| Title | Screening and Identification of ClpE Interaction Proteins in Streptococcus pneumoniae by a Bacterial Two-Hybrid System and Co-immunoprecipitation |
|---|---|
| Author | WenJuan Yan1, YingYing Cai1, Qun Zhang2, YuSi Liu1, WenChun Xu1, YiBing Yin1, YuJuan He1, Hong Wang1, and XueMei Zhang1* |
| Address | 1Key Laboratory of Diagnostic Medicine Designated by the Ministry of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing 400016, P. R. China, 2Clinical Laboratories Center, Affiliated Children's Hospital, Chongqing Medical University, Chongqing 400016, P. R. China |
| Bibliography | Journal of Microbiology, 51(4),453-460, 2013, |
| DOI | 10.1007/s12275-013-3001-4 |
| Key Words | Streptococcus pneumoniae, ClpE, interaction proteins, FtsW, bacterial two-hybrid, co-immunoprecipitation |
| Abstract | Hsp100/Clp proteins have crucial functions in the protein quality control, stress tolerance, and virulence of many pathogenic bacteria. ClpE is an important virulence factor involved in adherence and invasion in Streptococcus pneumoniae. To explore the underlying mechanism, we screened ClpE interaction proteins using a bacterial two-hybrid system and co-immunoprecipitation. We used ClpE as bait and constructed the pBT-ClpE bait plasmid for two-hybrid screening. Then, we constructed ClpE::GFP fusion for co-immunoprecipitation screening using anti-GFP monoclonal antibody. We obtained eight potential ClpE interaction proteins, including carbamoyl-phosphate synthase, pyruvate oxidase (SpxB), phosphoenolpyruvate-protein phosphotransferase, aminopeptidase N (pepN), L-lactate dehydrogenase, ribosomal protein S4, sensor histidine kinase (SPD_2019), and FtsW (a cell division protein). FtsW, SpxB, pepN, and SPD_2019 were confirmed to interact with ClpE using Bacterial Two-hybrid or Co-immunoprecipitation. Morphologic observations found that ΔclpE strain existed in abnormal division. β-Galactosidase activity assay suggested that ClpE contributed to the degradation of FtsW. Furthermore, FtsW could be induced by heat shock. The results suggested that ClpE might affect cell division by regulating the level of FtsW. These data may provide new insights in studying the role of ClpE in S. pneumoniae. |