title: Heterologous expression and characterization of the manganese-oxidizing protein from Erythrobacter sp. strain SD21.
abstract: The manganese (Mn)-oxidizing protein (MopA) from Erythrobacter sp. strain SD21 is part of a unique enzymatic family that is capable of oxidizing soluble Mn(II). This enzyme contains two domains, an animal heme peroxidase domain, which contains the catalytic site, followed by a C-terminal calcium binding domain. Different from the bacterial Mn-oxidizing multicopper oxidase enzymes, little is known about MopA. To gain a better understanding of MopA and its role in Mn(II) oxidation, the 238-kDa full-length protein and a 105-kDa truncated protein containing only the animal heme peroxidase domain were cloned and heterologously expressed in Escherichia coli. Despite having sequence similarity to a peroxidase, hydrogen peroxide did not stimulate activity, nor was activity significantly decreased in the presence of catalase. Both pyrroloquinoline quinone (PQQ) and hemin increased Mn-oxidizing activity, and calcium was required. The Km for Mn(II) of the full-length protein in cell extract was similar to that of the natively expressed protein, but the Km value for the truncated protein in cell extract was approximately 6-fold higher than that of the full-length protein, suggesting that the calcium binding domain may aid in binding Mn(II). Characterization of the heterologously expressed MopA has provided additional insight into the mechanism of bacterial Mn(II) oxidation, which will aid in understanding the role of MopA and Mn oxidation in bioremediation and biogeochemical cycling.
authors.affiliation: Center for Applied Biotechnology Studies, Department of Biological Science, California State University Fullerton, Fullerton, California, USA.
authors.forename: Katherine
authors.initials: K
authors.lastname: Nakama
authors.affiliation: Center for Applied Biotechnology Studies, Department of Biological Science, California State University Fullerton, Fullerton, California, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Medina
authors.affiliation: Center for Applied Biotechnology Studies, Department of Biological Science, California State University Fullerton, Fullerton, California, USA.
authors.forename: Ahn
authors.initials: A
authors.lastname: Lien
authors.affiliation: Center for Applied Biotechnology Studies, Department of Biological Science, California State University Fullerton, Fullerton, California, USA.
authors.forename: Jordan
authors.initials: J
authors.lastname: Ruggieri
authors.affiliation: Center for Applied Biotechnology Studies, Department of Biological Science, California State University Fullerton, Fullerton, California, USA.
authors.forename: Krystle
authors.initials: K
authors.lastname: Collins
authors.affiliation: Center for Applied Biotechnology Studies, Department of Biological Science, California State University Fullerton, Fullerton, California, USA hajohnson@fullerton.edu.
authors.forename: Hope A
authors.initials: HA
authors.lastname: Johnson
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D003001:Cloning, Molecular; D003067:Coenzymes / Q000378:metabolism; D020536:Enzyme Activators; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006427:Hemin / Q000378:metabolism; D007700:Kinetics; D008345:Manganese / Q000378:metabolism*; D008970:Molecular Weight; D010084:Oxidation-Reduction; D045542:PQQ Cofactor / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D042301:Sphingomonadaceae / Q000201:enzymology*
chemical_list: D001426:Bacterial Proteins; D003067:Coenzymes; D020536:Enzyme Activators; D011994:Recombinant Proteins; D008345:Manganese; D045542:PQQ Cofactor; D006427:Hemin; D002118:Calcium