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ToolsBrummett, Adam E, Schnicker, Nicholas J, Crider, Alexander, Todd, Jonathan and Dey, Mishtu (2015) Biochemical, kinetic, and spectroscopic characterization of Ruegeria pomeroyi DddW - A mononuclear iron-dependent DMSP lyase. PLoS One, 10 (5). ISSN 1932-6203
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Abstract
The osmolyte dimethylsulfoniopropionate (DMSP) is a key nutrient in marine environments and its catabolism by bacteria through enzymes known as DMSP lyases generates dimethylsulfide (DMS), a gas of importance in climate regulation, the sulfur cycle, and signaling to higher organisms. Despite the environmental significance of DMSP lyases, little is known about how they function at the mechanistic level. In this study we biochemically characterize DddW, a DMSP lyase from the model roseobacter Ruegeria pomeroyi DSS-3. DddW is a 16.9 kDa enzyme that contains a C-terminal cupin domain and liberates acrylate, a proton, and DMS from the DMSP substrate. Our studies show that as-purified DddW is a metalloenzyme, like the DddQ and DddP DMSP lyases, but contains an iron cofactor. The metal cofactor is essential for DddW DMSP lyase activity since addition of the metal chelator EDTA abolishes its enzymatic activity, as do substitution mutations of key metal-binding residues in the cupin motif (His81, His83, Glu87, and His121). Measurements of metal binding affinity and catalytic activity indicate that Fe(II) is most likely the preferred catalytic metal ion with a nanomolar binding affinity. Stoichiometry studies suggest DddW requires one Fe(II) per monomer. Electronic absorption and electron paramagnetic resonance (EPR) studies show an interaction between NO and Fe(II)-DddW, with NO binding to the EPR silent Fe(II) site giving rise to an EPR active species (g = 4.29, 3.95, 2.00). The change in the rhombicity of the EPR signal is observed in the presence of DMSP, indicating that substrate binds to the iron site without displacing bound NO. This work provides insight into the mechanism of DMSP cleavage catalyzed by DddW.
| Item Type: | Article |
|---|---|
| Additional Information: | © 2015 Brummett et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited |
| Uncontrolled Keywords: | dmsp,dms,biogeochemistry,biochemistry,sdg 14 - life below water ,/dk/atira/pure/sustainabledevelopmentgoals/life_below_water |
| Faculty \ School: | Faculty of Science > School of Biological Sciences |
| UEA Research Groups: | Faculty of Science > Research Groups > Molecular Microbiology |
| Depositing User: | Pure Connector |
| Date Deposited: | 24 Jul 2015 22:55 |
| Last Modified: | 21 Oct 2022 01:01 |
| URI: | https://ueaeprints.uea.ac.uk/id/eprint/53756 |
| DOI: | 10.1371/journal.pone.0127288 |
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