A thermophilic phosphatase from Methanothermobacter marburgensis and its application to in vitro biosynthesis.

Phosphatases catalyze the irreversible dephosphorylation of phosphate-containing compounds, and hence can be applied as the final enzymatic step for the synthesis of various biochemicals. However, the extensive substrate spectrums of phosphatases impose a great challenge for efficient biomanufacturing. Characterization of phosphatases is therefore of extreme importance. In this study, MmPase, a putative HAD phosphatase from Methanothermobacter marburgensis, was expressed, purified, and characterized. Recombinant MmPase was readily expressed in Escherichia coli, and required metal ions such as Mn2+ or Mg2+ to function. MmPase worked optimally at 50 °C, pH 6.5, and exhibited a half-life of 6.5 h under this condition. Among all substrates tested, MmPase established the highest dephosphorylation activity against D-tagatose 6-phosphate, and was relatively specific for this substrate than for D-glucose 1-phosphate, D-glucose 6-phosphate, and D-fructose 6-phosphate. Therefore, MmPase was integrated into an in vitro synthetic enzymatic biosystem for the one-pot production of D-tagatose from maltodextrin, and achieved a product yield of 37.6%. Our studies of MmPase provided a promising strategy for the economic and efficient production of D-tagatose in the future.