High-level secretory expression of immunologically active intact antibody from the yeast Pichia pastoris

We have produced a functional murine antibody to dioxin in the culture medi um of the methylotrophic yeast Pichia pastoris. Complementary DNA copies en coding the light (kappa) and heavy (gamma) chains of the dioxin monoclonal antibody, DD1, were each placed under the control of P. pastoris alcohol ox idase (AOX1) promoter and Saccharomyces cerevisiae alpha-mating factor secr etion signal sequence. The resulting expression cassettes were assembled in to a single plasmid (pPICZ alpha DD1) to permit co-expression of both light and heavy chains of the antibody molecule. P. pastoris SMD1168 (pep4, his4 ) transformed with pPICZ alpha DD1 was able to secrete intact antibody into the culture medium. As high as 36 mg l(-1) of the antibody was produced in shake-flask cultures after 96-h induction with methanol. Functional analys is using immunoassay confirmed murine nature of the recombinant antibody an d its ability to bind dioxin.