TY - JOUR
T1 - Discovering a novel d-xylonate-responsive promoter
T2 - the PyjhI-driven genetic switch towards better 1,2,4-butanetriol production
AU - Bañares, Angelo B.
AU - Valdehuesa, Kris Niño G.
AU - Ramos, Kristine Rose M.
AU - Nisola, Grace M.
AU - Lee, Won Keun
AU - Chung, Wook Jin
PY - 2019/10/1
Y1 - 2019/10/1
N2 - The capability of Escherichia coli to catabolize d-xylonate is a crucial component for building and optimizing the Dahms pathway. It relies on the inherent dehydratase and keto-acid aldolase activities of E. coli. Although the biochemical characteristics of these enzymes are known, their inherent expression regulation remains unclear. This knowledge is vital for the optimization of d-xylonate assimilation, especially in addressing the problem of d-xylonate accumulation, which hampers both cell growth and target product formation. In this report, molecular biology techniques and synthetic biology tools were combined to build a simple genetic switch controller for d-xylonate. First, quantitative and relative expression analysis of the gene clusters involved in d-xylonate catabolism were performed, revealing two d-xylonate-inducible operons, yagEF and yjhIHG. The 5′-flanking DNA sequence of these operons were then subjected to reporter gene assays which showed PyjhI to have low background activity and wide response range to d-xylonate. A PyjhI-driven synthetic genetic switch was then constructed containing feedback control to autoregulate d-xylonate accumulation and to activate the expression of the genes for 1,2,4-butanetriol (BTO) production. The genetic switch effectively reduced d-xylonate accumulation, which led to 31% BTO molar yield, the highest for direct microbial fermentation systems thus far. This genetic switch can be further modified and employed in the production of other compounds from d-xylose through the xylose oxidative pathway.
AB - The capability of Escherichia coli to catabolize d-xylonate is a crucial component for building and optimizing the Dahms pathway. It relies on the inherent dehydratase and keto-acid aldolase activities of E. coli. Although the biochemical characteristics of these enzymes are known, their inherent expression regulation remains unclear. This knowledge is vital for the optimization of d-xylonate assimilation, especially in addressing the problem of d-xylonate accumulation, which hampers both cell growth and target product formation. In this report, molecular biology techniques and synthetic biology tools were combined to build a simple genetic switch controller for d-xylonate. First, quantitative and relative expression analysis of the gene clusters involved in d-xylonate catabolism were performed, revealing two d-xylonate-inducible operons, yagEF and yjhIHG. The 5′-flanking DNA sequence of these operons were then subjected to reporter gene assays which showed PyjhI to have low background activity and wide response range to d-xylonate. A PyjhI-driven synthetic genetic switch was then constructed containing feedback control to autoregulate d-xylonate accumulation and to activate the expression of the genes for 1,2,4-butanetriol (BTO) production. The genetic switch effectively reduced d-xylonate accumulation, which led to 31% BTO molar yield, the highest for direct microbial fermentation systems thus far. This genetic switch can be further modified and employed in the production of other compounds from d-xylose through the xylose oxidative pathway.
KW - 1,2,4-Butanetriol
KW - d-Xylonate
KW - Dahms pathway
KW - Genetic switch
KW - yjhI promoter
UR - http://www.scopus.com/inward/record.url?scp=85072115573&partnerID=8YFLogxK
U2 - 10.1007/s00253-019-10073-0
DO - 10.1007/s00253-019-10073-0
M3 - Article
C2 - 31482281
AN - SCOPUS:85072115573
VL - 103
SP - 8063
EP - 8074
JO - Applied microbiology and biotechnology
JF - Applied microbiology and biotechnology
SN - 0175-7598
IS - 19
ER -