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Title 

Cobrotoxin inhibits NF-κB activation and target gene expression through reaction with NF-κB signal molecules

Authors 

M H ParkH S SongK H KimD J SonS H LeeDo Young YoonY KimI Y ParkS SongB Y HwangJ K JungJ T Hong

Publisher 

American Chemical Society

Issue Date 

2005

Citation 

Biochemistry, vol. 44, no. 23, pp. 8326-8336

Keywords 

activation energybinding energybiomaterialscarbotoxinsenzyme inhibitioncobrotoxinI kappa B kinaseI kappa B kinase alphaI kappa B kinase betaimmunoglobulin enhancer binding protein

Abstract 

Cobrotoxin is known to bind with cysteine residues of biological molecules such as nicotine acetylcholine receptor. Cobrotoxin may modify IKKs and p50 through protein-protein interaction since cysteine residues are present in the kinase domains of IKKα and IKKβ and in the p50 of NF-κB. Our surface plasmon resonance analysis showed that cobrotoxin directly binds to p50 (Kd = 1.54 × 10-5 M), IKKα (Kd = 3.94 × 10-9 M) and IKKβ (Kd = 3.4 × 10-8 M) with high binding affinity. Moreover, these protein-protein interactions suppressed the lipopolysaccharide (LPS, 1 μg/mL)- and the sodium nitroprusside (SNP, 200 μM)-induced DNA binding activity of NF-κB and NF-κB-dependent luciferase activity in astrocytes and Raw 264.7 macrophages. These inhibitory effects were correlated with the inhibition of IκB release and p50 translocation. Inhibition of NF-κB by cobrotoxin resulted in reductions in the LPS-induced expressions of COX-2, iNOS, cPLA 2, IL-4, and TNF-α in astrocytes and in COX-2 expression induced by SNP, LPS, and TNF-α in astrocytes. Moreover, these inhibitory effects of cobrotoxin were reversed by adding reducing agents, dithiothreitol and glutathione. In addition, cobrotoxin did not have any inhibitory effect on NF-κB activity in cells carrying mutant p50 (C62S), IKKα (C178A), and IKKβ (C179A), with the exception of IKKβ (K44A) mutant plasmid. Confocal microscopic analysis showed that cobrotoxin is uptaken into the nucleus of cells. These results demonstrate that cobrotoxin directly binds to the sulfhydryl groups of p50 and IKKs, and that this results in reduced IκB release and the translocation of p50, thereby inhibiting the activation of NF-κB.

ISSN 

0006-2960

Link 

http://dx.doi.org/10.1021/bi050156h

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2017-04-19


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