Abstract
In this study, we conducted the characterization and purification of the thermostable mannitol dehydrogenase (MtDH) from Caldicellulosiruptor hydrothermalis 108. Furthermore, a coupling-enzyme system was designed using (MtDH) from Caldicellulosiruptor hydrothermalis 108 and formate dehydrogenase (FDH) from Ogataea parapolymorpha. The biotransformation system was constructed using Escherichia coli whole cells. The purified enzyme native and subunit molecular masses were 76.7 and 38 kDa, respectively, demonstrating that the enzyme was a dimer. The purified and couple enzyme system results were as follows; the optimum pH for the reduction and the oxidation was 7.0 and 8.0, the optimum temperature was 60 °C, the enzyme activity was inhibited by EDTA and restored by zinc. Additionally, no activity was detected with NADPH and NADP. The purified enzyme showed high catalytic efficiency Kcat 385 s−1, Km 31.8 mM, and kcat/Km 12.1 mM−1 s−1 for D-fructose reduction. Moreover, the purified enzyme retained 80%, 75%, 60%, and 10% of its initial activity after 4 h at 55, 60, 65, and 75 °C, respectively. D-mannitol yield was achieved via HPLC. Escherichia coli are the efficient biotransformation mediator to produce D-mannitol (byproducts free) at high temperature and staple pH, resulting in a significant D-mannitol conversation (41 mg/mL) from 5% D-fructose.
Highlights
D-mannitol polyol is a safe choice when added to foods and beverages
D-mannitol production from D-fructose was concerned with the high bioconversion ratio, which reached ~100% [5]
D-mannitol, D-fructose, sodium formate (HCOONa), and other chemicals used in this study were obtained from Sigma-Aldrich (Shanghai, China)
Summary
D-mannitol polyol is a safe choice when added to foods and beverages. Many health problems are related to the daily consumption of added sugars, so the wise choice is to replace them with healthy alternatives. A possible option is D-mannitol, which is likely to be produced and other polyols at industrial levels by genetically modified microorganisms that offer exciting new future potentials, leading to outstanding results with enormous possibilities for the food and pharmaceutical industries [1]. Research has focused on improving polyols production using low-cost biotechnology protocols. The main challenge is to improve the overall production of D-mannitol without sorbitol combination through genetic engineering tools [2]. D-mannitol production from D-fructose was concerned with the high bioconversion ratio, which reached ~100% [5]
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