Purified I kappa B-beta is inactivated upon dephosphorylation.
In uninduced cells, the NF-kappa B transcription factor resides in the cytoplasm in complex with an inhibitory protein, I kappa B. I kappa B is a specific inhibitor of DNA binding and apparently prevents nuclear uptake of NF-kappa B. Stimulation of cells, for instance with the cytokine tumor necrosis factor, releases I kappa B and allows nuclear translocation and DNA binding of NF-kappa B to regulatory DNA sequences in many genes. We recently reported on the purification of a major form of I kappa B, referred to as I kappa B-alpha, with a molecular size of 37 kDa. Here, we purified and characterized I kappa B-beta, a chromatographically distinct second form of I kappa B. I kappa B-beta has a size of 43 kDa and, as I kappa B-alpha, an acidic isoelectric point between 4.8 and 5.0. Both forms of I kappa B were inactivated by a treatment with protein kinases A and C in vitro. In contrast to I kappa B-alpha, I kappa B-beta lost its inhibiting activity upon a treatment with phosphatase. Phosphatase treatment also released active NF-kappa B from its inactive complex with I kappa B-beta suggesting that the activation of NF-kappa B in intact cells might not only rely on phosphate transfer onto I kappa B but also on phosphate removal from one form of I kappa B.
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Laboratory for Molecular Biology, Gene Center, Martinsried, Federal Republic of Germany.
Kerr
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Schreck
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Verma
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Baeuerle
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D002853:Chromatography, Liquid; D004247:DNA / Q000235:genetics / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D007526:Isoelectric Point; D016328:NF-kappa B / Q000378:metabolism*; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D010766:Phosphorylation; D010920:Placenta / Q000378:metabolism; D011493:Protein Kinase C / Q000494:pharmacology; D011494:Protein Kinases / Q000494:pharmacology
D016328:NF-kappa B; D004247:DNA; D011494:Protein Kinases; D011493:Protein Kinase C; D010744:Phosphoric Monoester Hydrolases