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

Publication Type:

Journal Article

Authors:

Messens,J.; Martins,J.C.; Zegers,I.; Van,Belle K.; Brosens,E.; Wyns,L.

Source:

J. Chromatogr. B Analyt. Technol. Biomed. Life Sci., Volume 790, Issue 1-2, p.217 - 227 (2003)

URL:

PM:12767334

Keywords:

Arsenite Transporting ATPases; Crystallography; X-Ray; Electrophoresis; Polyacrylamide Gel; Ion Pumps; chemistry; isolation & purification; Kinetics; Models; Molecular; Multienzyme Complexes; Nuclear Magnetic Resonance; Biomolecular; Oxidation-Reduction; Prot

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

Notes:

DA - 20030527
IS - 1570-0232 (Print)
LA - eng
PT - Journal Article
PT - Research Support, Non-U.S. Gov't
RN - 0 (Ion Pumps)
RN - 0 (Multienzyme Complexes)
RN - EC 3.6.3.16 (Arsenite Transporting ATPases)
SB - IM