Secretion of Functional Antibody and Fab Fragment from Yeast Cells

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 85; no. 22; pp. 8678 - 8682
• Main Authors: Horwitz, Arnold H., Chang, C. Paul, Better, Marc, Hellstrom, Karl Erik, Robinson, Randy R.
• Format: Journal Article
• Language: English
• Published: Washington, DC National Academy of Sciences of the United States of America 01.11.1988; National Acad Sciences
• Subjects: Amino Acid Sequence; Antibodies; Antibody dependent cell cytotoxicity; Antiserum; Applied microbiology; Base Sequence; Biological and medical sciences; carcinoma; cDNA; Chimera; cloning; Cloning, Molecular; Escherichia coli - genetics; Fab; Fundamental and applied biological sciences. Psychology; Gels; Genes, Immunoglobulin; Growth, nutrition, metabolism, transports, enzymes. Molecular biology; Humans; Immunoglobulin Fab Fragments - analysis; Immunoglobulin Fab Fragments - genetics; Immunoglobulin G - analysis; Immunoglobulin G - genetics; Lymphocytes; Microbiology; Molecular Sequence Data; Mutagenesis; Mycology; Plasmids; Recombinant Proteins - analysis; Saccharomyces cerevisiae; Saccharomyces cerevisiae - genetics; Secretion; Stop codon; Vaccines, antisera, therapeutical immunoglobulins and monoclonal antibodies (general aspects); Yeasts; Antibody; Secretion; Fab-Fragment; Human; Immunoglobulins; Rodentia; Gene expression; Fungi; Vertebrata; Mammalia; Complementary DNA; Mouse; Production; Shuttle vector; Ascomycetes; Molecular cloning; Saccharomyces cerevisiae; Thallophyta; Chimera
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.85.22.8678

We have constructed yeast strains that secrete functional mouse--human chimeric antibody and its Fab fragment into the culture medium. For chimeric whole antibody, cDNA copies of the chimeric light-chain and heavy-chain genes of an anti-tumor antibody were inserted into vectors containing the yeast phosphoglycerate kinase promoter, invertase signal sequence, and phosphoglycerate kinase polyadenylylation signal. Simultaneous expression of these genes in yeast resulted in secretion of properly folded and assembled chimeric antibody that bound to target cancer cells. Yeast chimeric antibody exhibited antibody-dependent cellular cytotoxicity activity but not complement-dependent cytotoxicity activity. For production of Fab fragments, a truncated heavy-chain (Fd) gene was created by introducing a stop codon near the codon for the amino acid at which papain digestion occurs. Simultaneous expression of the resulting chimeric Fd and light-chain genes in yeast resulted in secretion of properly folded and assembled Fab fragment that bound to target cancer cells.