Sweets such as candies and cakes have gradually become popular due to their enticing flavors.
However, with the increasing amounts of sweeteners like lactose and fructose in these desserts,health issues have emerged.
According to surveys, the number of obese people around the world has increased dramatically year by year, and has even reached 890 million people. Also,422 million people with diabetes. In developing countries, on average, there are three diabetics in every four adults.
The diabetic community and obese people have to give up desserts for the sake of their health.
The fitness and body management crowd also shun sweets for the sake of being super fit.
Sugar seems to be the ‘worm’ in people's lives. When you eat sugar, it’s like you have become a sinner of your body.
But, is all sugar so harmful to one's health and figure?
Is there any sugar that can be enjoyed without the burden of sweetness?
Fortunately, there really is.
Tagatose, a natural sweetener and functional food ingredient, can perfectly balance sweetness and sugar content. It provides a delightful sweet taste without imposing additional health burdens.
However, tagatose is not widely used currently due to its high production cost and low yield. Fructose can be converted to tagatose through a single-step reaction at the C4 position, significantly reducing the production cost of tagatose.
To achieve efficient production of tagatose, we focused on tagatose-4-epimerase. Through genetic exploration, we identified four unknown functional proteins with potential tagatose-4-epimerase activity and employed directed evolution strategies to cultivate strains with higher activity and conversion rates.
Fructose is an isomer of tagatose and can be converted into tagatose through a one-step 4-position epimerization, making it an ideal substrate for tagatose production. However, a natural tagatose 4-epimerase has not yet been discovered. Using UxaE as a sequence template, we selected five protein sequences with potential tagatose-4-epimerase activity based on binding free energy and docking results, through homologous substrate modeling and molecular docking.
To address the common issue of inclusion body problems when expressing foreign genes in E. coli, we modified AJCT to increase its solubility, thereby achieving its efficient soluble expression. Plan 1: We insert the DNA of the target protein and fusion tag into a plasmid, followed by transcription and translation to produce the target protein attached to the fusion tag. Plan 2: We co-express molecular chaperones and the recombinant plasmid in BL21. Plan 3: We adjust the fermentation conditions to influence the growth and metabolic activities of the microorganisms or cells, thereby affecting the protein expression levels.
However, due to the low enzyme activity of the wild-type enzyme, our goal is to enhance its activity through site-directed mutagenesis. We begin with single-point mutations and continue with double, triple, quadruple, and quintuple mutations, ultimately aiming to improve its ability to catalyze fructose to produce tagatose.
Until now, Tagatose has not been able to come into the public eye due to its expensive price; we hope that our research can reduce the cost of Tagatose, so that everyone can enjoy the sweet taste without any burden at the same time.
