Phosphorylation at Ser²? in the ATP-binding site of Ca²?/calmodulin-dependent kinase II as a mechanism for switching off the kinase activity.

ABSTRACT: CaMKII (Ca²?/calmodulin-dependent kinase II) is a serine/threonine phosphotransferase that is capable of long-term retention of activity due to autophosphorylation at a specific threonine residue within each subunit of its oligomeric structure. The ? isoform of CaMKII is a significant regulator of vascular contractility. Here, we show that phosphorylation of CaMKII ? at Ser²?, a residue located within the ATP-binding site, terminates the sustained activity of the enzyme. To test the physiological importance of phosphorylation at Ser²?, we generated a phosphospecific Ser²? antibody and demonstrated an increase in Ser²? phosphorylation upon depolarization and contraction of blood vessels. To determine if the phosphorylation of Ser²? affects the kinase activity, we mutated Ser²? to alanine or aspartic acid. The S26D mutation mimicking the phosphorylated state of CaMKII causes a dramatic decrease in Thr²?? autophosphorylation levels and greatly reduces the catalytic activity towards an exogenous substrate (autocamtide-3), whereas the S26A mutation has no effect. These data combined with molecular modelling indicate that a negative charge at Ser²? of CaMKII ? inhibits the catalytic activity of the enzyme towards its autophosphorylation site at Thr²?? most probably by blocking ATP binding. We propose that Ser²? phosphorylation constitutes an important mechanism for switching off CaMKII activity.

SUBMITTER: Yilmaz M 

PROVIDER: S-EPMC3566533 | biostudies-literature | 2013 Feb

REPOSITORIES: biostudies-literature