High‐level production and secretion of recombinant proteins by the dimorphic yeast Arxula adeninivorans

• Published in: FEMS yeast research Vol. 2; no. 3; pp. 363 - 369
• Main Authors: Wartmann, Thomas, Böer, Erik, Pico, Almudena Huarto, Sieber, Heike, Bartelsen, Oliver, Gellissen, Gerd, Kunze, Gotthard
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Publishing Ltd 01.08.2002
• Subjects: Arxula adeninivorans; Blotting, Southern; Dimorphism; Fermentation; Genetic Markers; Genetic Vectors; Green Fluorescent Proteins; Heterologous gene expression; Hygromycin B - pharmacology; Luminescent Proteins - biosynthesis; Luminescent Proteins - genetics; Recombinant Proteins - biosynthesis; Saccharomycetales - genetics; Saccharomycetales - metabolism; Secretion; Serum Albumin - biosynthesis; Serum Albumin - genetics; Temperature; Transformation; Transformation, Genetic
• ISSN: 1567-1356; 1567-1364
• DOI: 10.1111/j.1567-1364.2002.tb00105.x

The non‐conventional dimorphic thermo‐ and salt‐resistant yeast Arxula adeninivorans has been developed as a host for heterologous gene expression. For assessment of the system two model genes have been selected: the GFP gene encoding the intracellular green fluorescent protein, and the HSA gene encoding the secreted human serum albumin. The expression system includes two host strains, namely A. adeninivorans LS3, which forms budding cells at 30°C and mycelia at >42°C, and the strain A. adeninivorans 135, which forms mycelia at temperatures as low as 30°C. For expression control the constitutive A. adeninivorans‐derived TEF1‐promoter and S. cerevisiae‐derived PHO5‐terminator were selected. The basic A. adeninivorans transformation/expression vector pAL‐HPH1 is further equipped with the Escherichia coli‐derived hph gene conferring hygromycin B resistance and the 25S rDNA from A. adeninivorans for rDNA targeting. Transformants were obtained for both budding cells and mycelia. In both cell types similar expression levels were achieved and the GFP was localised in the cytoplasm while more than 95% of the HSA accumulated in the culture medium. In initial fermentation trials on a 200‐ml shake flask scale maximal HSA product levels were observed after 96 h of cultivation.