Abstract
Malonyl-coenzyme A (malonyl-CoA) is a critical precursor for the biosynthesis of a variety of biochemicals. It is synthesized by the catalysis of acetyl-CoA carboxylase (Acc1p), which was demonstrated to be deactivated by the phosphorylation of Snf1 protein kinase in yeast. In this study, we designed a synthetic malonyl-CoA biosensor and used it to screen phosphorylation site mutations of Acc1p in Saccharomyces cerevisiae. Thirteen phosphorylation sites were mutated, and a combination of three site mutations in Acc1p, S686A, S659A, and S1157A, was found to increase malonyl-CoA availability. ACC1S686AS659AS1157A expression also improved the production of 3-hydroxypropionic acid, a malonyl-CoA-derived chemical, compared to both wild type and the previously reported ACC1S659AS1157A mutation. This mutation will also be beneficial for other malonyl-CoA-derived products.
Highlights
Saccharomyces cerevisiae is a very potential microbial cell factory for production of a wide range of fuels and chemicals due to its robustness and high tolerance to environmental stresses
FapR binds to fapO in the promoter region, which blocks the access of RNA polymerase II, repressing the transcription of downstream fatty acid pathway genes
We designed a malonyl-CoA sensor in S. cerevisiae and used yeGFP as a reporter to monitor the intracellular malonyl-CoA levels
Summary
Saccharomyces cerevisiae is a very potential microbial cell factory for production of a wide range of fuels and chemicals due to its robustness and high tolerance to environmental stresses. When engineering the cell factory to produce these products, the precursor availability is one of key limiting factors that affects the product titer. Malonyl-coenzyme A (malonyl-CoA) is a major precursor for the production of many important biochemicals and biofuels, such as polyketides, flavonoids, and fatty acid-derived chemicals (Dixon and Steele, 1999; Hara et al, 2006; Christian, 2009; Chooi and Tang, 2013). The intracellular level of malonyl-CoA is very low because malonyl-CoA synthesis is tightly regulated in S. cerevisiae (Li et al, 2014). Insufficient malonyl-CoA synthesis largely limits the production of malonyl-CoA-derived products. In S. cerevisiae, cytosolic malonyl-CoA is synthesized from acetyl-CoA by acetyl-CoA carboxylase (Acc1) and it is subsequently used for fatty acid synthesis. Eukaryotic ACCs contain two non-catalytic regions, the large central domain (CD) and the BC–CT interaction domain (BT)
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
