Academic Journal
Measurement of S-glutathionylated proteins by HPLC
| العنوان: | Measurement of S-glutathionylated proteins by HPLC |
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| المؤلفون: | Giustarini, Daniela, Milzani, Aldo, Dalle-Donne, Isabella, Rossi, Ranieri |
| المساهمون: | D. Giustarini, A. Milzani, I. Dalle-Donne, R. Rossi |
| بيانات النشر: | SPRINGER WIEN |
| سنة النشر: | 2022 |
| المجموعة: | The University of Milan: Archivio Istituzionale della Ricerca (AIR) |
| مصطلحات موضوعية: | Cell, HPLC, Protocol, Quantification, S-glutathionylation, Whole blood, Settore BIO/14 - Farmacologia, Settore BIO/06 - Anatomia Comparata e Citologia |
| الوصف: | S-glutathionylated proteins (GSSP), i.e., protein-mixed disulfides with glutathione (GSH), are considered a suitable biomarker of oxidative stress. In fact, they occur within cells at low level and their concentration increases markedly under pro-oxidant conditions. Plasma is something different, since it is physiologically rich in S-thiolated proteins (RSSP), i.e., protein-mixed disulfides with various types of low molecular mass thiols (LMM-SH). However, albumin, which is largely the most abundant plasma protein, possesses a cysteine residue at position 34 that is mostly reduced (about 60%) under physiological conditions, but easily involved in the formation of additional RSSP in the presence of oxidants. The quantification of GSSP requires special attention to sample handling, since their level can be overestimated as a result of artefactual oxidation of GSH. We have developed the present protocol to avoid this methodological problem. Samples should be treated as soon as possible after their collection with the alkylating agent N-ethylmaleimide that masks -SH groups and prevents their oxidation. The GSH released from mixed disulfides by reduction with dithiothreitol is then labeled with the fluorescent probe monobromobimane and quantified by HPLC. The method can be applied to many different biological samples, comprising blood components, red blood cell plasma membrane, cultured cells, and solid organs from animal models. |
| نوع الوثيقة: | article in journal/newspaper |
| اللغة: | English |
| Relation: | info:eu-repo/semantics/altIdentifier/pmid/34129091; info:eu-repo/semantics/altIdentifier/wos/WOS:000661791500001; volume:54; issue:4; firstpage:675; lastpage:686; numberofpages:12; journal:AMINO ACIDS; https://hdl.handle.net/2434/969558; info:eu-repo/semantics/altIdentifier/scopus/2-s2.0-85107924829 |
| DOI: | 10.1007/s00726-021-03015-6 |
| الاتاحة: | https://hdl.handle.net/2434/969558 https://doi.org/10.1007/s00726-021-03015-6 |
| Rights: | info:eu-repo/semantics/openAccess |
| رقم الانضمام: | edsbas.B592F002 |
| قاعدة البيانات: | BASE |
| DOI: | 10.1007/s00726-021-03015-6 |
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