An Introduction to Phosphorylation Site Specific Antibodies
Protein phosphorylation is a prevalent, versatile and widely studied reversible Post Translational Modification (PTM) and occurs principally on three amino acids: serine, threonine or tyrosine. It is mediated by kinases and phosphatases and plays a critical role in the regulation of many cellular processes including cell cycle, signal transduction pathways and apoptosis.
Phosphorylation site-specific antibodies which can discriminate between the phosphorylated and non-phosphorylated forms have emerged as reliable tools to enable quantitative and qualitative detection of phosphorylated proteins and can be used for Western blotting, Immunohistochemistry and Immunoprecipitation, ELISA analysis and flow cytometry.
Anti-phosphorylated CDK (pY15) Antibody
Cyclin-dependent kinase-2 (CDK2) is a serine/threonine kinase that plays an essential role in coordinating cell cycle. Monomeric CDK2 is inactive, and requires interaction with cyclins for functional activation. The CDK2/Cyclin E complexes are involved in controlling the G1/S transition and initiation of DNA synthesis. Whereas, the CDK2/Cyclin A complexes are involved in the control of progression through S phase and mitosis. The CDK2 is the catalytic subunit of the heterodimeric complexes.
The CDK2 kinase activity is tightly regulated and depends on the phosphorylation state at 2 major phosphorylation sites, Thr14/Tyr15 (T14/Y15) and Thr160 (T160). CDK2 activation occurs when CDK2 is dephosphorylated at the inhibitory site T14/Y15 and phosphorylated at activating site T160.
CDK2 contains an N-terminal lobe structure and a C-terminal lobe structure which sandwich the active site. The N-terminal lobe contains a glycine-rich inhibitory element (G-loop) and a unique major helix (C-helix). T14/Y15 residues are located in the flexible G-loop that forms the roof of the ATP-binding site and is important for ATP interaction. Structural studies of pY15/pT160 CDK2/Cyclin/ATP complex show that phosphorylation at Y15 perturbs protein substrate binding affinity due to steric hindrance. This affects correct ATP alignment and binding at the catalytic site.
Wee1 and Myt1 kinases phosphorylate CDK2 at Y15 and inhibit its activity by preventing ATP binding. The phosphorylation at Y15 of CDK2 is independent of cyclin interaction and precedes the activating CDK2 T160 phosphorylation by CDK-activating kinase complex (CAK), as the CDK2 T160 phosphorylation is dependent on cyclin binding. Phosphorylation at CDK2 T14/Y15 has been shown to stabilize Cyclin E and prevent its degradation.
The subsequent dephosphorylation of CDK2 at T14/Y15 by the cell division cycle 25 (CDC25) phosphatase promotes CDK2 activation and is the rate-limiting step of the CDK2/Cyclin complex activation process. The CDK2 inhibitory phosphorylation plays an important role during replication stress signaling and in maintaining genome integrity.
Antibodies to purchase via DISCOVERY Antibodies
Peptide and Antibody bundles to purchase via DISCOVERY Antibodies
Featuring: Anti-CDK2 antibody and CDK2 peptide
Featuring: Anti-phosphorylated CDK2 (pY15) antibody, Phosphorylated CDK2 (pY15) peptide and CDK2 peptide