d-Allulose 3-epimerase of Bacillus sp. origin manifests profuse heat‐stability and noteworthy potential of d-fructose epimerization

<jats:title>Abstract</jats:title><jats:sec> <jats:title>Background</jats:title> <jats:p><jats:sc>d</jats:sc>-Allulose is an ultra-low calorie sugar of multifarious health benefits, including anti-diabetic and anti-obesity potential. <jats:sc>d</jats:sc>-Allulose 3-epimerase family enzymes catalyze biosynthesis of <jats:sc>d</jats:sc>-allulose <jats:italic>via</jats:italic> epimerization of <jats:sc>d</jats:sc>-fructose.</jats:p> </jats:sec><jats:sec> <jats:title>Results</jats:title> <jats:p>A novel <jats:sc>d</jats:sc>-allulose 3-epimerase (DaeB) was cloned from a plant probiotic strain, <jats:italic>Bacillus</jats:italic> sp. KCTC 13219, and expressed in <jats:italic>Bacillus subtilis</jats:italic> cells. The purified protein exhibited substantial epimerization activity in a broad pH spectrum, 6.0–11.0. DaeB was able to catalyze <jats:sc>d</jats:sc>-fructose to <jats:sc>d</jats:sc>-allulose bioconversion at the temperature range of 35 °C to 70 °C, exhibiting at least 50 % activity. It displaced excessive heat stability, with the half-life of 25 days at 50 °C, and high turnover number (<jats:italic>k</jats:italic><jats:sub>cat</jats:sub> 367 s<jats:sup>− 1</jats:sup>). The coupling of DaeB treatment and yeast fermentation of 700 g L<jats:sup>− 1</jats:sup><jats:sc>d</jats:sc>-fructose solution yielded approximately 200 g L<jats:sup>− 1</jats:sup><jats:sc>d</jats:sc>-allulose, and 214 g L<jats:sup>− 1</jats:sup> ethanol.</jats:p> </jats:sec><jats:sec> <jats:title>Conclusions</jats:title> <jats:p>The novel <jats:sc>d</jats:sc>-allulose 3-epimerase of <jats:italic>Bacillus</jats:italic> sp. origin discerned a high magnitude of heat stability along with exorbitant epimerization ability. This biocatalyst has enormous potential for the large-scale production of <jats:sc>d</jats:sc>-allulose.</jats:p> </jats:sec>