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Merck

Manganese Superoxide Dismutase Activity Assay in the Yeast Saccharomyces cerevisiae.

Bio-protocol (2021-03-05)
Louise Thines, Pierre Morsomme
RESUMO

Superoxide dismutases (SODs) act as a primary defence against reactive oxygen species (ROS) by converting superoxide anion radicals (O2 -) into molecular oxygen (O2) and hydrogen peroxide (H2O2). Members of this enzyme family include CuZnSODs, MnSODs, FeSODs, and NiSODs, depending on the nature of the cofactor that is required for proper activity. Most eukaryotes, including yeast, possess CuZnSOD and MnSOD. This protocol aims at assessing the activity of the yeast Saccharomyces cerevisiae MnSOD Sod2p from cellular extracts using nitroblue tetrazolium staining. This method can be used to estimate the cellular bioavailability of Mn2+ as well as to evaluate the redox state of the cell.

MATERIAIS
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Sigma-Aldrich
Cloreto de sódio, ACS reagent, ≥99.0%
Sigma-Aldrich
Glicerol, for molecular biology, ≥99.0%
Sigma-Aldrich
N,N,N′,N′-tetrametiletilenodiamina, BioReagent, suitable for electrophoresis, ≥99.0%
Sigma-Aldrich
Glicina, BioUltra, Molecular Biology, ≥99.0% (NT)
Sigma-Aldrich
Ácido clorídrico, 36.5-38.0%, BioReagent, Molecular Biology
Sigma-Aldrich
Ácido etilenoglicol-bis(2-aminoetiléter)-N,N,N′,N′-tetracético, Molecular Biology, ≥97.0%
Sigma-Aldrich
Fosfato de sódio, BioReagent, Molecular Biology, anhydrous, ≥98%
Supelco
Ácido bicinconínico
Sigma-Aldrich
Triton X-100, laboratory grade
Sigma-Aldrich
Ácido etilenodiamino tetra-acético, ACS reagent, 99.0-101.0%
Sigma-Aldrich
Bromophenol Blue, titration: suitable
Sigma-Aldrich
Cloreto de nitrotetrazólio azul, ≥90.0% (HPLC)
Sigma-Aldrich
Fosfato de sódio, BioXtra, ≥99.0%