[P003] Poster

12:10 PM - 1:00 PM JST(3:10 AM - 4:00 AM UTC)

[P02-213] Toward Reductive Fermentation using the NOG strain

○Shao-Hsiang Chien¹, Ying-Hsuan Lin¹, Paul P. Lin¹ (1. National Yang Ming Chiao Tung University (Taiwan))

Keywords:

Non-Oxidative Glycolysis,Reductive Fermentation,Adaptive Laboratory Evolution

Acetyl-CoA is a versatile precursor for numerous bioproducts, including fatty acids, alcohols, alkanes, isoprenoids, and polyketides. Most organisms metabolize sugars into pyruvate through the Embden-Meyerhof-Parnas (EMP) pathway, followed by the oxidative decarboxylation of pyruvate into acetyl-CoA, a process that releases CO₂. Consequently, the maximum carbon yield for acetyl-CoA-derived products is capped at 67% unless CO₂ or formate is reassimilated through carbon fixation. To overcome this limitation, we previously constructed and evolved an Escherichia coli NOG strain capable of converting nearly 90% of glucose-derived carbon into acetate. However, to fully realize the potential of NOG for more reduced compounds, additional reducing equivalents must be generated from the carbon source or provided externally.
Here, we present a NOG-based reductive fermentation strategy for converting glucose to ethanol with yields exceeding the standard EMP theoretical limit. To direct carbon flux, we deleted acetate kinase to redirect acetyl phosphate toward acetyl-CoA rather than acetate. We also expressed CoA-acylating aldehyde dehydrogenase (PduP) and alcohol dehydrogenase (AdhA) for ethanol production. Initially, the resulting engineered strain (NOGe1) exhibited severe growth defects in minimal medium, requiring casamino acid supplementation to bypass an extended lag phase. Through adaptive laboratory evolution (ALE), we successfully developed a robust strain capable of utilizing glucose and formate in minimal medium. We are currently evaluating ethanol titers and yields from glucose in bioreactors.