identifying-protein-protein-interaction-sites-using-peptide-arrays

Identifying Protein-protein Interaction Sites Using Peptide Arrays

Protein-protein interactions mediate most of the processes in the living cell and control homeostasis of the organism. Impaired protein interactions may ...

The Hebrew University of Jerusalem

CagA is a virulence factor that Helicobacter pylori inject into gastric epithelial cells through a type IV secretion system where it can cause gastric adenocarcinoma. Translocation is dependent on the presence of secretion signals found in both the N- and C-terminal domains of CagA and an interaction with the accessory protein CagF. However, the molecular basis of this essential protein-protein interaction is not fully understood. Herein we report, using isothermal titration calorimetry, that CagA forms a 1:1 complex with a monomer of CagF with nM affinity. Peptide arrays and isothermal titration calorimetry both show that CagF binds to all five domains of CagA, each with μM affinity. More specifically, a coiled coil domain and a C-terminal helix within CagF contacts domains II-III and domain IV of CagA, respectively. In vivo complementation assays of H. pylori with a double mutant, L36A/I39A, in the coiled coil region of CagF showed a severe weakening of the CagA-CagF interaction to such an extent that it was nearly undetectable. However, it had no apparent effect on CagA translocation. Deletion of the C-terminal helix of CagF also weakened the interaction with CagA but likewise had no effect on translocation. These results indicate that the CagA-CagF interface is distributed broadly across the molecular surfaces of these two proteins to provide maximal protection of the highly labile effector protein CagA.

Anat Iosub-Amir

Institute of Chemistry

The Hebrew University of Jerusalem

Identifying Protein-protein Interaction Sites Using Peptide Arrays

Anat Iosub-Amir

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Institute of Chemistry

The Hebrew University of Jerusalem

Identifying Protein-protein Interaction Sites Using Peptide Arrays

Institute of Chemistry