Manganese-oxide minerals (MnOx) are widely distributed Over the Earth's surface, and their geochemical cycling is globally important. A multicopper oxidase (MCO) MnxG protein from marine Bacillus bacteria plays an essential role in producing MnOx minerals by oxidizing Mn2+(aq) at rates that ate 3 to 5 orders of magnitude faster than abiotic rates. The MnxG protein is isolated as part of a mtiltiprotein complex denoted as "Mnx" that includes accessory protein subunits MnxE and MnxF, with an estimated stoichiometry of MnxE(3)F(3)G and corresponding molecular weight of approximate to 211 kDa. Herein, we report successful expression and isolation of the MCO MnxG protein without the E3F3 hexamer. This isolated MnxG shows activity for Mn2+(aq) oxidation to form manganese oxides. The complement of paramagnetic Cu(II) ions in the Mnx protein complex was examined by electron paramagnetic resonance (EPR) spectroscopy. Two distnict classes of type 2 Cu sites were detected. One class of Cu(II) site (denoted as T2Cu-A), located in the MnxG subunit, is identified by the magnetic parameters g(parallel to)= 2.320 and A(parallel to) = 510 MHz. The other class of Cu(II) sites (denoted as T2Cu-B) is characterized by g(parallel to) = 2.210 and A(parallel to) = 615 MHz and resides in the putative hexameric MnxE(3)F(3) subunit. These different magnetic properties correlate with the differences in the reduction potentials of the respective Cu(II) centers. These studies provide. new insights into the molecular mechanism of manganese biomineralization.