Dmytruk K. Yeast metabolic engineering for construction of efficient producers of riboflavin and ethanol

Object of the research: the metabolic pathways for the construction of riboflavin and ethanol producers on yeast. The aim of the work: to develop approaches for the construction of riboflavin overproducers based on yeast Candida famata, ethanol producers of the first and second generations on yeast Saccharomyces cerevisiae, Pichia stipitis and Hansenula polymorpha using metabolic engineering approaches. Methods: genetic, molecular biological, biochemical and microbiological. For the first time transcription factor Sef1 was identified. Sef1 is involved in the regulation of the vitamin B2 synthesis. Sef1 determines the stability of industrial riboflavin producing strain dep8 of C. famata. Overexpression of regulatory SEF1 and structural RIB1 and RIB7 genes possessed riboflavin accumulation over 16 g/l of vitamin B2. Genes PHO8 and apy were overexpressed in yeast S. cerevisiae for improvement of ethanol production. Derepression of initial stages of xylose catabolism allowed to select the H. polymorpha mutants with increased ethanol yield during fermentation of xylose at 45 °C. Deletion of genes ATG13 and CAT8 increases the production of ethanol during xylose fermentation. Peroxisomal enzymes dihydroxyacetone synthase and transaldolase are involved in the process of xylose alcoholic fermentation in H. polymorpha, and derepression of the corresponding genes leads to increased production of ethanol to 12 g/l.