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Functional Reconstitution of Staphylococcus aureus Truncated AgrC Histidine Kinase in a Model Membrane System
Wang, Lina1; Quan, Chunshan2,3; Liu, Baoquan2,3; Wang, Jianfeng2,3; Xiong, Wen2,3; Zhao, Pengchao1; Fan, Shengdi2,3
刊名PLOS ONE
2013-11-26
DOI10.1371/journal.pone.0080400
8期:11
收录类别SCI
文章类型Article
WOS标题词Science & Technology
类目[WOS]Multidisciplinary Sciences
研究领域[WOS]Science & Technology - Other Topics
关键词[WOS]SIZE-EXCLUSION CHROMATOGRAPHY ; ESCHERICHIA-COLI ; 2-DIMENSIONAL CRYSTALLIZATION ; PROTEIN RECONSTITUTION ; AUTOINDUCING PEPTIDES ; RECEPTOR ; SENSOR ; DETERGENTS ; LIPOSOMES ; VIRULENCE
英文摘要The integral membrane protein AgrC is a histidine kinase whose sensor domains interact with an autoinducing peptide, resulting in a series of downstream responses. In this study, truncated AgrC(TM5-6C) and AgrC(TM5-6C)-GFP with GFP as a reporter gene were produced using a bacterial system. Purified AgrC(TM5-6C) and AgrC(TM5-6C)-GFP were reconstituted into liposomes by a detergent-mediated method. To achieve high-yield protein incorporation, we investigated the effect of different detergents on protein reconstitution efficiency. The highest incorporation was found with N, N-dimethyldode-cylamine Noxide during complete liposome solubilization, which resulted in a yield of 85 +/- 5%. The COOH-terminus of the protein AgrC(TM5-6C) was almost exclusively oriented towards the inside of the vesicles. AgrC(TM5-6C) in proteoliposomes exhibited approximately a 6-fold increase in constitutive activity compared with AgrC(TM5-6C) in detergent micelles. The reconstitution of AgrC(TM5-6C) or AgrC(TM5-6C)-GFP was characterized using dynamic light scattering, fluorescence microscopy, and transmission electron microscopy. Based on the results, the optimal conditions for protein incorporation were defined. These findings contribute to the study of membrane protein structure and function in vitro using a reconstitution system.
语种英语
WOS记录号WOS:000327546400027
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文献类型期刊论文
条目标识符http://cas-ir.dicp.ac.cn/handle/321008/137718
专题中国科学院大连化学物理研究所
作者单位1.Chinese Acad Sci, Dalian Inst Chem Phys, Dalian, Peoples R China
2.Dalian Nationalities Univ, Dept Life Sci, Dalian, Peoples R China
3.State Ethn Affairs Commiss, Minist Educ, Dalian, Peoples R China
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Wang, Lina,Quan, Chunshan,Liu, Baoquan,et al. Functional Reconstitution of Staphylococcus aureus Truncated AgrC Histidine Kinase in a Model Membrane System[J]. PLOS ONE,2013,8(11).
APA Wang, Lina.,Quan, Chunshan.,Liu, Baoquan.,Wang, Jianfeng.,Xiong, Wen.,...&Fan, Shengdi.(2013).Functional Reconstitution of Staphylococcus aureus Truncated AgrC Histidine Kinase in a Model Membrane System.PLOS ONE,8(11).
MLA Wang, Lina,et al."Functional Reconstitution of Staphylococcus aureus Truncated AgrC Histidine Kinase in a Model Membrane System".PLOS ONE 8.11(2013).
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