Cag type IV secretion system (CagT4SS) mediates the translocation of a wide spectrum of substrates including oncoprotein CagA to host cells and plays an indispensable role in the pathogenesis of Helicobacter pylori. Central cylinder components, including CagM, CagX and CagY, are responsible for initiating the assembly of CagT4SS. Here, we performed differential HDX-MS analysis on CagX-CagY complex to map the interface upon binding. Our findings reveal the conformational changes of CagX and CagY during assembly.