Aims: Characterization of a mutated Geobacillus stearothermophilus L-arabinose isomerase used to increase the production rate of D-tagatose. Methods and Results: A mutated gene was obtained by an error-prone polymerase chain reaction using L-arabinose isomerase gene from G. stearothermophilus as a template and the gene was expressed in Escherichia coli. The expressed mutated L-arabinose isomerase exhibited the change of three amino acids (Met(322) -> Val, Ser(393) -> Thr, and Val(408) -> Ala), compared with the wild-type enzyme and was then purified to homogeneity. The mutated enzyme had a maximum galactose isomerization activity at pH 8(.)0, 65 degrees C, and 1(.)0 mM Co2+, while the wild-type enzyme had a maximum activity at pH 8(.)0, 60 degrees C, and 1(.)0-mM Mn2+. The mutated L-arabinose isomerase exhibited increases in D-galactose isomerization activity, optimum temperature, catalytic efficiency (k(cat)/K-m) for D-galactose, and the production rate of D-tagatose from D-galactose. Conclusions: The mutated L-arabinose isomerase from G. stearothermophilus is valuable for the commercial production of D-tagatose. Significance and Impact of the Study: This work contributes knowledge on the characterization of a mutated L-arabinose isomerase, and allows an increased production rate for D-tagatose from D-galactose using the mutated enzyme.