MAPPING THE RIGID AND FLEXIBLE REGIONS OF A NOVEL SECRETED PROTEIN EVPP FROM THE T6SS OF EDWARDSIELLA TARDA

Abstract

Many gram negative bacteria utilize secretion systems to deliver effectors into the environment or directly into their hosts. Here we characterized an important virulence protein EvpP, which is unique to the Type VI Secretion System (T6SS) of Edwardsiella tarda. We found that EvpP exists as a dimer in solution (by DLS and FPLC) and we mapped its rigid (N-terminal part) and flexible (C-ternimal part) region by NMR backbone assignment (3D and 4D), limited protease digestion as well as amide proton exchange followed by Mass Spectrometry. In addition, we examined its interaction with another secreted protein EvpC also from T6SS gene cluster. Moreover, we co-expressed and purified the translocator-chaperone complex AcrH-AopB from T3SS of Aeromonas hydrophila. We optimized the boundary for hanging-drop protein crystallization by limited protease digestion. Future work will lead to the investigation of interaction between EvpP and other T6SS members and a high quality X-ray structure of AcrH-AopB crystal complex.