TY - JOUR
T1 - STO-609, a specific inhibitor of the CA2+/calmodulin-dependent protein kinase kinase
AU - Tokumitsu, Hiroshi
AU - Inuzuka, Hiroyuki
AU - Ishikawa, Yumi
AU - Ikeda, Masahiko
AU - Saji, Ikutaro
AU - Kobayashi, Ryoji
PY - 2002/5/3
Y1 - 2002/5/3
N2 - 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.
AB - 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.
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U2 - 10.1074/jbc.M201075200
DO - 10.1074/jbc.M201075200
M3 - Article
C2 - 11867640
AN - SCOPUS:0037013222
VL - 277
SP - 15813
EP - 15818
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 18
ER -