STO-609, a specific inhibitor of the CA2+/calmodulin-dependent protein kinase kinase

Hiroshi Tokumitsu, Hiroyuki Inuzuka, Yumi Ishikawa, Masahiko Ikeda, Ikutaro Saji, Ryoji Kobayashi

Research output: Contribution to journalArticlepeer-review

212 Citations (Scopus)

Abstract

STO-609, a selective inhibitor of Ca2+/calmodulin-dependent protein kinase kinase (CaM-KK) was synthesized, and its inhibitory properties were investigated both in vitro and in vivo. STO-609 inhibits the activities of recombinant CaM-KKα and CaM-KKβ isoforms, with Ki values of 80 and 15 ng/ml, respectively, and also inhibits their autophosphorylation activities. Comparison of the inhibitory potency of the compound against various protein kinases revealed that STO-609 is highly selective for CaM-KK without any significant effect on the downstream CaM kinases (CaM-KI and -IV), and the IC50 value of the compound against CaM-KII is ∼10 μg/ ml. STO-609 inhibits constitutively active CaM-KKα (glutathione S-transferase (GST)-CaM-KK-(84-434)) as well as the wild-type enzyme. Kinetic analysis indicates that the compound is a competitive inhibitor of ATP. In transfected HeLa cells, STO-609 suppresses the Ca2+-induced activation of CaM-KIV in a dose-dependent manner. In agreement with this observation, the inhibitor significantly reduces the endogenous activity of CaM-KK in SH-SY5Y neuroblastoma cells at a concentration of 1 μg/ml (∼80% inhibitory rate). Taken together, these results indicate that STO-609 is a selective and cell-permeable inhibitor of CaM-KK and that it may be a useful tool for evaluating the physiological significance of the CaM-KK-mediated pathway in vivo as well as in vitro.

Original languageEnglish
Pages (from-to)15813-15818
Number of pages6
JournalJournal of Biological Chemistry
Volume277
Issue number18
DOIs
Publication statusPublished - May 3 2002
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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