Hello,
I want to study the effect of in-vitro S-nitrosylation (using GSNO) on the enzymatic activity of a recombinant (expressed and purified) redox enzyme.
Can anyone provide me the detailed protocol for the GSNO treatment of purified enzyme.
Thank you,
Vandan
Hello Vandan, S-nitrosocysteine and S-nitrosoglutathione are synthesized by mixing equimolar volumes of NaNO2/1 mM EDTA in deionized water with thiol in 0.5 M HCl. For synthesis of S-nitroso-4-
mercaptophenylacetic acid, the thiol is dissolved in isopropanol
containing 0.5 M HCl. After mixing with NaNO2, PASNO is immediately neutralized with NaOH and diluted further with HEN buffer [250 mM Hepes, 1 mM EDTA, 0.1 mM neocuproine (pH 7.7)].
Please go through the attached documents. For detailed protocol, you will be led to supplementary information when you open the second document. I hope these serve the purpose.
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