Triacetic acid lactone (TAL) is a promising platform polyketide with broad applications, especially it can be used as a precursor for the synthesis of various organic compounds.This study characterized the repeating sequences on the yeast genome to integrate the TAL biosynthesis pathway into these sites for enhanced gene expression and TAL production by Saccharomyces cerevisiae.Firstly, this study used the green fluorescent protein as the reporter to characterize multi-copy integration by the Delta1 site.It showed that truncating the promoter of a selection marker gene (for geneticin or hygromycin B) or increasing the antibiotic concentration, were effective in improving the integration efficiency and copy numbers.The highest copy number of multi-copy integration was obtained when truncating the antibiotic gene promoter to 15 bp, and using an antibiotic at 160 μg/mL.The optimization system was subsequently used to characterize the second multi-copy integration site, Delta2, with similar results.Then, the optimized multi-copy integration system was applied to introduce the TAL biosynthesis pathway into S.cerevisiae.The highest pathway copies using Delta1 and Delta2 sequences were 10 and 7, respectively, with Delta1 slightly better than Delta2.After 48 h fermentation in YTD medium, TAL titers of the Delta1-integration strain and the Delta2-integration strain were 1.50 mmol/L and 1.17 mmol/L, respectively, 460% and 337% higher than the single-copy integration strain.This study demonstrates that the multi-copy integration system is an efficient approach to introducing heterologous pathways into S.cerevisiae to improve biocatalytic efficiency.
WEI Huan
,
LIN Pingxin
,
LIU Xiuxia
,
LIU Chunli
,
YANG Yankun
,
LI Ye
,
BAI Zhonghu
. Efficient production of triacetic acid lactone using a multi-copy integration system in Saccharomyces cerevisiae[J]. Food and Fermentation Industries, 2025
, 51(4)
: 1
-10
.
DOI: 10.13995/j.cnki.11-1802/ts.038860
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