Figure 1
Expression and purification of the fusion protein LALF 32-51 -E7. (A) SDS-PAGE (15%). (B) Western Blot using anti-HPV-16 E7 monoclonal antibody: initial crude extract of E. coli proteins containing the expressed (about 18%) fusion protein LALF32-51-E7 (lane 1); soluble fraction after cell disruption (lane 2); insoluble fraction after cell disruption (lane 3); soluble fraction after treatment with 6 M urea (lane 4); insoluble fraction after treatment with 6 M urea (lane 5); IMAC purification, initial sample (lane 6); IMAC purification, wash with 10 mM imidazole (lane 7); IMAC purification, elution with 300 mM imidazole (lane 8); 300 mM imidazole elution fraction after desalting (lane 9). Molecular mass markers are indicated on the left. (C) Analytical gel filtration chromatogram of purified LALF32-51-E7 (continuous line). Dash line is representative of the retention time for (from left to right) bovine thyroglobulin (670 kDa), bovine gamma globulin (158 kDa), ovalbumin (44 kDa), myoglobin (17 kDa) and vitamin B-12 (1.35 kDa).