Adaptive mutations in sugar metabolism restore growth on glucose in a pyruvate decarboxylase negative yeast strain
Journal article, 2015

Background: A Saccharomyces cerevisiae strain carrying deletions in all three pyruvate decarboxylase (PDC) genes (also called Pdc negative yeast) represents a non-ethanol producing platform strain for the production of pyruvate derived biochemicals. However, it cannot grow on glucose as the sole carbon source, and requires supplementation of C2 compounds to the medium in order to meet the requirement for cytosolic acetyl-CoA for biosynthesis of fatty acids and ergosterol. Results: In this study, a Pdc negative strain was adaptively evolved for improved growth in glucose medium via serial transfer, resulting in three independently evolved strains, which were able to grow in minimal medium containing glucose as the sole carbon source at the maximum specific rates of 0.138, 0.148, 0.141 h(-1), respectively. Several genetic changes were identified in the evolved Pdc negative strains by genomic DNA sequencing. Among these genetic changes, 4 genes were found to carry point mutations in at least two of the evolved strains: MTH1 encoding a negative regulator of the glucose-sensing signal transduction pathway, HXT2 encoding a hexose transporter, CIT1 encoding a mitochondrial citrate synthase, and RPD3 encoding a histone deacetylase. Reverse engineering of the non-evolved Pdc negative strain through introduction of the MTH1(81D) allele restored its growth on glucose at a maximum specific rate of 0.053 h(-1) in minimal medium with 2% glucose, and the CIT1 deletion in the reverse engineered strain further increased the maximum specific growth rate to 0.069 h(-1). Conclusions: In this study, possible evolving mechanisms of Pdc negative strains on glucose were investigated by genome sequencing and reverse engineering. The non-synonymous mutations in MTH1 alleviated the glucose repression by repressing expression of several hexose transporter genes. The non-synonymous mutations in HXT2 and CIT1 may function in the presence of mutated MTH1 alleles and could be related to an altered central carbon metabolism in order to ensure production of cytosolic acetyl-CoA in the Pdc negative strain.

Genomic DNA sequencing

Pyruvate decarboxylase

Yeast

MTH1

Reverse engineering

Hexose

Author

Yiming Zhang

Chalmers, Biology and Biological Engineering, Systems and Synthetic Biology

Guodong Liu

Chalmers, Biology and Biological Engineering, Systems and Synthetic Biology

Martin Engqvist

Chalmers, Biology and Biological Engineering, Systems and Synthetic Biology

Anastasia Krivoruchko

Chalmers, Biology and Biological Engineering, Systems and Synthetic Biology

B. M. Hallstrom

Royal Institute of Technology (KTH)

Yun Chen

Chalmers, Biology and Biological Engineering, Systems and Synthetic Biology

Verena Siewers

Chalmers, Biology and Biological Engineering, Systems and Synthetic Biology

Jens B Nielsen

Chalmers, Biology and Biological Engineering, Systems and Synthetic Biology

Microbial Cell Factories

14752859 (eISSN)

Vol. 14 116 116

Industrial Systems Biology of Yeast and A. oryzae (INSYSBIO)

European Commission (EC) (EC/FP7/247013), 2010-01-01 -- 2014-12-31.

Areas of Advance

Life Science Engineering (2010-2018)

Subject Categories

Bioinformatics and Systems Biology

DOI

10.1186/s12934-015-0305-6

More information

Latest update

2/26/2018