| Summary: | <i>Mau</i> DNA polymerase is a family A DNA polymerase isolated from <i>Massilia aurea</i>. In this study, a recombinant plasmid, His<sub>6</sub>-tagged Mau-pET28c, was constructed. His-tagged <i>Mau</i> was expressed in <i>Escherichia coli</i> Rosseta 2 (DE3) competent cells and, after optimization of purification conditions, was successfully isolated via a two-step purification system by Ni<sup>2+</sup>-chelating affinity chromatography followed by heparin affinity chromatography. The biochemical properties of <i>Mau</i> DNA polymerase were investigated next. This polymerase showed maximal polymerase activity at 30 °C, pH 8.4–8.8, 2–10 mM MgCl<sub>2</sub>, and 10–40 mM KCl. Kinetic parameters of correct and incorrect dNTP incorporation as well as DNA-binding affinity were determined too. <i>K</i><sup>dNTP</sup><sub>d,app</sub> values were found to be 16 µM for correct dNTP and 200–500 µM for incorrect dNTP. The kinetic parameter <i>k</i><sub>cat</sub> turned out to be 0.2 s<sup>−1</sup> for correct dNTP incorporation and an order of magnitude less for incorrect dNTP incorporation. It was demonstrated that <i>Mau</i> DNA polymerase has 5′→3′ and 3′→5′ exonuclease activities associated with the main activity.
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