Manganese peroxidase (MnP) from the white rot fungus Phanerochaete chrysosporium plays a vital role in lignin degradation. MnP functions by oxidizing Mn(II) to Mn(III) which serves as a diffusible oxidant. To test the current understanding of the structure and function of MnP and to find an alternative catalyst for oxidative delignification, we employed a new approach involving the design and engineering of a MnP using cytochrome c peroxidase (CcP) from bakers' yeast. Based on structural comparisons and computer modeling, we created a CcP mutant (MnCcP) that binds Mn(II) in a manner similar to the native enzyme and showed that the incorporation of the Mn(II)-binding site facilitates Mn(II) oxidation. Further mutations of key active site residues (W191 and W51) in MnCcP to the corresponding Phe in MnP conferred even more MnP activity of our protein model. The two mutations do not contribute equally to the activity increase. A much larger increase arises from the W51F mutation because W51 stabilizes compound II. Since the reaction of compound II with substrate is rate-limiting, a more reactive compound II increases MnP activity. Thus our approach is also capable of offering new insight into the structure↔function relationships of MnP and CcP.
