克雷伯氏菌Z3还原Cr(Ⅵ)的细胞定位及相关功能蛋白分析  

Cellular localization and related functional protein analysis of Cr (Ⅵ) reduced by Klebsiella Z3

作　　者：曹子敬 张智超 高明 杜文涛[1] 张继冉[1] 张世敏[1] 吴坤[1] 徐淑霞[1] CAO Zijing;ZHANG Zhichao;GAO Ming;DU Wentao;ZHANG Jiran;ZHANG Shimin;WU Kun;XU Shuxia(College of Life Sciences,Henan Agricultural University,Zhengzhou 450002,China)

机构地区：[1]河南农业大学生命科学学院,河南郑州450002

出　　处：《河南农业大学学报》2021年第4期736-743,共8页Journal of Henan Agricultural University

基　　金：国家自然科学基金项目(31801672);河南省高等学校重点科研项目计划支持项目(19A180017)。

摘　　要：为了探究微生物还原Cr(Ⅵ)的细胞定位以及还原过程中相关蛋白的功能,以嗜碱性Cr(Ⅵ)高效还原细菌克雷伯氏菌(Klebsiella sp.)Z3为研究对象,结合扫描电镜(scanning electron microscope,SEM)、X射线光电子能谱分析(X-ray photoelectron spectroscopy,XPS)、能量色散X射线谱(energy dispersive spectroscopy,EDS)以及双向电泳联合质谱,考察了铬(Cr)在菌株Z3内的分布特征、细胞各组分对Cr(Ⅵ)的还原能力以及Cr胁迫条件下菌株Z3的全细胞蛋白表达差异。结果表明,100 mg·L^(-1) Cr(Ⅵ)处理组,菌体胞外多聚物、细胞膜、细胞壁和细胞质的总Cr质量浓度分别为8.00、1.49、1.84和3.05 mg·L^(-1)。细胞各组分的还原能力由高到低依次为胞外多聚物>细胞质>细胞膜>细胞壁。SEM结果表明,菌体还原Cr(Ⅵ)的主要位置为胞外多聚物。还原过程及还原结束菌体细胞的EDS检测结果进一步证明,Cr(Ⅵ)的还原过程发生在胞外。XPS分析表明,Cr(Ⅵ)被还原为Cr(Ⅲ),产生的蓝灰色沉淀为Cr(OH)_(3)。双向电泳联合质谱对相关功能蛋白的鉴定结果表明,Cr(Ⅵ)胁迫条件下,菌株Z3有23个蛋白的表达量上调1.5倍以上,功能上分别聚类于蛋白质合成、能量代谢、氨基酸代谢、胁迫应答、免疫应答、信号传导和脂肪酸代谢共7大类。In order to explore the cellular localization of Cr(Ⅵ)reduction by microorganisms and the function of related proteins in the process of reduction,Klebsiella sp.Z3,an alkaline Cr(Ⅵ)reducing bacterium,was used as the research object.The cellular distribution of Cr in strain Z3,the reduction ability of each component to Cr(Ⅵ)and the whole cell protein expression of strain Z3 under Cr stress were investigated by scanning electron microscope(SEM),X-ray photoelectron spectroscopy(XPS),energy dispersive spectroscopy(EDS)and two-dimensional electrophoresis combined with mass spectrometry.The results showed that the total Cr mass concentration of extracellular polymer,cell membrane,cell wall and cytoplasm in 100 mg·L^(-1) Cr(Ⅵ)treatment group was 8.00,1.49,1.84 and 3.05 mg·L^(-1),respectively.The reduction ability of cell components from high to low was as follows:extracellular polymer>cytoplasm>cell membrane>cell wall.SEM results showed that the main position of Cr(Ⅵ)reduction was extracellular polymer.The results of EDS test of the reduction process and the end of reduction further proved that Cr(Ⅵ)reduction occurred outside the cell.XPS analysis showed that Cr(Ⅵ)was reduced to Cr(Ⅲ),and the resulting blue gray precipitation was Cr(OH)_(3).The results of the identification of related functional proteins by twodimensional electrophoresis and mass spectrometry showed that under Cr(Ⅵ)stress,the expression of 23 proteins in strain Z3 was increased by more than 1.5 times,and the functional clustering was divided into seven categories:protein synthesis,energy metabolism,amino acid metabolism,stress response,immune response,signal conduction and fatty acid metabolism.

关 键 词：克雷伯氏菌 铬 三价铬 六价铬 还原机制 细胞定位 功能蛋白 

分 类 号：S182[农业科学—农业基础科学] Q932[生物学—微生物学]