@article{mbs:/content/journal/micro/10.1099/00221287-148-1-41, author = "Swennen, Dominique and Paul, Marie-Françoise and Vernis, Laurence and Beckerich, Jean-Marie and Fournier, Alain and Gaillardin, Claude", title = "Secretion of active anti-Ras single-chain Fv antibody by the yeasts Yarrowia lipolytica and Kluyveromyces lactis", journal= "Microbiology", year = "2002", volume = "148", number = "1", pages = "41-50", doi = "https://doi.org/10.1099/00221287-148-1-41", url = "https://www.microbiologyresearch.org/content/journal/micro/10.1099/00221287-148-1-41", publisher = "Microbiology Society", issn = "1465-2080", type = "Journal Article", keywords = "scFv, single-chain antibody fragment(s)", keywords = "heterologous secretion", keywords = "glucoamylase", keywords = "GST, glutathione S-transferase", keywords = "AEP, Yarrowia lipolytica alkaline extracellular protease", keywords = "Kex2p, Kex2 protease (Kexin, EC 3 . 4 . 21 . 61)", abstract = " Yarrowia lipolytica and Kluyveromyces lactis secretion vectors were constructed and assessed for the expression of heterologous proteins. An anti-Ras single-chain antibody fragment (scFv) coding sequence was fused in-frame to different pre- or prepro-regions, or downstream from a reporter secretory gene (Arxula adeninivorans glucoamylase), separated by a Kex2 protease (Kex2p)-like processing sequence. Both organisms are able to secrete soluble scFv, with yields depending on the nature of the expression cassette, up to levels ranging from 10 to 20 mg l−1. N-terminal sequence analysis of the purified scFv showed that fusions are correctly processed to the mature scFv by a signal peptidase or a Kex2p-type endoprotease present in Y. lipolytica and K. lactis. The scFv protein also retains the capacity to bind to a glutathioneS-transferase (GST)–Harvey-RasVal12 fusion, indicating that the antibody is functional. These results indicate that the yeasts Y. lipolytica and K. lactis have potential for industrial production of soluble and active scFv.", }