Chaplin, AK and Svistunenko, DA and Hough, MA and Wilson, MT and Vijgenboom, E and Worrall, JAR (2017) Active site maturation and activity of the copper-radical oxidase GlxA is governed by a tryptophan residue. The Biochemical Journal, 474 (5). pp. 809-825. DOI https://doi.org/10.1042/BCJ20160968
Chaplin, AK and Svistunenko, DA and Hough, MA and Wilson, MT and Vijgenboom, E and Worrall, JAR (2017) Active site maturation and activity of the copper-radical oxidase GlxA is governed by a tryptophan residue. The Biochemical Journal, 474 (5). pp. 809-825. DOI https://doi.org/10.1042/BCJ20160968
Chaplin, AK and Svistunenko, DA and Hough, MA and Wilson, MT and Vijgenboom, E and Worrall, JAR (2017) Active site maturation and activity of the copper-radical oxidase GlxA is governed by a tryptophan residue. The Biochemical Journal, 474 (5). pp. 809-825. DOI https://doi.org/10.1042/BCJ20160968
Abstract
GlxA from Streptomyces lividans is a mononuclear copper-radical oxidase and a member of the auxiliary activity family 5 (AA5). Its domain organisation and low sequence homology make it a distinct member of the AA5 family in which the fungal galactose 6-oxidase (Gox) is the best-characterized. GlxA is a key cuproenzyme in the copper-dependent morphological development of S. lividans with a function that is linked to the processing of an extracytoplasmic glycan. The catalytic site in GlxA and Gox contain two distinct one-electron acceptors comprising the copper ion and a 3'-(S-cysteinyl) tyrosine. The latter is formed post-translationally through a covalent bond between a cysteine and a copper coordinating tyrosine ligand and houses a radical. In GlxA and Gox a second coordination sphere tryptophan residue (Trp288 in GlxA) is present, but the orientation of the indole ring differs between the two enzymes creating a marked difference in the ?-? stacking interaction of the benzyl ring with the 3'-(S-cysteinyl) tyrosine. Differences in the spectroscopic and enzymatic activity have been reported between GlxA and Gox with the indole orientation suggested as a reason. Here we report a series of in vivo and in vitro studies using the W288F and W288A variants of GlxA to assess the role of Trp288 on the morphology, maturation, spectroscopic and enzymatic properties. Our findings point towards a salient role for Trp288 in the kinetics of copper loading and maturation of GlxA, with its presence essential for stabilising the metalloradical site required for coupling catalytic activity and morphological development.
Item Type: | Article |
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Uncontrolled Keywords: | Radical-copper oxidase post translational modification EPR spectroscopy free radicals |
Subjects: | Q Science > QD Chemistry Q Science > QH Natural history > QH301 Biology |
Divisions: | Faculty of Science and Health Faculty of Science and Health > Life Sciences, School of |
SWORD Depositor: | Unnamed user with email elements@essex.ac.uk |
Depositing User: | Unnamed user with email elements@essex.ac.uk |
Date Deposited: | 27 Jan 2017 16:17 |
Last Modified: | 30 Oct 2024 20:26 |
URI: | http://repository.essex.ac.uk/id/eprint/18926 |
Available files
Filename: BCJ20160968.full.pdf