Purification of an oxidation-sensitive enzyme, pI258 arsenate reductase from Staphylococcus aureus.

TitlePurification of an oxidation-sensitive enzyme, pI258 arsenate reductase from Staphylococcus aureus.
Publication TypeJournal Article
Year of Publication2003
AuthorsMessens, J., J. C. Martins, I. Zegers, K. Van Belle, E. Brosens, and L. Wyns
JournalJ Chromatogr B Analyt Technol Biomed Life Sci
Volume790
Issue1-2
Pagination217-27
Date Published2003 Jun 25
ISSN1570-0232
KeywordsArsenite Transporting ATPases, Crystallography, X-Ray, Electrophoresis, Polyacrylamide Gel, Ion Pumps, Kinetics, Models, Molecular, Multienzyme Complexes, Nuclear Magnetic Resonance, Biomolecular, Oxidation-Reduction, Protein Conformation, Staphylococcus aureus
Abstract

Arsenate reductase (ArsC) from Staphylococcus aureus pI258 is extremely sensitive to oxidative inactivation. The presence of oxidized ArsC forms was not that critical for NMR, but kinetics and crystallization required an extra reversed-phase purification to increase sample homogeneity. The salt ions observed in the X-ray electron density of ArsC were investigated. Carbonate was found to have the lowest dissociation constant for activation (K(a)=1.1 mM) and potassium was stabilizing ArsC (DeltaT(m)=+6.2 degrees C). Also due to the use of these salt ions, the final yield of the purification had improved with a factor of four, i.e. 73 mg/l culture.

Alternate JournalJ. Chromatogr. B Analyt. Technol. Biomed. Life Sci.
PubMed ID12767334
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