Figure 3
PAR1b-binding activity of Hp_TH2099 CagA. (a) A schematic view of ABC-CagA and ABC’-CagA proteins. ABC-CagA contains the canonical EPIYA-A, EPIYA-B and EPIYA-C segments as well as canonical CM sequences. ABC’-CagA contains canonical EPIYA-A and EPIYA-B segments followed by a variant EPIYA-C segment (EPIYA-C’) and a variant CM sequence derived from Hp_TH2099 CagA. (b) AGS cells were transiently transfected with an ABC’-CagA-FLAG vector, an ABC-CagA-FLAG vector, or an empty control vector (SRα). Cell lysates were immunoprecipitated with an anti-FLAG antibody and the immunoprecipitates were subjected to immunoblotting analysis with the indicated antibodies. TCL: total cell lysate. IP: immunoprecipitate. Full-length blots are presented in Supplementary Figure S12. (c) Structural prediction of the Hp_TH2099 CagA-PAR1b interaction was carried out by using MacPyMOL v1.8.6.2 based on the crystal structure of Western CagA-PAR1b complex (PDB ID: 3IEC)38. Interface structure of the Western CagA-PAR1b complex is shown with yellow stick and grey ribbon models for CagA and PAR1b, respectively (left). The conserved Arg residue in the CM sequence of Western CagA was replaced with Gly as observed in Hp_TH2099 CagA (right). Cyan dashed lines indicate polar contacts.
