Abstract:
:MAPK-activated protein kinase-2 (MAPKAPK2) regulates the synthesis of tumor necrosis factor and other cytokines and is a potential drug target for inflammatory diseases. Five protein constructs were produced in 4-10mg quantities per liter of culture media using baculovirus-infected insect cells and characterized for kinase activity, thermal stability, and ligand-binding affinity. Compared to construct 1-370, removal of the C-terminal autoinhibitory peptide in 1-338 resulted in a destabilized but partially active nonphosphorylated enzyme; phosphorylation of 1-338 by p38alpha further increased activity 12-fold. A putative constitutively active mutant, 1-370/T222E/T334E, was 6.3-fold less active than phosphorylated 1-370. ThermoFluor, an equilibrium ligand-binding assay, was used to measure nucleotide analogue affinity for various constructs. Binding of phosphorylated nucleotides was Mg(2+)-dependent. Residues 1-40 were required for high-affinity binding of ADP, ATPgammaS, staurosporine, and K252a. A mutation M138A rendered 1-370 susceptible to p38-inhibitors SB-203580 and SB-202190 with IC50 values of 17.4 and 14.1 microM, respectively. Taken together, these studies provide information on the mechanism of ligand-binding to MAPKAPK2 that can be used in the search for selective small-molecule inhibitors.
journal_name
Arch Biochem Biophysjournal_title
Archives of biochemistry and biophysicsauthors
Kervinen J,Ma H,Bayoumy S,Schubert C,Milligan C,Lewandowski F,Moriarty K,Desjarlais RL,Ramachandren K,Wang H,Harris CA,Grasberger B,Todd M,Springer BA,Deckman Idoi
10.1016/j.abb.2006.03.018subject
Has Abstractpub_date
2006-05-15 00:00:00pages
47-56issue
1-2eissn
0003-9861issn
1096-0384pii
S0003-9861(06)00112-3journal_volume
449pub_type
杂志文章abstract::The in vivo turnover of several rat liver microsomal proteins was studied using techniques designed to maximize antibody recognition specificity and minimize reutilization of radioactive labels. The kinetics of degradation of seven cytochrome P-450 isozymes, NADPH-cytochrome P-450 reductase, and epoxide hydrolase were...
journal_title:Archives of biochemistry and biophysics
pub_type: 杂志文章
doi:10.1016/0003-9861(84)90257-1
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abstract::To confirm and extend our previous microspectrophotometric observations of 30-week-old male Long-Evans Cinnamon (LEC) rats, an animal model of human Wilson's disease, we analyzed the porphyrin patterns of the organs, urine, and plasma of LEC rats. Abnormal accumulation of porphyrins, especially highly carboxylated por...
journal_title:Archives of biochemistry and biophysics
pub_type: 杂志文章
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journal_title:Archives of biochemistry and biophysics
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journal_title:Archives of biochemistry and biophysics
pub_type: 杂志文章
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journal_title:Archives of biochemistry and biophysics
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journal_title:Archives of biochemistry and biophysics
pub_type: 杂志文章
doi:10.1006/abbi.1994.1407
更新日期:1994-10-01 00:00:00
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journal_title:Archives of biochemistry and biophysics
pub_type: 杂志文章
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journal_title:Archives of biochemistry and biophysics
pub_type: 杂志文章
doi:10.1006/abbi.1993.1332
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journal_title:Archives of biochemistry and biophysics
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pub_type: 杂志文章
doi:10.1016/j.abb.2011.11.024
更新日期:2012-02-01 00:00:00
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journal_title:Archives of biochemistry and biophysics
pub_type: 杂志文章
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pub_type: 杂志文章
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pub_type: 杂志文章
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pub_type: 杂志文章
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journal_title:Archives of biochemistry and biophysics
pub_type: 杂志文章
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更新日期:2004-11-15 00:00:00
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journal_title:Archives of biochemistry and biophysics
pub_type: 杂志文章
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更新日期:2001-11-01 00:00:00
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journal_title:Archives of biochemistry and biophysics
pub_type: 杂志文章
doi:10.1006/abbi.1995.1278
更新日期:1995-05-10 00:00:00
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journal_title:Archives of biochemistry and biophysics
pub_type: 杂志文章
doi:10.1006/abbi.1997.9936
更新日期:1997-05-01 00:00:00
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journal_title:Archives of biochemistry and biophysics
pub_type: 杂志文章
doi:10.1006/abbi.1993.1290
更新日期:1993-06-01 00:00:00
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journal_title:Archives of biochemistry and biophysics
pub_type: 杂志文章
doi:10.1006/abbi.1993.1417
更新日期:1993-09-01 00:00:00