1. Heterologous expression and characterization of the manganese-oxidizing protein from Erythrobacter sp. strain SD21.
- Author
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Nakama K, Medina M, Lien A, Ruggieri J, Collins K, and Johnson HA
- Subjects
- Bacterial Proteins chemistry, Bacterial Proteins genetics, Calcium metabolism, Cloning, Molecular, Coenzymes metabolism, Enzyme Activators, Escherichia coli genetics, Escherichia coli metabolism, Gene Expression, Hemin metabolism, Kinetics, Molecular Weight, Oxidation-Reduction, PQQ Cofactor metabolism, Recombinant Proteins chemistry, Recombinant Proteins genetics, Recombinant Proteins metabolism, Bacterial Proteins metabolism, Manganese metabolism, Sphingomonadaceae enzymology
- 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., (Copyright © 2014, American Society for Microbiology. All Rights Reserved.)
- Published
- 2014
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