Journal
FEBS JOURNAL
Volume 277, Issue 4, Pages 973-988Publisher
WILEY
DOI: 10.1111/j.1742-4658.2009.07540.x
Keywords
LxCxE motif; natively unfolded proteins; phosphorylation; retinoblastoma protein; viral oncoprotein
Categories
Funding
- Fundacion YPF
- University of Buenos Aires
- CONICET
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DNA tumor viruses ensure genome amplification by hijacking the cellular replication machinery and forcing infected cells to enter the S phase. The retinoblastoma (Rb) protein controls the G1/S checkpoint, and is targeted by several viral oncoproteins, among these the E7 protein from human papillomaviruses (HPVs). A quantitative investigation of the interaction mechanism between the HPV16 E7 protein and the RbAB domain in solution revealed that 90% of the binding energy is determined by the LxCxE motif, with an additional binding determinant (1.0 kcal.mol(-1)) located in the C-terminal domain of E7, establishing a dual-contact mode. The stoichiometry and subnanomolar affinity of E7 indicated that it can bind RbAB as a monomer. The low-risk HPV11 E7 protein bound 2.0 kcal.mol(-1) more weakly than the high-risk HPV16 and HPV18 type counterparts, but the modularity and binding mode were conserved. Phosphorylation at a conserved casein kinase II site in the natively unfolded N-terminal domain of E7 affected the local conformation by increasing the polyproline II content and stabilizing an extended conformation, which allowed for a tighter interaction with the Rb protein. Thus, the E7-RbAB interaction involves multiple motifs within the N-terminal domain of E7 and at least two conserved interaction surfaces in RbAB. We discussed a mechanistic model of the interaction of the Rb protein with a viral target in solution, integrated with structural data and the analysis of other cellular and viral proteins, which provided information about the balance of interactions involving the Rb protein and how these determine the progression into either the normal cell cycle or transformation.
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