Members of the PDZ domain family of proteins play a key role in development and transmission of signals between cells. Viruses will therefore target these proteins in order to hijack their host cells as a means to enhance replication and transmit to new hosts. NMR spectroscopy provides a powerful technique to analyze the atomic-level interaction between PDZ domains and their intrinsically disordered viral ligands in solution. We are specifically interested in the interaction between the PDZ-1 domain from Zonula Occludens Protein 1 (ZO-1) and the major regulatory nonstructured protein 5 (NS5) from both tick borne encephalitis virus (TBEV) and Dengue virus (DENV). In order to investigate these interactions at the atomic level by NMR spectroscopy, the necessary first step is to fully assign the 1H-15N HSQC spectrum for isotopically 15N-labeled PDZ-1 from ZO-1. This HSQC spectrum acts as a footprint for the protein and once each resonance has been assigned to the appropriate amino acid in the protein backbone, it will be possible to monitor changes to the chemical shift position of these resonances upon addition of viral ligand. This will ultimately enable the binding interface to be mapped to the PDZ-1 domain of ZO-1. This project therefore focuses on the full assignment of the 1H-15N HSQC, utilizing uniformly isotopically labeled 15N/13C protein and a battery of 3-dimensional NMR experiments.

Analysis and assignment of the 1H-15N HSQC of the PDZ-1 domain from zonula occludens protein 1

253rd National Meeting (2017)

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