Expression of the Kappa (K) Light Chain of Mouse Anticreatine Kinase-M (MAK33) Antibody in the Yeast Hansenula Polymorpha

An expression plasmid was constructed and transformed to the methylotrophic yeast Hansenulapolyinorpha to produce the kappa chain peptide of IgG antibody (MAK33). Hansenula polymorpha appears to be one of the most efficient host cells for the expression of genetically engineered antibody genes. Although the rate of transformation of this yeast by polyethylene glycol was low (3-5 cells/1.0μg plasmid DNA), yet the transformants showed high mitotic stability for more than 100 generations. The expression plasmid was integrated within the yeast genome at one or more integrations site(s) with low and multicopy-number plasmids via a non-homologous integration mechanism. An N-terminal glucoamylase gene fragment was linked to the light chain (kappa) gene of the Fab derivative of the MAK33 antibody. Prepro-alpha factor of the yeast Saceharomyces cerevisiae was inserted into the plasmid between the glucoamylase and light chain genes as a secretion signal sequence for the light chain peptide. The kappa chain was produced at 50mg/L free protein in the culture medium and 500mg/L entrapped within the cells, amounting to 550mg/L and representing about 10% of the total cell protein. The prepro-a factor was shown to be incompletely processed in Hansenulapolyinorpha and the pro-segment accompanied the light chain peptide.