摘要
The membrane fusion protein CmeA and the outer membrane channel CmeC,two important components of CmeABC system in Campylobacter jejuni,were expressed in Escherichia coli.After Ni-NTA and ion exchange columns purification,size exclusion chromatography showed that CmeA primarily existed as trimer and CmeC existed as monomer.Then the interaction between CmeA and CmeC was analyzed by dithiobis(succinimidyl propionate)(DSP)chemical crosslinking,pull-down assay on a Ni-NTA column,and isothermal titration calorimetry(ITC)measurement.The results clearly showed the CmeA–CmeC complex band,which confirmed the interaction in vitro and this interaction is independent of substrate and CmeB.It suggests that the mechanism underlying CmeABC function in C.jejuni is similar to that of AcrABTolC in E.coli.
The membrane fusion protein CmeA and the outer membrane channel CmeC, two important compo- nents of CrneABC system in Campylobacter jejuni, were expressed in Escherichia coll. After Ni-NTA and ion exchange columns purification, size exclusion chromatography showed that CmeA primarily existed as trimer and CmeC existed as monomer. Then the interaction between CmeA and CmeC was analyzed by dithiobis （succinimidyl propionate） （DSP） chemical crosslinking, pull-down assay on a Ni-NTA column, and isothermal titration calorimetry （ITC） measurement. The results clearly showed the CmeACmeC complex band, which confirmed the interaction in vitro and this interaction is independent of substrate and CmeB. It suggests that the mechanism underlying CmeABC function in C. jejuni is similar to that of AcrAB TolC in E. coli.
基金
supported by the National Basic Research Program of China(2009CB918602)
the National Natural Science Foundation of China(30970575 and 31011120381)
Shanghai Leading Academic Discipline Project(B111)
Qi Qiwas supported by the National Talent Training Fund in Basic Research of China(J1210012)
