ,gamma subunit.Phylogenetic analysis showed that amino acid sequences of MCR I gene were conservative in the evolvement of methanogen,and consistent with the relationships that were established by 16S rRNA gene analysis on the whole.To examine the role（s） of MCR I of M.marburgensis DX01 in response to temperature,the MCR I mRNA expressions at different temperature were conducted using real-time PCR analysis.The MCR I mRNA expression increased highly at the culture temperatures 70 ℃ and was significantly lower at 50 ℃.The results suggest that MCR I in M.marburgensis DX01 is temperature-dependent transcript and conservative in the evolvement of methanogen.

关 键 词：古菌 嗜热自养甲烷杆菌 甲基辅酶M还原酶 胁迫 

分 类 号：Q936[生物学—微生物学]