The same genetic regulation strategy produces inconsistent effects in different Saccharomyces cerevisiae strains for 2-phenylethanol production

A CRISPR/Cas9 system with gene editing efficiency of 100% in the industrial diploid Saccharomyces cerevisiae CWY-132 strain for 2-phenylethanol (2-PE) production was constructed. The effect of deletion of acetyltransferase gene ATF1 in the Ehrlich pathway on 2-PE synthesis was studied for the first time in S. cerevisiae. Laboratory and industrial strains were compared for the deletion effect of ATF1 and acetaldehyde dehydrogenase genes ALD2 and ALD3 involved in competing branches of the Ehrlich pathway on the 2-PE titer. The results showed that in 2-PE low-yielding haploid strain PK-2C, the ATF1 increment mutant produced 2-PE of 0.45 g/L, an increase of 114%, whereas in CWY-132, the 2-PE yield of ATF1 increment decreased significantly from 3.50 to 0.83 g/L. In PK-2C, the 2-PE yield of ALD2 increment increased from 0.21 to 1.20 g/L, whereas in CWY-132, it decreased from 3.50 to 3.02 and 2.93 g/L in ALD2 increment and ALD3 increment mutants, respectively, and to 1.65 g/L in ALD2 increment ALD3 increment . These results indicate that the same genetic manipulation strategy used for strains with different 2-PE yield backgrounds produces significantly different or even opposite effects. Moreover, we found that a supply of NADH or GSH increased the 2-PE production in S. cerevisiae. The correlation between the synthesis of 2-PE and ethanol was also revealed, and the tolerance of cells to 2-PE and ethanol was suggested to be a key limiting factor for further increase of 2-PE production in high-yielding strains.

Automated summary

Investigated the effects of gene deletion on the production of 2-phenylethanol (2-PE) in yeast, revealing that different strains showed varying responses to the same genetic manipulation strategy. Furthermore, the supply of specific substances was found to increase 2-PE production, and the tolerance of cells to 2-PE and ethanol was identified as a key limiting factor for further enhancing production.