Analysis of protein kinases by Phos-tag SDS-PAGE

Protein kinases regulate almost all biological processes including cell proliferation, differentiation, apoptosis, and gene expression. Dysregulation of protein phosphorylation caused by abnormal activity and expression of protein kinases results in the onset of various diseases such as cancer and metabolic syndromes. The activities of a large number of protein kinases are regulated by phosphorylation. Therefore, analysis of the phosphorylation status of protein kinases is important for elucidation of biological phenomena and the pathogenesis of diseases. To investigate protein phosphorylation, phosphate-binding tag molecule "Phos-tag" was developed. In addition, various techniques and tools using Phos-tag such as Phos-tag SDS-PAGE, have been developed for analysis and profiling of protein phosphorylation. Here, we describe the methods and analytical techniques that use Phos-tag for investigation of protein kinase phosphorylation and the applications of phosphorylation analysis. SIGNIFICANCE: Protein kinases play pivotal roles in regulating many biological processes and pathogenesis of diseases. Determination of phosphorylation status of protein kinases can provide the essential information for their activation. This review provides analytical techniques for analysis of phosphorylation status of protein kinases by Phos-tag SDS-PAGE. We believe that this review would help readers to study in kinomics research.