Effects of mutations in active site heme ligands on the spectroscopic and catalytic properties of SoxAX cytochromes

Kilmartin, James R., Bernhardt, Paul V., Dhouib, Rabeb, Hanson, Graeme R., Riley, Mark J. and Kappler, Ulrike (2016) Effects of mutations in active site heme ligands on the spectroscopic and catalytic properties of SoxAX cytochromes. Journal of Inorganic Biochemistry, 162 309-318. doi:10.1016/j.jinorgbio.2016.04.015


Author Kilmartin, James R.
Bernhardt, Paul V.
Dhouib, Rabeb
Hanson, Graeme R.
Riley, Mark J.
Kappler, Ulrike
Title Effects of mutations in active site heme ligands on the spectroscopic and catalytic properties of SoxAX cytochromes
Journal name Journal of Inorganic Biochemistry   Check publisher's open access policy
ISSN 1873-3344
0162-0134
Publication date 2016-04-15
Year available 2016
Sub-type Article (original research)
DOI 10.1016/j.jinorgbio.2016.04.015
Open Access Status Not Open Access
Volume 162
Start page 309
End page 318
Total pages 10
Place of publication Philadelphia, PA, United States
Publisher Elsevier
Collection year 2017
Language eng
Formatted abstract
By attaching a sulfur substrate to a conserved cysteine of the SoxYZ carrier protein SoxAX cytochromes initiate the reaction cycle of the Sox (sulfur oxidation) multienzyme complex, which is the major pathway for microbial reoxidation of sulfur compounds in the environment. Despite their important role in this process, the reaction mechanism of the SoxAX cytochromes has not been fully elucidated. Here we report the effects of several active site mutations on the spectroscopic and enzymatic properties of the type II SoxAX protein from Starkeya novella, which in addition to two heme groups also contains a Cu redox centre. All substituted proteins contained these redox centres except for His231Ala which was unable to bind Cu(II). Substitution of the SoxA active site heme cysteine ligand with histidine resulted in increased microheterogeneity around the SoxA heme as determined by CW-EPR, while a SnSoxAXC236A substituted protein revealed a completely new, nitrogenous SoxA heme ligand. The same novel ligand was present in SnSoxAXH231A CW-EPR spectra, the first time that a ligand switch of the SoxA heme involving a nearby amino acid has been demonstrated. Kinetically, SnSoxAXC236A and SnSoxAXC236H showed reduced turnover, and in assays containing SoxYZ these mutants retained only ~25% of the wildtype activity. Together, these data indicate that the Cu redox centre can mediate a low level of activity, and that a possible ligand switch can occur during catalysis. It also appears that the SoxA heme cysteine ligand (and possibly the low redox potential) is important for an efficient reaction with SnSoxYZ/thiosulfate.
Keyword Cytochromes
EPR
Heme thiolate proteins
Redox properties
SoxAX
Thiosulfate oxidation
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: HERDC Pre-Audit
School of Chemistry and Molecular Biosciences
Centre for Advanced Imaging Publications
 
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