The Library
O2-independent demethylation of trimethylamine N-oxide by Tdm of Methylocella silvestris
Tools
Zhu, Yijun , Ksibe, Amira Z., Schäfer, Hendrik, Blindauer, Claudia A., Bugg, Tim and Chen, Yin (2016) O2-independent demethylation of trimethylamine N-oxide by Tdm of Methylocella silvestris. FEBS Journal, 283 (21). pp. 3979-3993. doi:10.1111/febs.13902 ISSN 1742-464X.
PDF
WRAP_Zhu_et_al-2016-The_FEBS_Journal.pdf - Published Version - Requires a PDF viewer. Available under License Creative Commons Attribution 4.0. Download (1176Kb) |
|
PDF
WRAP_0380721-lf-160916-zhu_2016_febsj.pdf - Accepted Version Embargoed item. Restricted access to Repository staff only - Requires a PDF viewer. Download (3002Kb) |
Official URL: http://dx.doi.org/10.1111/febs.13902
Abstract
Bacterial trimethylamine N-oxide (TMAO) demethylase, Tdm, carries out an unusual oxygen-independent demethylation reaction, resulting in the formation of dimethylamine and formaldehyde. In this study, sitedirected mutagenesis, homology modelling and metal analyses by inorganic mass spectrometry have been applied to gain insight into metal stoichiometry and underlying catalytic mechanism of Tdm of Methylocella silvestris BL2. Herein, we demonstrate that active Tdm has 1 molar equivalent of Zn2+ and 1 molar equivalent of non-heme Fe2+. We further investigated Zn2+ and Fe2+-binding sites through homology modelling and sitedirected mutagenesis and found that Zn2+ is coordinated by a 3-sulfur-1-O motif. An aspartate residue (D198) likely bridges Fe2+ and Zn2+ centres, either directly or indirectly via H-bonding through a neighbouring H2O molecule. H276 contributes to Fe2+ binding, mutation of which results in an inactive enzyme, and the loss of iron, but not zinc. Site-directed mutagenesis of Tdm also led to the identification of three hydrophobic aromatic residues likely involved in substrate coordination (F259, Y305, W321), potentially through a cation- interaction. Furthermore, a cross-over experiment using a substrate analogue gave direct evidence that a trimethylamine-alike intermediate was produced during the Tdm catalytic cycle, suggesting TMAO has a dual role of being both a substrate and an oxygen donor for formaldehyde formation. Together, our results provide novel insight into the role of Zn2+ and Fe2+ in the catalysis of TMAO demethylation by this unique oxygenindependent enzyme.
Item Type: | Journal Article | ||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|
Divisions: | Faculty of Science, Engineering and Medicine > Science > Chemistry Faculty of Science, Engineering and Medicine > Science > Life Sciences (2010- ) |
||||||||||
Journal or Publication Title: | FEBS Journal | ||||||||||
Publisher: | Wiley-Blackwell Publishing Ltd. | ||||||||||
ISSN: | 1742-464X | ||||||||||
Official Date: | November 2016 | ||||||||||
Dates: |
|
||||||||||
Volume: | 283 | ||||||||||
Number: | 21 | ||||||||||
Page Range: | pp. 3979-3993 | ||||||||||
DOI: | 10.1111/febs.13902 | ||||||||||
Status: | Peer Reviewed | ||||||||||
Publication Status: | Published | ||||||||||
Access rights to Published version: | Open Access (Creative Commons) | ||||||||||
Date of first compliant deposit: | 19 September 2016 | ||||||||||
Date of first compliant Open Access: | 26 October 2016 | ||||||||||
Related URLs: |
Request changes or add full text files to a record
Repository staff actions (login required)
View Item |
Downloads
Downloads per month over past year