Oligomerization mechanism of the HPV 16 oncoprotein E7

C. SMAL; WETZLER DE; ALONSO LEONARDO G.; DE PRAT-GAY GONZALO

Salta

Congreso; XXXIX Annual Meeting of the Argentinean Biophysical Society (SAB 2010); 2010

 @font-face { font-family: "Times New Roman"; }p.MsoNormal, li.MsoNormal, div.MsoNormal { margin: 0cm 0cm 0.0001pt; font-size: 10pt; font-family: "Times New Roman"; }p.MsoBodyTextIndent, li.MsoBodyTextIndent, div.MsoBodyTextIndent { margin: 0cm 0cm 0.0001pt; text-align: justify; text-indent: 14.2pt; font-size: 10pt; font-family: "Times New Roman"; }table.MsoNormalTable { font-size: 10pt; font-family: "Times New Roman"; }div.Section1 { page: Section1; } The oncoprotein E7 is the major transforming activity in human papillomaviruses, a causal agent for cervical cancer. More than 40 specific cellular targets were described for this protein that represents a prototypical oncoprotein among small DNA tumor viruses. The HPV16 E7 oncoprotein is an extended dimer, with a stable and cooperative fold, but it displays properties of intrinsically disorder proteins. The protein can form spherical oligomers with amyloid-like properties that display chaperone activity. It was previously shown that oligomers assemble through an ordered process. The oligomers assemble can be started in vitro by chelating the structural Zn by EDTA 1,2. In this work we characterize the kinetic mechanism of oligomers assembly using different experimental techniques such as circular dichroism and fluorescence spectroscopy, dynamic light scattering measurements and binding to specific dyes that report different conformational and structural events. We could observe three phases in the oligomerization process: a slow lag phase, a rapid elongation step and a final steady state. A two state process with an initial monomeric specie and a final homogeneous spherical oligomer without populated intermediaries was found. Using a kinetic model proposed by Zlotnick et al.3 for spherical oligomeric assemblies we propose a simple mechanism where the formation of E7 monomer species after chelating Zn corresponds to lag phase observed, followed by the elongation phase where monomeric E7 species are added one by one to form the final state.