Highly efficient assimilation of lactose by a metabolically engineered strain of Saccharomyces cerevisiae.
A diploid strain of Saccharomyces cerevisiae able to metabolize lactose with high efficiency has been obtained. Haploid strains of Saccharomyces able to grow on lactose were constructed by cotransformation with two genes of Kluyveromyces lactis required for the utilization of the sugar, LAC4 and LAC12, encoding beta-galactosidase and lactose permease respectively. Both genes were placed under the control of a galactose-inducible promoter and targeted to the rDNA encoding region (RDN1 locus) of the Saccharomyces genome. Lac+ transformants were selected on medium with lactose as the only carbon source. These transformants were mitotically stable, they maintained the Lac+ phenotype after growing in non-selective medium for more than 60 generations, but their growth was slow. We found that this lack of vigour was caused by their genetic background and not by a deficient expression of the heterologous genes. Therefore, their performance could be improved by crossing with a wild-type strain. Among the offspring of the crosses, two strains of opposite mating type were selected and mated to obtain a fast-growing Lac+ diploid. This diploid strain showed the typical fermentative behaviour of S. cerevisiae when it was grown in aerated liquid medium with glucose. In lactose medium, it exhibited a respiro-fermentative metabolism similar to that of K. lactis, with low ethanol production and high biomass yield.
Rubio-Texeira
M
M
Instituto de Agroquímica y Tecnología de Alimentos, Consejo Superior de Investigaciones Científicas, Valencia, Spain.
Castrillo
J I
JI
Adam
A C
AC
Ugalde
U O
UO
Polaina
J
J
D018533:Biomass; D001709:Biotechnology; D003470:Culture Media; D004275:DNA, Ribosomal; D029968:Escherichia coli Proteins; D005285:Fermentation; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal; D049770:Genes, Mating Type, Fungal; D007716:Kluyveromyces / Q000201:enzymology / Q000235:genetics / Q000254:growth & development; D007785:Lactose / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism; D009004:Monosaccharide Transport Proteins; D020161:Physical Chromosome Mapping; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D027981:Symporters; D014170:Transformation, Genetic; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D003470:Culture Media; D004275:DNA, Ribosomal; D029968:Escherichia coli Proteins; C050221:LacY protein, E coli; D026901:Membrane Transport Proteins; D009004:Monosaccharide Transport Proteins; D027981:Symporters; C021499:lactose permease; D001616:beta-Galactosidase; D007785:Lactose