Trehalose synthase which catalyze inexpensive maltose to trehalose by one-step hydrolysis reaction, was considered to be a potential biocatalyst for trehalose production. In our work, the DNA encoding trehalose synthase with 6×his-tag was ligated with pGEM-T easy vector, transformed into E.coli DH5á and screened. After sequence analysis, the PCR products was digested by BamHI and AatII and ligated into pHT01 by using T4 DNA ligase to generate the recombinant vector pHT01-TreS. The purified pHT01-TreS was electrotransformed into B.subtilis W800N. The recombinant TreS protein was expressed and purified by nickel affinity chromatogaraphy. The optimum temperature and pH were 25°C and pH 8.0, respectively. The pure enzyme was stable in pH 6.0 to 9.0. It activity increased by Na⁺, K⁺ and Mg²⁺, but it was greater inhibited by Zn²⁺ and Ca²⁺.