Characterization of a mutated Geobacillusstearothermophilusl-arabinose isomerase that increases the productionrate of d-tagatose

摘要

Aims:Characterization of a mutated Geobacillus stearothermophilusl-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 super(322) arrow right Val, Ser super(393) arrow right Thr, and Val super(408) arrow right 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 degree C, and 1.0 mM Co super(2+), while the wild-type enzyme had a maximum activity at pH 8.0, 60 degree C, and 1.0-mM Mn super(2+). The mutated l-arabinose isomerase exhibited increases in d-galactose isomerization activity, optimum temperature, catalytic efficiency (k sub(cat)/K sub(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.