Multimodal analysis of metals in copper-zinc superoxide dismutase isoforms separated on electrophoresis gels.
It has been suggested that copper-zinc superoxide dismutase (CuZnSOD) isoforms of distinct isoelectric point (pI) could result from differences in their metallation state. Our aim was then to develop and validate analytical methods for the determination and understanding of metallation states in human CuZnSOD isoforms. To avoid metal losses during sample preparation steps, CuZnSOD isoforms were separated according to their pI using non-denaturing isoelectric focusing (IEF) gel electrophoresis. Metal quantification was directly performed in-gel. Cu/Zn ratios of CuZnSOD isoforms were quantified by Particle-Induced X-ray Emission (PIXE) and Laser Ablation-Inductively Coupled Plasma-Mass Spectrometry (LA-ICP-MS). Cu/Zn ratios were measured close to the value of 1 as expected from the known stoichiometry of CuZnSOD with slight, but statistically significant, differences between acidic and basic isoforms. Overall, this study demonstrates that metal quantification can be performed directly on metalloproteins separated on electrophoresis gels.
Chevreux
Sylviane
S
Groupe d'Imagerie Chimique Cellulaire et Spéciation, Laboratoire de Chimie Nucléaire Analytique et Bioenvironnementale (CNAB), CNRS UMR5084, Université de Bordeaux 1, chemin du Solarium, 33175 Gradignan Cedex, France.
Roudeau
Stéphane
S
Fraysse
Aurélien
A
Carmona
Asunción
A
Devès
Guillaume
G
Solari
Pier Lorenzo
PL
Mounicou
Sandra
S
Lobinski
Ryszard
R
Ortega
Richard
R
D004586:Electrophoresis / Q000379:methods*; D006801:Humans; D007525:Isoelectric Focusing; D007527:Isoenzymes / Q000302:isolation & purification*; D013058:Mass Spectrometry; D013482:Superoxide Dismutase / Q000302:isolation & purification*
D007527:Isoenzymes; D013482:Superoxide Dismutase