THE RECOVERY OF THE HEPATITIS-B VIRUS SURFACE-ANTIGEN (HBSAG) FROM A RECOMBINANT P-PASTORIS STRAIN - DISRUPTION AND PRECIPITATION STUDIES

Cell disruption studies for the extraction of HBsAg from a recombinant P. pastoris strain (r-HBsAg) were done using a bead mill disintegrato r. Three sequential passages (4 min retention time each) were enough t o disrupt the cells and extract most of the r-HBsAg and soluble protei ns. An acid precipitation step was performed just after cell disruptio n to precipitate proteins together with the cell debris. Different pre cipitation pH values (2.5 to 6.0) were investigated. A pH value of 4.2 was selected as a compromise between recovery and improvement of spec ific activity. A 6 to 8-fold enhancement of the specific activity was obtained, having a r-HBsAg overall yield of about 80%. The influencing presence of a chaotropic salt (potassium thiocyanate) during the acid precipitation step was also studied.