Targeting mechanism of the retinoblastoma tumor suppressor by a prototypical viral oncoprotein. Structural modularity, intrinsic disorder and phosphorylation of human papillomavirus E7.
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.
Chemes
Lucía B
LB
Protein Structure-Function and Engineering Laboratory, Fundación Instituto Leloir and IIBBA-CONICET, Buenos Aires, Argentina.
Sánchez
Ignacio E
IE
Smal
Clara
C
de Prat-Gay
Gonzalo
G
D000595:Amino Acid Sequence; D047390:Casein Kinase II / Q000235:genetics; D017124:Conserved Sequence; D052162:Human papillomavirus 16; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009856:Oncogene Proteins, Viral / Q000235:genetics / Q000378:metabolism*; D050725:Papillomavirus E7 Proteins; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D017434:Protein Structure, Tertiary; D016160:Retinoblastoma Protein / Q000235:genetics / Q000378:metabolism*
D009856:Oncogene Proteins, Viral; D050725:Papillomavirus E7 Proteins; D010446:Peptide Fragments; D016160:Retinoblastoma Protein; C059731:oncogene protein E7, Human papillomavirus type 16; D047390:Casein Kinase II