3-Hydroxypropionate (3HP) is one of top 12 value-added chemicals from biomass ranked in the list released by the US Department of Energy. The nature of the molecular structure makes it capable of converting to various compounds. The microbial production of 3HP with inexpensive bio-based stocks is one of the attractive researches in the field of metabolic engineering.  

Currently, important progress in 3HP biosynthesis via malonyl-CoA pathway with low cost and high efficiency has been made by the Bio-based Bulk Chemicals Group, Qingdao Institute of Bioenergy and Bioprocess Technology (QIBEBT), Chinese Academy of Sciences. 

Malonyl-CoA reductase (MCR), a key enzyme in themalonyl-CoA pathway, was derived from the thermophilic bacterium Chloroflexus aurantiacus. It had been inferred that the enzyme could complete a two-step reaction from malonyl-CoA to 3HP, but the reaction mechanism remained a mystery. 

To answer the puzzle, researchers tried dividing the MCR protein into two parts named MCR-C and MCR-N, and each part was discovered to catalyze an independent reaction. The MCR-C could catalyze malonyl-CoA into an intermediate which was further converted to 3HP by the MCR-N. However, the dissection of MCR brought about a higher substrate affinity and enzyme activity, which made 3HP production promoted in recombinant Escherichia coli. This result shows feasibility to develop a new strategy different from constructing fusion proteins to improve catalytic efficiency and product yield. Also, it has a certain reference value for the use of multi-functional enzymes in metabolic engineering. 

Emerson said, “all is riddle, and the key to a riddle is another riddle.” After the dissection of MCR, a serious functional imbalance between the MCR-C and MCR-N was found to be a contribution to the low 3HP yield. To resolve the problem, the catalytic efficiency of MCR-C was increased about 15 times higher than before through laboratory evolution, and the functional match was achieved by fine-tuning of protein expression level. Combined with culture conditions optimization, the engineering approaches increased the 3HP concentration by hundreds of times. Finally, the production of 3HP rose from 0.15 g/L to 40.6 g/L with glucose as the sole carbon source. Recently, the findings have been published in Metabolic Engineering (Figure 1). 

Figure 1. The malonyl-CoA route for 3-hydroxypropionate biosynthesis and the laboratory evolutionprocess of MCR-C protein. (Image by QIBEBT)