Engineering Phosphofructokinase-deficient Escherichia coli for Improved Production of 3-Hydroxypropic Acid

Abstract

3-Hydroxypropionic acid (3HP) is a valuable platform chemical. In parallel with concerns about global climate change, it is highly desirable to bio-synthesize such valuable compound from affordable and renewable lignocellulosic feedstock. Among 3HP biosynthesis pathways from glucose, one via malonyl-CoA takes advantage of no consumption of ATP, short pathway length, and thermodynamic feasibility. Conversion from a malonyl-CoA to a 3HP requires two NADPH, thus regeneration of NADPH is one of key factors for efficient 3HP production. A list of the strategies for the issue include utilizing Embden-Meyerhof-Parnas pathway disrupted Escherichia coli mutants (e.g. ΔpfkA or Δpgi mutant). Their glycolytic flux is redirected to pentose phosphate pathway (PPP) where 2 moles of NADPH are produced per mole of glucose. Especially mutant with no phosphofructokinase activity (i.e. ΔpfkA ΔpfkB) is a potent NADPH regenerator as theoretically 12 moles of NADPH can be generated per mole of glucose by partial cyclization of PPP. However, these mutants showed significant reduced growth rate. Here, we engineered the phosphofructokinase-deficient mutant to overcome severe growth defect so that we can exploit its potential regarding increased NADPH availability and the resulting mutant was applied to 3HP biosynthesis.