{"title":"Recent developments in Phos-tag electrophoresis for the analysis of phosphoproteins in proteomics","authors":"H. Hirano, Jun Shirakawa","doi":"10.1080/14789450.2022.2052850","DOIUrl":null,"url":null,"abstract":"ABSTRACT Introduction Phosphate-binding tag (Phos-tag) sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) is an important development capable of analyzing the phosphorylation state of proteins. Conventionally, proteins were separated via SDS-PAGE and Phos-tag SDS-PAGE that use different gels to identify phosphorylated proteins. However, it was often difficult to compare the electrophoretic mobility of the proteins in the different gels used. The recently developed Phos-tag diagonal electrophoresis has been able to solve this problem. It can indicate the SDS-PAGE and Phos-tag SDS-PAGE patterns on a single gel; therefore, phosphorylated proteins can be distinguished easily from non-phosphorylated proteins. Areas covered This review assesses the importance of Phos-tag electrophoresis, which enables the analysis of protein phosphorylation states, in the field of proteomics. Additionally, this review describes the significance and actual experimental technique of Phos-tag diagonal electrophoresis, which was recently developed to overcome the drawbacks of Phos-tag SDS-PAGE. Expert opinion Although shotgun analysis of proteins allows detecting many phosphorylation sites, it is challenging to clarify the differences in the phosphorylation states of protein molecules using this technique. Therefore, Phos-tag SDS-PAGE is frequently used to determine the phosphorylation state of proteins. This technique has become more powerful with the recent development of Phos-tag diagonal electrophoresis. Abbreviations: BIS, N,N’-methylenebis(acrylamide); CBB, Coomassie brilliant blue R250; ESI, electrospray ionization; hnRNP, heterogeneous ribonucleoprotein K; LTQ–Orbitrap, Linear trap quadrupole–Orbitrap; LC, liquid chromatography; MS, mass spectrometry; MALDI, matrix-assisted laser desorption ionization; Phos-tag, phosphate-binding tag [1,3-bis [bis (pyridine-2-ylmethyl) amino] propane-2-olate]; SDS-PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis; TOF, time of flight; 2D-DIGE, fluorescence-labeled two-dimensional difference gel electrophoresis; 2-DE, two-dimensional gel electrophoresis","PeriodicalId":50463,"journal":{"name":"Expert Review of Proteomics","volume":null,"pages":null},"PeriodicalIF":3.8000,"publicationDate":"2022-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"2","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Expert Review of Proteomics","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1080/14789450.2022.2052850","RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"BIOCHEMICAL RESEARCH METHODS","Score":null,"Total":0}
引用次数: 2
Abstract
ABSTRACT Introduction Phosphate-binding tag (Phos-tag) sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) is an important development capable of analyzing the phosphorylation state of proteins. Conventionally, proteins were separated via SDS-PAGE and Phos-tag SDS-PAGE that use different gels to identify phosphorylated proteins. However, it was often difficult to compare the electrophoretic mobility of the proteins in the different gels used. The recently developed Phos-tag diagonal electrophoresis has been able to solve this problem. It can indicate the SDS-PAGE and Phos-tag SDS-PAGE patterns on a single gel; therefore, phosphorylated proteins can be distinguished easily from non-phosphorylated proteins. Areas covered This review assesses the importance of Phos-tag electrophoresis, which enables the analysis of protein phosphorylation states, in the field of proteomics. Additionally, this review describes the significance and actual experimental technique of Phos-tag diagonal electrophoresis, which was recently developed to overcome the drawbacks of Phos-tag SDS-PAGE. Expert opinion Although shotgun analysis of proteins allows detecting many phosphorylation sites, it is challenging to clarify the differences in the phosphorylation states of protein molecules using this technique. Therefore, Phos-tag SDS-PAGE is frequently used to determine the phosphorylation state of proteins. This technique has become more powerful with the recent development of Phos-tag diagonal electrophoresis. Abbreviations: BIS, N,N’-methylenebis(acrylamide); CBB, Coomassie brilliant blue R250; ESI, electrospray ionization; hnRNP, heterogeneous ribonucleoprotein K; LTQ–Orbitrap, Linear trap quadrupole–Orbitrap; LC, liquid chromatography; MS, mass spectrometry; MALDI, matrix-assisted laser desorption ionization; Phos-tag, phosphate-binding tag [1,3-bis [bis (pyridine-2-ylmethyl) amino] propane-2-olate]; SDS-PAGE, sodium dodecyl sulfate-polyacrylamide gel electrophoresis; TOF, time of flight; 2D-DIGE, fluorescence-labeled two-dimensional difference gel electrophoresis; 2-DE, two-dimensional gel electrophoresis
期刊介绍:
Expert Review of Proteomics (ISSN 1478-9450) seeks to collect together technologies, methods and discoveries from the field of proteomics to advance scientific understanding of the many varied roles protein expression plays in human health and disease.
The journal coverage includes, but is not limited to, overviews of specific technological advances in the development of protein arrays, interaction maps, data archives and biological assays, performance of new technologies and prospects for future drug discovery.
The journal adopts the unique Expert Review article format, offering a complete overview of current thinking in a key technology area, research or clinical practice, augmented by the following sections:
Expert Opinion - a personal view on the most effective or promising strategies and a clear perspective of future prospects within a realistic timescale
Article highlights - an executive summary cutting to the author''s most critical points.