MopA, the Mn Oxidizing Protein From Erythrobacter sp. SD-21, Requires Heme and NAD+ for Mn(II) Oxidation

Abstract

Bacterial manganese (Mn) oxidation is catalyzed by a diverse group of microbes and can affect the fate of other elements in the environment. Yet, we understand little about the enzymes that catalyze this reaction. The Mn oxidizing protein MopA, from Erythrobacter sp. strain SD-21, is a heme peroxidase capable of Mn(II) oxidation. Unlike Mn oxidizing multicopper oxidase enzymes, an understanding of MopA is very limited. Sequence analysis indicates that MopA contains an N-terminal heme peroxidase domain and a C-terminal calcium binding domain. Heterologous expression and nickel affinity chromatography purification of the N-terminal peroxidase domain (MopA-hp) from Erythrobacter sp. strain SD-21 led to partial purification. MopA-hp is a heme binding protein that requires heme, NAD+, and calcium (Ca2+) for activity. Mn oxidation is also stimulated by the presence of pyrroloquinoline quinone. MopA-hp has a KM for Mn(II) of 154 ± 46 μM and kcat = 1.6 min−1. Although oxygen requiring MopA-hp is homologous to peroxidases based on sequence, addition of hydrogen peroxide and hydrogen peroxide scavengers had little effect on Mn oxidation, suggesting this is not the oxidizing agent. These studies provide insight into the mechanism by which MopA oxidizes Mn.

Manganese-contaminated groundwater treatment by novel bacterial isolates: kinetic study and mechanism analysis using synchrotron-based techniques

“…This incomplete oxidation issue could be governed by manganese-oxidizing activity of SBP1. Manganese oxidation by bacteria is typically from two enzymes including multicopper oxidases or peroxidase cyclooxygenases 33 , 45 , 46 . Multicopper oxidases have been widely identified in numerous bacterial species 33 , 46 , 47 .…”

Initial Formation and Accumulation of Manganese Deposits in Drinking Water Pipes: Investigating the Role of Microbial-Mediated Processes

“…Mn­(II) oxidation by microbes is perceived as an enzymatic process. Multicopper oxidase is an important enzyme responsible for MnO 2 formation, and peroxidase cyclooxygenase has been identified as an additional enzyme capable of Mn­(II) oxidation. − Although much work has been conducted to investigate Mn­(II) oxidation by MOB, previous research has mainly focused on Mn­(II) oxidation in natural aquatic environments and water treatment processes. ,− For example, a Bacillus strain (SG-1), a salt-requiring bacterium isolated from sea sediments, has been found to oxidize Mn­(II) in seawater rapidly. , In addition to marine SG-1, some other MOB strains, including Pseudomonas putida MnB1 and GB-1, Leptothrix discophora SS-1, Pedomicrobium sp. ACM 3067, and so forth, found in freshwater systems have been implicated in Mn­(II) oxidation. ,,, In drinking water systems, species of Leptothrix, Crenothrix, Hyphomicrobium, Siderocapsa, and Metallogenium have been identified in mediating the oxidative removal of Mn­(II) by using a filter. − …”

Comparative Genomics on Cultivated and Uncultivated Freshwater and Marine “CandidatusManganitrophaceae” Species Implies Their Worldwide Reach in Manganese Chemolithoautotrophy

“…Mn(H2O)6 2+ vs MnCO3, Mn(II) vs Mn(III) complexes) that have different solubilities. In comparison, known heterotrophs that catalyze Mn(II) oxidation often employ multicopper oxidase (MCO) or heme peroxidase homologs to oxidise this metal (45)(46)(47). However, the nature of these enzymes is to couple the oxidation of Mn to the direct reduction of oxygen, without a clear path for conserving any of the potential free energy energy for use by the cell.…”

“…In that case, Cyc2 and PCC_1 might serve to react with different species of Mn(II) [e.g., soluble Mn(H 2 O) 6 2+ , soluble or insoluble MnCO 3 , or Mn(HCO 3 ) 2 ] or different oxidations of Mn(II) [e.g., Mn(II) versus Mn(III)]. Employing Cyc2 and PCC_1 would differ from well-studied nonlithotrophic heterotrophs that catalyze direct Mn(II) oxidations with O 2 or reactive oxygen species, e.g., via multicopper oxidase (MCO) or heme peroxidase homologs ( 45 – 47 ), involving mechanisms without a clear path for free energy conservation. While members of “ Ca.…”