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The metabolic burden of the PGK1 and ADH2 promoter systems for heterologous xylanase production by Saccharomyces cerevisiae in defined medium
Author(s) -
Görgens Johann F.,
van Zyl Willem H.,
Knoetze Johannes H.,
HahnHägerdal Bärbel
Publication year - 2001
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.1056
Subject(s) - heterologous , biology , heterologous expression , recombinant dna , saccharomyces cerevisiae , gene , xylanase , plasmid , promoter , trichoderma reesei , gene expression , yeast , microbiology and biotechnology , biochemistry , genetics , enzyme , cellulase
Five recombinant S. cerevisiae strains were cultivated under identical conditions to quantify the molecular basis of the metabolic burden of heterologous gene expression, and to evaluate mechanisms for the metabolic burden. Two recombinant S. cerevisiae strains, producing Trichoderma reesei xylanase II under control of either the PGK1 or ADH2 promoters, were compared quantitatively with three references strains, where either the heterologous xylanase II ( XYN2 ) gene, or the heterologous gene and the promoter and terminator were omitted from the recombinant plasmid. Neither the replication of multiple copies of the 2‐μm‐based YEp352 plasmid nor the replication the foreign XYN2 gene represented a metabolic burden to the cell, as the growth of the host organism was not affected. The inclusion of a glycolytic promoter on the recombinant plasmid, however, reduced the maximum specific growth rate (12% to 15%), biomass yield on glucose (8% to 11%), and specific glucose consumption rate (6% to 10%) of the recombinant strains. The presence of the heterologous XYN2 gene on the recombinant plasmid caused a further reduction in the maximum specific growth rate (11% to 14%), biomass yield (4%), and specific glucose consumption rate (12%) of the host strain during active gene expression, which was dictated by the regulatory characteristics of the promoter utilized. The metabolic effect of foreign gene expression was disproportionally large, with respect to on the amount of heterologous protein produced. This was most likely due to an increased energetic demand for the expression of a foreign gene and/or a competition for limiting amounts of transcription or translation factors, biosynthetic precursors or metabolic energy. © 2001 John Wiley & Sons, Inc. Biotechnol Bioeng 73: 238–245, 2001.