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Measurement of mixed disulfides including glutathionylated proteins

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posted on 2023-06-07, 15:34 authored by Raffaella Priora, Lucia Coppo, Sonia Salzano, Paolo Di Simplicio, Pietro Ghezzi
Mixed disulfides between protein cysteines and low-molecular-weight thiol cysteine or glutathione lead to the formation of cysteinylated proteins or glutathionylated proteins. These types of posttranslational modification are of great importance in the so-called redox regulation, by which changes in the redox state of the cell regulate a number of biochemical processes. We describe the methods for quantitatively measuring the various redox states of cellular thiols including protein cysteines and these mixed disulfides. These include spectrophotometric methods, which do not distinguish between protein-cysteine and protein-glutathione disulfides, and HPLC methods that make such distinction. Finally, we report a method for labeling proteins susceptible to glutathionylation with biotin, to allow their visualization by Western blot after electrophoretic separation, which is used to identify proteins undergoing this posttranslational modification.

History

Publication status

  • Published

File Version

  • Published version

Journal

Methods in Enzymology

Publisher

Elsevier/Academic Press

Volume

473

Page range

149-159

Pages

359.0

Book title

Thiol redox transitions in cell signaling. Part A, Chemistry and biochemistry of low molecular weight and protein thiols

Place of publication

Amsterdam; Boston

ISBN

9780123813459

Series

Methods in enzymology

Department affiliated with

  • Clinical and Experimental Medicine Publications

Full text available

  • No

Peer reviewed?

  • Yes

Editors

Lester Packer, Enrique Cadenas

Legacy Posted Date

2010-10-25

First Compliant Deposit (FCD) Date

2017-12-01

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