Related ArticlesPhosphorylation and flexibility of cyclic-AMP-dependent protein kinase (PKA) using (31)P NMR spectroscopy.
Biochemistry. 2002 May 14;41(19):5968-77
Authors: Seifert MH, Breitenlechner CB, Bossemeyer D, Huber R, Holak TA, Engh RA
Cell signaling pathways rely on phosphotransfer reactions that are catalyzed by protein kinases. The protein kinases themselves are typically regulated by phosphorylation and concurrent structural rearrangements, both near the catalytic site and elsewhere. Thus, physiological function requires posttranslational modification and deformable structures. A prototypical example is provided by cyclic AMP-dependent protein kinase (PKA). It is activated by phosphorylation, is inhomogeneously phosphorylated when expressed in bacteria, and exhibits a wide range of dynamic properties. Here we use (31)P nuclear magnetic resonance (NMR) spectroscopy to characterize the phosphorylation states and to estimate the flexibility of the phosphorylation sites of 2-, 3-, and 4-fold phosphorylated PKA. The phosphorylation sites Ser10, Ser139, Thr197, and Ser338 are assigned to individual NMR resonances, assisted by complexation with AMP-PNP and dephosphorylation with alkaline phosphatase. Rotational diffusion correlation times estimated from resonance line widths show progressively increasing flexibilities for phosphothreonine 197, phosphoserines 139 and 338, and disorder at phosphoserine 10, consistent with crystal structures of PKA. However, because the apparent rotational diffusion correlation time fitted for phosphothreonine 197 of the activation loop is longer than the overall PKA rotational diffusion time, microsecond to millisecond time scale conformational exchange effects involving motions of phosphothreonine 197 are probable. These may represent "open"-"closed" transitions of the uncomplexed protein in solution. These data represent direct measurements of flexibilities also associated with functional properties, such as ATP binding and membrane association, and illustrate the applicability of (31)P NMR for functional and dynamic characterization of protein kinase phosphorylation sites.
[NMR paper] Rapid and accurate structure determination of coiled-coil domains using NMR dipolar couplings: application to cGMP-dependent protein kinase Ialpha.
Rapid and accurate structure determination of coiled-coil domains using NMR dipolar couplings: application to cGMP-dependent protein kinase Ialpha.
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Protein Sci. 2005 Sep;14(9):2421-8
Authors: Schnell JR, Zhou GP, Zweckstetter M, Rigby AC, Chou JJ
Coiled-coil motifs play essential roles in protein assembly and molecular recognition, and are therefore the targets of many ongoing...
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[NMR paper] A novel view of domain flexibility in E. coli adenylate kinase based on structural mo
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J Mol Biol. 2002 Jan 11;315(2):155-70
Authors: Tugarinov V, Shapiro YE, Liang Z, Freed JH, Meirovitch E
Adenylate kinase from Escherichia coli (AKeco), consisting of a single 23.6 kDa polypeptide chain folded into domains CORE, AMPbd and LID, catalyzes the reaction AMP+ATP-->2ADP. In the...
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[NMR paper] Isoform-specific differences between the type Ialpha and IIalpha cyclic AMP-dependent
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J Biol Chem. 2000 Nov 10;275(45):35146-52
Authors: Banky P, Newlon MG, Roy M, Garrod S, Taylor SS, Jennings PA
Cyclic AMP dependent protein kinase (PKA) is controlled, in part, by the subcellular localization of the enzyme (). Discovery of...
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[NMR paper] Insights into tyrosine phosphorylation control of protein-protein association from th
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Biochemistry. 1998 Jan 20;37(3):867-77
Authors: Eisenmesser EZ, Post CB
A protein-protein association regulated by phosphorylation of tyrosine is examined by NMR...
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[NMR paper] NMR studies on the flexibility of nucleoside diphosphate kinase.
NMR studies on the flexibility of nucleoside diphosphate kinase.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--www3.interscience.wiley.com-aboutus-images-wiley_interscience_pubmed_logo_120x27.gif Related Articles NMR studies on the flexibility of nucleoside diphosphate kinase.
Proteins. 1997 Jun;28(2):150-2
Authors: Xu Y, Lecroisey A, Veron M, Delepierre M, Janin J
Human NDP kinase B, product of the nm23-H2 gene, binds DNA. It has been suggested that a helix hairpin on the protein surface, part of the nucleotide substrate binding...
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[NMR paper] NMR studies on the flexibility of nucleoside diphosphate kinase.
NMR studies on the flexibility of nucleoside diphosphate kinase.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--www3.interscience.wiley.com-aboutus-images-wiley_interscience_pubmed_logo_120x27.gif Related Articles NMR studies on the flexibility of nucleoside diphosphate kinase.
Proteins. 1997 Jun;28(2):150-2
Authors: Xu Y, Lecroisey A, Veron M, Delepierre M, Janin J
Human NDP kinase B, product of the nm23-H2 gene, binds DNA. It has been suggested that a helix hairpin on the protein surface, part of the nucleotide substrate binding...
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[NMR paper] Stimulation of cGMP-dependent protein kinase I alpha by a peptide from its own sequen
Stimulation of cGMP-dependent protein kinase I alpha by a peptide from its own sequence. An investigation by enzymology, circular dichroism and 1H NMR of the activity and structure of cGMP-dependent protein kinase I alpha-(546-576)-peptide amide.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--www3.interscience.wiley.com-aboutus-images-wiley_interscience_pubmed_logo_FREE_120x27.gif Related Articles Stimulation of cGMP-dependent protein kinase I alpha by a peptide from its own sequence. An investigation by enzymology, circular dichroism and 1H NMR of the activity and...
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[NMR paper] Stimulation of cGMP-dependent protein kinase I alpha by a peptide from its own sequen
Stimulation of cGMP-dependent protein kinase I alpha by a peptide from its own sequence. An investigation by enzymology, circular dichroism and 1H NMR of the activity and structure of cGMP-dependent protein kinase I alpha-(546-576)-peptide amide.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--www3.interscience.wiley.com-aboutus-images-wiley_interscience_pubmed_logo_FREE_120x27.gif Related Articles Stimulation of cGMP-dependent protein kinase I alpha by a peptide from its own sequence. An investigation by enzymology, circular dichroism and 1H NMR of the activity and...
Phosphorylation and flexibility of cyclic-AMP-dependent protein kinase (PKA) using (3
Linear Mode
