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Transforming Flask Reaction into Cell-Based Synthesis: Production of Polyhydroxylated Molecules via Engineered Escherichia coli

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posted on 02.07.2015, 00:00 by Mohui Wei, Zijie Li, Tiehai Li, Baolin Wu, Yunpeng Liu, Jingyao Qu, Xu Li, Lei Li, Li Cai, Peng George Wang
Dihydroxyacetone phosphate (DHAP)-dependent aldolases have been intensively studied and widely used in the synthesis of carbohydrates and complex polyhydroxylated molecules. However, strict specificity toward donor substrate DHAP greatly hampers their synthetic utility. Here, we transformed DHAP-dependent aldolases-mediated by in vitro reactions into bioengineered Escherichia coli (E. coli). Such flask-to-cell transformation addressed several key issues plaguing in vitro enzymatic synthesis: (1) it solves the problem of DHAP availability by in vivo-hijacking DHAP from the glycolysis pathway of the bacterial system, (2) it circumvents purification of recombinant aldolases and phosphatase, and (3) it dephosphorylates the resultant aldol adducts in vivo, thus eliminating the additional step for phosphate removal and achieving in vivo phosphate recycling. The engineered E. coli strains tolerate a wide variety of aldehydes as acceptor and provide a set of biologically relevant polyhydroxylated molecules in gram scale.