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Title: Identification of proteins capable of metal reduction from the proteome of the Gram‐positive bacterium D esulfotomaculum reducens   MI ‐1 using an NADH ‐based activity assay

Journal Article · · Environmental Microbiology
 [1];  [2];  [2];  [3];  [3];  [3];  [4];  [5]
  1. Department of Microbiology Cornell University Ithaca NY USA
  2. Department of Proteomics and Mass Spectrometry Facility Cornell University Ithaca NY USA
  3. Department of Chemistry and Chemical Biology Cornell University Ithaca NY USA
  4. Pacific Northwest National Laboratory Richland WA USA
  5. Department of Civil and Environmental Engineering Cornell University Ithaca NY USA
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Summary Understanding of microbial metal reduction is based almost solely on studies of G ram‐negative organisms. In this study, we focus on D esulfotomaculum reducens   MI ‐1, a G ram‐positive metal reducer whose genome lacks genes with similarity to any characterized metal reductase. Using non‐denaturing separations and mass spectrometry identification, in combination with a colorimetric screen for chelated F e( III )‐ NTA reduction with NADH as electron donor, we have identified proteins from the D . reducens proteome not previously characterized as iron reductases. Their function was confirmed by heterologous expression in E scherichia coli . Furthermore, we show that these proteins have the capability to reduce soluble C r( VI ) and U ( VI ) with NADH as electron donor. The proteins identified are NADH  : flavin oxidoreductase ( D red_2421) and a protein complex composed of oxidoreductase flavin adenine dinucleotide/ NAD ( P )‐binding subunit ( D red_1685) and dihydroorotate dehydrogenase 1B ( D red_1686). D red_2421 was identified in the soluble proteome and is predicted to be a cytoplasmic protein. D red_1685 and D red_1686 were identified in both the soluble as well as the insoluble protein fraction, suggesting a type of membrane association, although PSORTb predicts both proteins are cytoplasmic. This study is the first functional proteomic analysis of D . reducens and one of the first analyses of metal and radionuclide reduction in an environmentally relevant G ram‐positive bacterium.

Sponsoring Organization:
USDOE
OSTI ID:
1400889
Journal Information:
Environmental Microbiology, Journal Name: Environmental Microbiology Vol. 17 Journal Issue: 6; ISSN 1462-2912
Publisher:
Wiley-BlackwellCopyright Statement
Country of Publication:
United Kingdom
Language:
English
Citation Metrics:
Cited by: 11 works
Citation information provided by
Web of Science

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