基于同位素标记相对和绝对定量技术研究耐铬(VI)菌株CM01的蛋白定量组学  

Quantitative proteomic in hexavalent chromium resistance CM01 by isobaric tags for relative and absolute quantitation techniques(iTRAQ)

作　　者：刘媛[1] 顾芮嘉 邱衍伦 李星龙 李迎丽[1] 高洁莹[1] 肖虹[1] LIU Yuan;GU Rui-Jia;QIU Yan-Lun;LI Xing-Long;LI Ying-Li;GAO Jie-Ying;XIAO Hong(School of Public Health and Management,Chongqing Medical University,Chongqing 400016,China;Mianyang Fucheng District Center for Disease Control and Prevention,Mianyang,Sichuan 621000,China;Chongqing Beibei District Center for Disease Control and Prevention,Chongqing 400700,China)

机构地区：[1]重庆医科大学公共卫生与管理学院,重庆400016 [2]绵阳市涪城区疾病预防控制中心,四川绵阳621000 [3]重庆市北碚区疾病预防控制中心,重庆400700

出　　处：《微生物学通报》2020年第10期3183-3195,共13页Microbiology China

基　　金：重庆市自然科学基金(cstc2019jcyj-msxmX0177)。

摘　　要：【背景】六价铬[Cr(VI)]作为一种高毒性的重金属污染物,用微生物学方法还原Cr(VI)既经济又环保。【目的】探究耐铬菌株CM01中与耐受或还原铬Cr(VI)有关的蛋白或基因,从分子水平上阐述其耐铬Cr(VI)机制。【方法】运用同位素标记相对和绝对定量技术(isobarictagsforrelativeand absolute quantitation techniques,iTRAQ)分析耐铬菌株CM01在有无铬Cr(VI)胁迫下的蛋白表达差异。运用RT-qPCR技术验证4种与代谢途径有关的上调蛋白的m RNA表达量。运用紫外分光光度计测定细胞表面疏水性。【结果】共鉴定到2570个蛋白,其中存在显著差异的蛋白数为646个,上调蛋白数量343个,下调蛋白数量303个。CM01中的Fe/Mn超氧化物歧化酶、甜菜碱醛脱氢酶、磷酸戊糖途径、磷酸肌醇代谢途径、氨基酸代谢共同参与了菌株对于外源性Cr(VI)的响应机制。RT-qPCR实验结果表明,与代谢途径有关的4种蛋白其基因转录水平和蛋白水平一致。对照组CM01的细胞表面疏水性高于实验组。【结论】初步探索了耐铬菌的Cr(VI)响应机制,为利用微生物治理环境污染提供了理论支持和新的研究思路。[Background]As a highly toxic heavy metal pollutant,the reduction of hexavalent chromium(Cr(VI))by microbiological methods is not only economical but also environmentally friendly.[Objective]To explore the proteins or genes related the tolerance or reduction of Cr(VI)in chromium-resistant strain CM01,and to explain the tolerance mechanism of the strain at the molecular level.[Methods]The difference of protein expression in chromium-resistant strain CM01 under the Cr(VI)stress or not was analyzed by isobaric tags for relative and absolute quantitation techniques(iTRAQ).The m RNA expression of 4 up-regulated proteins related to metabolic pathway was verified by RT-qPCR technique.The hydrophobicity of cell surface was determined by UV spectrophotometer.[Results]In this study,2570 proteins were identified and 646 were significantly different,of which 343 were up-regulated and 303 were down-regulated.Fe/Mn superoxide dismutase,betaine aldehyde dehydrogenase,pentose phosphate pathway,inositol phosphate pathway and amino acid metabolism in CM01 were involved in the response mechanism to exogenous Cr(VI).The results of RT-qPCR experiments showed that the gene transcription levels and protein level of the four proteins involved in metabolic pathways were consistent.The cell surface hydrophobicity of CM01 in the control group was higher than that in the experimental group.[Conclusion]Cr(VI)response mechanism of chromium-resistant bacteria was preliminarily explored in this study,which provides theoretical support and new research ideas for the use of microorganisms to control environmental pollution.

关 键 词：同位素标记相对和绝对定量技术 差异蛋白 六价铬 细菌 耐受 

分 类 号：X172[环境科学与工程—环境科学]