Biophysical characterization of the complex between human papillomavirus E6 protein and synapse-associated protein 97.
The E6 protein of human papillomavirus (HPV) exhibits complex interaction patterns with several host proteins, and their roles in HPV-mediated oncogenesis have proved challenging to study. Here we use several biophysical techniques to explore the binding of E6 to the three PDZ domains of the tumor suppressor protein synapse-associated protein 97 (SAP97). All of the potential binding sites in SAP97 bind E6 with micromolar affinity. The dissociation rate constants govern the different affinities of HPV16 and HPV18 E6 for SAP97. Unexpectedly, binding is not mutually exclusive, and all three PDZ domains can simultaneously bind E6. Intriguingly, this quaternary complex has the same apparent hydrodynamic volume as the unliganded PDZ region, suggesting that a conformational change occurs in the PDZ region upon binding, a conclusion supported by kinetic experiments. Using NMR, we discovered a new mode of interaction between E6 and PDZ: a subset of residues distal to the canonical binding pocket in the PDZ(2) domain exhibited noncanonical interactions with the E6 protein. This is consistent with a larger proportion of the protein surface defining binding specificity, as compared with that reported previously.
Department of Medical Biochemistry and Microbiology, Uppsala University, SE-75123 Uppsala, Sweden.
Celestine N
CN
Chi
Anders
A
Bach
Åke
Å
Engström
Kristian
K
Strømgaard
Patrik
P
Lundström
Neil
N
Ferguson
Per
P
Jemth
D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D004268:DNA-Binding Proteins / Q000378:metabolism*; D000074288:Discs Large Homolog 1 Protein; D006801:Humans; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D009856:Oncogene Proteins, Viral / Q000378:metabolism*; D054731:PDZ Domains; D011485:Protein Binding; D011487:Protein Conformation; D012097:Repressor Proteins / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; C513314:DLG1 protein, human; D004268:DNA-Binding Proteins; D000074288:Discs Large Homolog 1 Protein; C068651:E6 protein, Human papillomavirus type 16; C052603:E6 protein, Human papillomavirus type 18; D008565:Membrane Proteins; D009856:Oncogene Proteins, Viral; D012097:Repressor Proteins