Is the papillomavirus HPV 16 E7 oncoprotein a dimeric natively unfolded protein?

LEONARDO ALONSO; M. GARCIA-ALAI; G. DE PRAT GAY

Buenos Aires (Argentina)

Congreso; XIV International Biophysics Congress (IUPAB-SAB); 2002

IUPAB-SAB

The E7 oncoprotein exerts its cellular transforming activity by interfering with the tumor suppressor retinoblastoma (Rb). Several other targets have been reported related to transformation, gene transcription, DNA synthesis, and glycolytic enzymes, suggesting a broad specificity. We present the characterization of the unfolding and dissociation of HPV16 E7 dimer by spectroscopic techniques. The extended E7 dimer becomes globular at 0.4 M Gdm.Cl, undergoes a dissociation to structured momers, which finally unfold cooperatively between 2.0 and 6.0 M denaturant. Chemical crosslinking confirms the uncoupling between dissocaition and monomer unfolding. Monitoring fluorescence anisotropy change upon dilution of a fluoresceinated derivative of E7, we obtained a Kd of 1.0 x 10-6 M, in agreement with equilibrium sedimentation experiments. We had previously shown that E7 shares several properties with "natively unfolded” proteins: stability to heat denaturation; highly charged amino acid sequence, apparently low secondary structure content, and non-globular Stokes radius. However, the results of cooperative unfolding, binding of ANS, and the ability to form a stable dimer, indicate that it cannot be unfolded in solution