Characteristics of Saccharomyces cerevisiae gal1Δ and gal1Δhxk2Δ mutants expressing recombinant proteins from the GAL promoter

• Published in: Biotechnology and bioengineering Vol. 89; no. 6; pp. 619 - 629
• Main Authors: Ah Kang, Hyun, Kyu Kang, Woo, Go, Su-Min, Rezaee, Abbas, Hari Krishna, Sajja, Ki Rhee, Sang, Kim, Jeong-Yoon
• Format: Journal Article
• Language: English
• Published: Hoboken Wiley Subscription Services, Inc., A Wiley Company 20.03.2005; Wiley
• Subjects: Biological and medical sciences; Biotechnology; DNA-Binding Proteins - genetics; DNA-Binding Proteins - metabolism; Ethanol - metabolism; Fermentation; Fundamental and applied biological sciences. Psychology; GAL promoter; GAL1; Galactose - genetics; Galactose - metabolism; Genes, Fungal; Genetic Engineering; Genetic technics; Hexokinase - genetics; Hexokinase - metabolism; Humans; HXK2; Kinetics; Methods. Procedures. Technologies; Modification of gene expression level; Mutation; Promoter Regions, Genetic; recombinant protein; Recombinant Proteins - metabolism; Repressor Proteins - genetics; Repressor Proteins - metabolism; Saccharomyces cerevisiae; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae - growth & development; Saccharomyces cerevisiae - metabolism; Saccharomyces cerevisiae Proteins; Serum Albumin - genetics; Serum Albumin - metabolism; Yeast; Transcription promoter; Gene expression; Fungi; HXK2; GAL promoter; Ascomycetes; Mutation; Recombinant protein; Galactose; Saccharomyces cerevisiae; GAL1; Thallophyta
• ISSN: 0006-3592; 1097-0290
• DOI: 10.1002/bit.20240

Galactose can be used not only as an inducer of the GAL promoters, but also as a carbon source by Saccharomyces cerevisiae, which makes recombinant fermentation processes that use GAL promoters complicated and expensive. To overcome this problem during the cultivation of the recombinant strain expressing human serum albumin (HSA) from the GAL10 promoter, a gal1Δ mutant strain was constructed and its induction kinetics investigated. As expected, the gal1Δ strain did not use galactose, and showed high levels of HSA expression, even at extremely low galactose concentrations (0.05–0.1 g/L). However, the gal1Δ strain produced much more ethanol, in a complex medium containing glucose, than the GAL1 strain. To improve the physiological properties of the gal1Δ mutant strain as a host for heterologous protein production, a null mutation of either MIG1 or HXK2 was introduced into the gal1Δ mutant strain, generating gal1Δmig1Δ and gal1Δhxk2Δ double strains. The gal1Δhxk2Δ strain showed a decreased rate of ethanol synthesis, with an accelerated rate of ethanol consumption, compared to the gal1Δ strain, whereas the gal1Δmig1Δ strain showed similar patterns to the gal1Δ strain. Furthermore, the gal1Δhxk2Δ strain secreted much more recombinant proteins (HSA and HSA fusion proteins) than the other strains. The results suggest that the gal1Δhxk2Δ strain would be useful for the large‐scale production of heterologous proteins from the GAL10 promoter in S. cerevisiae. © 2005 Wiley Periodicals, Inc.