A GENERAL PURIFICATION PROTOCOL FOR E7 PROTEINS FROM HIGH-RISK AND LOW-RISK HUMAN PAPILLOMAVIRUS TYPES EXPRESSED IN THE YEAST SCHIZOSACCHAROMYCES-POMBE
J. Braspenning et al., A GENERAL PURIFICATION PROTOCOL FOR E7 PROTEINS FROM HIGH-RISK AND LOW-RISK HUMAN PAPILLOMAVIRUS TYPES EXPRESSED IN THE YEAST SCHIZOSACCHAROMYCES-POMBE, Protein expression and purification, 10(2), 1997, pp. 192-201
A purification protocol was developed to obtain human papillomavirus (
HPV) type 16 E7 protein expressed in the yeast Schizosaccharomyces pom
be. Only three chromatographic steps were necessary to purify the unfu
sed HPV 16 E7 protein to homogeneity (95-99%) as shown by silver stain
ing after polyacrylamide gel electrophoresis. Approximately 0.8 mg of
highly purified E7 was obtained from 5 x 10(10) yeast cells. The purif
ied HPV 16 E7 phosphoprotein (Ser 31/32) was refolded and assayed for
functionality. Binding to the proteins Rbl and p107 in vitro and induc
tion of DNA synthesis after microinjection into serum-deprived NIH 3T3
cells suggest that the E7 protein retains some of its biological acti
vities. Most importantly, the purification strategy is also applicable
for different HPV 16 E7 mutants and for E7 proteins from other HPV ty
pes such as HPV 18 and 11. (C) 1997 Academic Press.