Rational Design of <i>Bacillus coagulans</i> NL01 <scp>l</scp>-Arabinose Isomerase and Use of Its F279I Variant in <scp>d</scp>-Tagatose Production

Abstract

d-Tagatose is a prospective functional sweetener that can be produced by l-arabinose isomerase (AI) from d-galactose. To improve the activity of AI toward d-galactose, the AI of Bacillus coagulans was rationally designed on the basis of molecular modeling and docking. After alanine scanning and site-saturation mutagenesis, variant F279I that exhibited improved activity toward d-galactose was obtained. The optimal temperature and pH of F279I were determined to be 50 °C and 8.0, respectively. This variant possessed 1.4-fold catalytic efficiency compared with the wild-type (WT) enzyme. The recombinant Escherichia coli overexpressing F279I also showed obvious advantages over the WT in biotransformation. Under optimal conditions, 67.5 and 88.4 g L^-1 d-tagatose could be produced from 150 and 250 g L^-1 d-galactose, respectively, in 15 h. The biocatalyst constructed in this study presents a promising alternative for large-scale d-tagatose production.