Trehalose is a unique disaccharide capable of protecting proteins against environmental stress. A novel trehalose synthase (TreS) gene from Rhodococcus opacus was cloned and expressed in Escherichia coli Top10 and BL21 (DE3) pLysS, respectively. The recombinant TreS showed a molecular mass of 79 kDa. Thin layer chromatography (TLC) result suggested that this enzyme had the ability to catalyze the mutual conversion of maltose and trehalose. Moreover, high-performance liquid chromatography (HPLC) result suggested that glucose appeared as a byproduct with a conversion rate of 12 %. The purified recombinant enzyme had an optimum temperature of 25 degrees C and pH optimum around 7.0. Kinetic analysis revealed that the K-m for trehalose was around 98 mM, which was a little higher than that of maltose. The preferred substrate of TreS was maltose according to the analysis of k(cat)/K-m. Both 1 and 10 mM of Hg2+, Cu2+ and Al3+ could inhibit the TreS activity, while only 1 mM of Ca2+ and Mn2+ could increase its activity. Five amino acid residues, Asp(244), Glu(286), Asp(354), His(147) and His(353), were shown to be conserved in R. opacus TreS, which were also important for a-amylase family enzyme catalysis.
