Sandbox Reserved 826: Difference between revisions

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OCA, Dimitri Feltrin

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-<Structure load='2w9f' size='500' frame='true' align='right' caption='3D structure of CDK4 (cyan) with cyclin D1 (orange) in complex' scene='56/568024/3d_structure_cdk4complex/2' />
+<Structure load='2w9f' size='500' frame='true' align='right' caption='3D structure of CDK4 (cyan) with cyclin D1 (orange) in complex' scene='56/568024/3d_structure_cdk4complex/3' />
 == '''CDK4 structure''' ==
-In order to gain structural information about CDKs, a crystallographic analysis of CDK in complex with cyclin D was performed. CDK4 and cyclin D1, were overexpressed in insect cells for crystallisation and slightly modified for better crystallization. CDK 4 possesses the typical <scene name='56/568024/Bilobal_cdk4/1'>bilobal structure</scene> of kinases, containing a 5-strandet ß-sheet and a N-terminal domain (residues 1-96). Within the N-terminal domain the helix alpha-C is packed against the ß-sheet. Furthermore an ATP binding site is located in between these domains, which might bind ATP after minimal conformational rearrangement of residues Asp-99, Asp-140, Lys-142, and Tyr-17. Residues 161-171 form a <scene name='56/568024/T_loop/1'>T loop</scene> containing alpha-helices. Phosphorylation of this T-loop on residue Thr-172 or cyclin D binding may change its conformation in order to activate kinase activity of CDK4. Lambda-phosphatase incubation studies confirm the importance of phosphorylation of the T-loop for full kinase activity <ref> PMID: 19237565 </ref>.
+In order to gain structural information about CDKs, a crystallographic analysis of CDK in complex with cyclin D was performed. CDK4 and cyclin D1, were overexpressed in insect cells for crystallisation and slightly modified for better crystallization. CDK 4 possesses the typical <scene name='56/568024/Bilobal_cdk4/2'>bilobal structure</scene> of kinases, containing a 5-strandet ß-sheet and a N-terminal domain (residues 1-96). Within the N-terminal domain the helix alpha-C is packed against the ß-sheet. Furthermore an ATP binding site is located in between these domains, which might bind ATP after minimal conformational rearrangement of residues Asp-99, Asp-140, Lys-142, and Tyr-17. Residues 161-171 form a <scene name='56/568024/T_loop/2'>T loop</scene> containing alpha-helices. Phosphorylation of this T-loop on residue Thr-172 or cyclin D binding may change its conformation in order to activate kinase activity of CDK4. Lambda-phosphatase incubation studies confirm the importance of phosphorylation of the T-loop for full kinase activity <ref> PMID: 19237565 </ref>.
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 == '''Cyclin D1 structure''' ==
 Cyclin D1 consists of a double glycin box domain fold, containing 11 alpha helices. Its secondary structure is generally equivalent to the structure of other cyclins. Furthermore, it contains 2 Rb binding sites comprising an LxCxE motive t its N-terminal site. Structural conservation of the RxL motive, responsible for peptide binding was observed on Cyclin D1 and Cyclin A. Peptide binding studies substantiate this observation <ref> PMID: 19237565 </ref>.
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 == '''Cyclin D1-CDK4 complex''' ==
-CDK4 in complex with Cyclin D1 shows an engagement of the CDK4 alpha-helix with cyclin D1. Nevertheless, the helix does perform the conformational switch, normally known for CDK activation in other CDK/cyclin complexes <ref> PMID: 7630397 </ref>. Surprisingly, the CDK4/cyclin D1 structure reminds to the structures of inactive structures of non cyclin bound CDK2 and CDK7 <ref> PMID: 8510751 </ref>. Further stabilization of the CDK4 T-loop in the inactive confirmation is achieved by interactions of C- and N- terminal lobes of the kinase. These residues, containing an Asp158–Phe159–Gly160 <scene name='56/568024/Dfg_motif/1'>(DFG) motif</scene> are condensed into a helix, which is stabilized by interactions with the alpha-C-helix, ß4-strand, ß6-strand as well as the apex of the T-loop. The architecture of the T-Loop is similar to the one observed at CDK7 and CDK6. CDK4 kinase activation could be achieved due to movement of the alpha-C-helix. Although the C-alpha-lobe of CDK4 seems to be bound by Cyclin D1, the rotation of the C-lobe of CDK4 is not maximal. This reduces the buried surface area of of the CDK4/cyclin D1 interface in comparison of the buried surface of other CDK/cyclin complexes.
+CDK4 in complex with Cyclin D1 shows an engagement of the CDK4 alpha-helix with cyclin D1. Nevertheless, the helix does perform the conformational switch, normally known for CDK activation in other CDK/cyclin complexes <ref> PMID: 7630397 </ref>. Surprisingly, the CDK4/cyclin D1 structure reminds to the structures of inactive structures of non cyclin bound CDK2 and CDK7 <ref> PMID: 8510751 </ref>. Further stabilization of the CDK4 T-loop in the inactive confirmation is achieved by interactions of C- and N- terminal lobes of the kinase. These residues, containing an Asp158–Phe159–Gly160 <scene name='56/568024/Dfg_motif/2'>(DFG) motif</scene> are condensed into a helix, which is stabilized by interactions with the alpha-C-helix, ß4-strand, ß6-strand as well as the apex of the T-loop. The architecture of the T-Loop is similar to the one observed at CDK7 and CDK6. CDK4 kinase activation could be achieved due to movement of the alpha-C-helix. Although the C-alpha-lobe of CDK4 seems to be bound by Cyclin D1, the rotation of the C-lobe of CDK4 is not maximal. This reduces the buried surface area of of the CDK4/cyclin D1 interface in comparison of the buried surface of other CDK/cyclin complexes.
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