# Heterologous Production of Acrylic Acid: Current Challenges and Perspectives
## 5 Concluding Remarks and Future Perspectives  
AA has numerous important applications for our day-to-day life. The market is expected to grow as the population increases and ages. Therefore, more sustainable and efficient production processes ought to be developed. In the past years, there have been remarkable advances in developing artificial biosynthetic pathways for AA heterologous production in _E. coli_. Nevertheless, the heterologous production of AA is still in an initial stage and has a long way to go. In this paper, the reported pathways for AA biosynthesis in _E. coli_ were thoroughly reviewed, together with 3-HP ones, as the lessons learnt during the construction of this pathway were essential to develop new ones for AA biosynthesis. The most promising pathways for AA production so far are probably the ones that use glycerol as a substrate. There are currently several variations of the glycerol route, and all have high potential, with concentrations so far reaching up to 144 mg/L in shake-flask culture. However, routes that bypass the 3-HP production step appear more promising as fewer steps are needed to reach the goal. Although the \(\beta\)-alanine pathway reported concentrations of up to 237 mg/L, these are obtained in bioreactors using fed-batch operation mode and the concentrations obtained in batch mode in the same study are significantly less competitive (up to 60 mg/L). Therefore, in the future, the use of a larger-scale and fed-batch process is expected to highly increase the concentrations obtained using the glycerol route. Of course, it is important to not forget the limitations of these pathways first step performed so far by a B\({}_{12}\)-dependent glycerol dehydatase that represents one of the most important bottlenecks of AA (and also 3-HP) biosynthesis. As discussed in Section 4.1, it is mandatory to optimize these pathways using a B\({}_{12}\)-independent glycerol dehydatase previously reported as efficient for the production of 1,3-propanediol. This is absolutely required to move to an industrial setting using this pathway, as it most probably makes no sense to select a pathway that can use the by-product crude glycerol but needs the supplementation of the expensive vitamin B\({}_{12}\). Several enzymes should be evaluated to find the best candidate to perform this step. In addition, the glycerol route that produces acrolein as an intermediary also needs to be more explored as it was the one that allowed obtaining higher concentrations and still has a large room for improvement. It is important to find the most adequate enzymes for this pathway as only two enzymes were evaluated to convert 3-HPA to acrolein and the step that converts acrolein to AA was left to an endogenous enzyme that was not overexpressed.  
In the future, as some of the identified bottlenecks related to enzyme selection resolve, others will emerge and metabolic engineering and synthetic biology strategies will continue to be essential to balance the pathway (e.g., CRISPR/a) and optimize AA production levels. Synthetic biology tools will continue to evolve and to respond to the biotechnological field challenges and contribute to the development of a sustainable, efficient and fast process for AA biosynthesis.  
This study was funded by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of the UIDB/04469/2020 unit.  
Not applicable.  
The author declares no conflict of interest. The funders had no role in the design and writing of the manuscript.