Discovery and characterization of a novel tagatose-4-epimerase

Abstract:

d-tagatose is a natural and rare ketohexose with blood glucose-lowering, anti-caries, gut microbiome-regulating, blood circulation-promoting, and anti-aging activities, and thus it is widely applied in food, pharmaceutical, and cosmetic industries. Though the enzymatic conversion of galactose into tagatose is a simple process, the high cost of galactose limits its industrial application. Fructose, an isomer of tagatose, is an economical alternative with stable supply. Through 4-epimerization, tagatose can be produced from fructose, which is an ideal substrate for tagatose production. [Objective] To discover new tagatose-4-epimerases essential for the efficient industrial production of tagatose. [Methods] Gene mining was performed to identify an unknown protein HCZ0 with 4-epimerization activity from Thermotogae bacterium. HCZ0 was heterologously expressed, purified, and characterized. [Results] HCZ0 had a molecular weight of 57.9 kDa and the specific activity of 0.9 U/mg. The enzyme showed the highest activity at 70 ℃ and pH 9.0, and it activity was significantly improved by 2 mmol/L Ni²⁺. HCZ0 exhibited the half-life of 180, 67, and 9 h at 60, 70, and 80 ℃, respectively. Under optimal conditions, the HCZ0 was able to catalyze 200 g/L fructose to produce 28 g/L tagatose within 2 h. [Conclusion] HCZ0 is an alkaline high-temperature enzyme with good thermal stability, which can provide inspiration for future theoretical research and industrial production.

Key words:d-tagatose;tagatose-4-epimerase;fructose isomerization;d-fructose