Source:http://linkedlifedata.com/resource/pubmed/id/10823818
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rdf:type | |
lifeskim:mentions | |
pubmed:issue |
34
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pubmed:dateCreated |
2000-9-25
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pubmed:abstractText |
Nuclear factor kappa B (NF-kappaB) is a ubiquitous, inducible transcription factor that regulates the initiation and progression of immune and inflammatory stress responses. NF-kappaB activation depends on phosphorylation and degradation of its inhibitor protein, IkappaB, initiated by an IkappaB kinase (IKK) complex. This IKK complex includes a catalytic heterodimer composed of IkappaB kinase 1 (IKK1) and IkappaB kinase 2 (IKK2) as well as a regulatory adaptor subunit, NF-kappaB essential modulator. To better understand the role of IKKs in NF-kappaB activation, we have cloned, expressed, purified, and characterized the physiological isoform, the rhIKK1/rhIKK2 heterodimer. We compared its kinetic properties with those of the homodimers rhIKK1 and rhIKK2 and a constitutively active rhIKK2 (S177E, S181E) mutant. We demonstrate activation of these recombinantly expressed IKKs by phosphorylation during expression in a baculoviral system. The K(m) values for ATP and IkappaBalpha peptide for the rhIKK1/rhIKK2 heterodimer are 0.63 and 0.60 micrometer, respectively, which are comparable to those of the IKK2 homodimer. However, the purified rhIKK1/rhIKK2 heterodimer exhibits the highest catalytic efficiency (k(cat)/K(m)) of 47.50 h(-1) micrometer(-1) using an IkappaBalpha peptide substrate compared with any of the other IKK isoforms, including rhIKK2 (17.44 h(-1) micrometer(-1)), its mutant rhIKK2 (S177E, S181E, 1.18 h(-1) micrometer(-1)), or rhIKK1 (0.02 h(-1) micrometer(-1)). Kinetic analysis also indicates that, although both products of the kinase reaction, ADP and a phosphorylated IkappaBalpha peptide, exhibited competitive inhibitory kinetics, only ADP with the low K(i) of 0.77 micrometer may play a physiological role in regulation of the enzyme activity.
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pubmed:language |
eng
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pubmed:journal | |
pubmed:citationSubset |
IM
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pubmed:chemical |
http://linkedlifedata.com/resource/pubmed/chemical/CHUK protein, human,
http://linkedlifedata.com/resource/pubmed/chemical/I-kappa B Kinase,
http://linkedlifedata.com/resource/pubmed/chemical/IKBKB protein, human,
http://linkedlifedata.com/resource/pubmed/chemical/IKBKE protein, human,
http://linkedlifedata.com/resource/pubmed/chemical/Protein-Serine-Threonine Kinases,
http://linkedlifedata.com/resource/pubmed/chemical/Recombinant Proteins
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pubmed:status |
MEDLINE
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pubmed:month |
Aug
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pubmed:issn |
0021-9258
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pubmed:author |
pubmed-author:BoddupalliHH,
pubmed-author:CouchN WNW,
pubmed-author:Diaz-CollierJ AJA,
pubmed-author:HallTT,
pubmed-author:HauserS DSD,
pubmed-author:HoodB LBL,
pubmed-author:HuynhQ KQK,
pubmed-author:KilpatrickB FBF,
pubmed-author:KoboldtC MCM,
pubmed-author:PierceJ LJL,
pubmed-author:ReitzB ABA,
pubmed-author:RouwS ASA,
pubmed-author:SommersCC,
pubmed-author:TrippC SCS,
pubmed-author:WeinbergR ARA
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pubmed:issnType |
Print
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pubmed:day |
25
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pubmed:volume |
275
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pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
25883-91
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pubmed:dateRevised |
2009-11-19
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pubmed:meshHeading |
pubmed-meshheading:10823818-Catalysis,
pubmed-meshheading:10823818-Dimerization,
pubmed-meshheading:10823818-Electrophoresis, Polyacrylamide Gel,
pubmed-meshheading:10823818-Humans,
pubmed-meshheading:10823818-I-kappa B Kinase,
pubmed-meshheading:10823818-Kinetics,
pubmed-meshheading:10823818-Molecular Weight,
pubmed-meshheading:10823818-Protein-Serine-Threonine Kinases,
pubmed-meshheading:10823818-Recombinant Proteins
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pubmed:year |
2000
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pubmed:articleTitle |
Characterization of the recombinant IKK1/IKK2 heterodimer. Mechanisms regulating kinase activity.
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pubmed:affiliation |
Discovery Research, G. D. Searle and Company, the Monsanto Life Science Company, St. Louis, Missouri 63167, USA. quang.k.huynh@monsanto.com
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pubmed:publicationType |
Journal Article
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