“…The gene candidates ( SMU_118c, SMU_400, SMU_643, SMU_1443c, SMU_1678 ) were PCR amplified from S. mutans UA159 genomic DNA by using designed primers (Table 1), then cloned into the p15Tv-LIC vector as previously described (empty vector was used as a control) [40], providing an N-terminal His6-tag fusion followed by a TEV protease cleavage site. The esterases were expressed in E. coli BL21 (DE3) then harvested for further protein isolation and purification [41]. Cells were resuspended in binding buffer [50 mM Hepes (pH 7.5), 100/300 mM NaCl, 10 mM imidazole and 2% glycerol (v/v)], lysed using a sonicator, and cell debris was removed via centrifugation at 30,000G.…”