iDEC 2024 | NJTECH-CHINA-A

Background:

D-Allose is a rare monosaccharide that occurs very rarely in the natural environment. D-Allose is 80% sweeter than table sugar, but is extremely low in calories and non-toxic, making it an ideal replacement for table sugar in food. D-Allose is a low-calorie, rare sugar that has sweetness properties similar to those of sucrose but accounts for only 38% of the energy of sucrose. D-allulose has anticancer, antioxidant, anti-inflammatory, anti-hypertensive, anti-osteoporotic, neuroprotective, cryoprotective and immunosuppressive properties, and therefore has a promising future for use in food systems, clinical treatments and healthcare.

Challenge:

Unfortunately, most rare sugars, including D-allose, are prohibitively expensive, and their limited natural availability may impede their industrial potential. The rapid advancement of synthetic biology has facilitated the biosynthesis of D-allose. However, challenges remain, including the low activity and stability of the enzymes, as well as the low productivity and yield of strains involved in D-allose synthesis. To enhance the performance of enzymes for D-allose production, directed evolution presents a promising strategy. However, there is currently a lack of high-throughput selection methods specifically for D-allose.

Methods:

Genetically encoded whole-cell biosensors are capable of sensing changes in metabolite concentrations inside and outside the cell as well as fluctuations in the external environment, generating measurable signal outputs or regulating gene expression levels in pathways. In this study, we developed a D-allose detection biosensor system utilizing the specific D-allose-responsive transcriptional regulator alsR in Escherichia coli to dynamically monitor D-allose levels. Further directed evolution of alsR yielded a biosensor with enhanced sensitivity, thereby providing a potential selection tool for the directed evolution of enzymes related to D-allose synthesis.

Fig. 1 D-Allose biosensor based on a single plasmid system

References:

[1] FENG Zai-Ping, Gong Hui-Ling, YUAN Hui-Jun et al. Research progress on the properties and bioproduction of the rare sugar D-allulose[J]. Food and Fermentation Industry, 2015, (6):227-233.

[2] Boada Y, Vignoni A, Pic6 J, et al. Extended metabolic biosensordesign for dynamic pathway regulation of cell factories. iScience2020 , 23( 7) : 101305.

[3] Yu WW, Xu X H, Jin K, e al. Genetically encoded biosensors formicrobial synthetic biology : from conceptual frameworks to practicalapplications. Biotechnology Advances , 2023 , 62 : 108077.