Identification of direct tyrosine kinase substrates based on protein kinase assay-linked phosphoproteomics.

ABSTRACT: Protein kinases are implicated in multiple diseases such as cancer, diabetes, cardiovascular diseases, and central nervous system disorders. Identification of kinase substrates is critical to dissecting signaling pathways and to understanding disease pathologies. However, methods and techniques used to identify bona fide kinase substrates have remained elusive. Here we describe a proteomic strategy suitable for identifying kinase specificity and direct substrates in high throughput. This approach includes an in vitro kinase assay-based substrate screening and an endogenous kinase dependent phosphorylation profiling. In the in vitro kinase reaction route, a pool of formerly phosphorylated proteins is directly extracted from whole cell extracts, dephosphorylated by phosphatase treatment, aft

SUBMITTER: Xue L 

PROVIDER: S-EPMC3790304 | biostudies-literature | 2013 Oct

REPOSITORIES: biostudies-literature