Original Article

Inhibitory effect and its kinetic analysis of tyrphostin AG1478 on recombinant human protein kinase CK2 holoenzyme.

Authors: Xin-Guang LIU, Nian-Ci LIANG


AIM: To study the direct effect of tyrphostin AG1478
[4-(3-chloroanilino)-6,7-dimethoxyquinazoline] on recombinant human protein
kinase CK2 holoenzyme and its kinetics.
METHODS: Recombinant human protein kinase CK2 alpha and beta subunits were mixed
at equal molar ratio and CK2 holoenzyme were reconstituted. The CK2 activity was
assayed by detecting incorporation of [gamma-32P]ATP or [gamma-32P]GTP into
substrates in various conditions.
RESULTS: These results demonstrated that the recombinant human CK2 was a second
messengers (Ca2+, cAMP, and cGMP)-independent protein kinase, the
characterization and function of the reconstituted holoenzyme were consistent
with those of native CK2. AG1478 strongly inhibited the holoenzyme activity of
recombinant human protein kinase CK2 with IC50 of 25.9 micromol/L, the inhibition
is very close to that of N-(2-aminoethyl)-5-chloronaphthalene-1-sulfonamide (A3),
but less potent than that of 5,6-dichloro-1- beta-D-ribofuranosylbenzimidazole
(DRB), known as CK2 special inhibitors with IC50 of 25.5 micromol/L and 10.4
micromol/L respectively. Kinetic studies of AG1478 on recombinant human CK2
showed that inhibitions were competitive with both GTP and casein, thus AG1478
was as bisubstrate inhibitor.
CONCLUSION: The present study indicates that AG1478 is not only an effective
inhibitor of protein tyrosine kinases of epidermal growth factor receptor (EGFR),
but also a novel potent inhibitor of protein kinase CK2. The recombinant human
protein kinase CK2 might be used as a molecular target for simpler screening and
development of more effective inhibitors of CK2.

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