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High Level 3-Hydroxypropionate Production Via Malonyl-CoA Pathway


3-Hydroxypropionate (3HP) is an important platform chemical, and malonyl-CoA reductase (MCR) of Chloroflexus aurantiacus can convert malonyl-CoA into 3HP. In our study, MCR was dissected into two functional distinct fragments, and malonyl-CoA was reduced with NADPH by MCR-C into malonate semialdehyde, which was further reduced into 3HP by MCR-N. It is discovered that the 3HP production via malonyl-CoA pathway was severely depressed by low activity of MCR-C and metabolic burden caused by excessive overexpression of MCR-N and MCR-C. To increase 3HP productivity, the enzymatic activity of MCR-C was improved by directed evolution, and a series of promoters was tested to achieve balance between enzyme activity required by 3HP synthesis and metabolic burden caused by heterologous protein expression. Finally, our engineered E. coli strain accumulated 40.6 g/L 3HP using glucose as sole carbon source under fed-batch fermentation conditions, about 270 times higher than that of our original strain. Compared with other 3HP biosynthetic routes, the malonyl-CoA pathway has some advantages, such as broad raw material spectrum and free of cofactor, and will be widely used in 3HP production in the future.