User:Iris To/Retinoblastoma Protein Regulation
From Proteopedia
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From previous studies, it has been determined that the phosphorylation state and activity of Rb are controlled by a balance of kinase and phosphatase activity, in which Cdks '''phosphorylate''' Rb from late G1 to mitosis and PP1 '''dephosphorylates''' Rb for mitotic exit<ref name=Hirschi>PMID: 20694007</ref>. An enzyme-docking site for PP1c (the catalytic subunit of PP1) was determined to overlap with the docking site for Cdks, more specifically the '''RxL Cyclin binding site'''<ref name=Lowe>PMID:12501191</ref> that brings Cdk-Cyclins to Rb, through a crystal structure; this crystal structure initiated studies on phosphatase and kinase competition for the docking site. | From previous studies, it has been determined that the phosphorylation state and activity of Rb are controlled by a balance of kinase and phosphatase activity, in which Cdks '''phosphorylate''' Rb from late G1 to mitosis and PP1 '''dephosphorylates''' Rb for mitotic exit<ref name=Hirschi>PMID: 20694007</ref>. An enzyme-docking site for PP1c (the catalytic subunit of PP1) was determined to overlap with the docking site for Cdks, more specifically the '''RxL Cyclin binding site'''<ref name=Lowe>PMID:12501191</ref> that brings Cdk-Cyclins to Rb, through a crystal structure; this crystal structure initiated studies on phosphatase and kinase competition for the docking site. | ||
| - | It has been found that PP1c '''directly inhibits phosphorylation''' of RbC from Cdk2-CycA[http://www.neb.com/nebecomm/products/productp6025.asp]; it is not affected by phosphatase activity but by the presence of the '''KLRF docking site''', which binds the catalytic subunit of PP1c to a specific site of Rb<ref name=Hirschi>PMID: 20694007</ref>. In addition, it has been observed that PP1c makes complexes with RbC when there are many phosphatases produced, thus overthrowing Cdk activity and inhibiting cell progression from the G1 phase. This competition is important in terms of '''cell signaling''', which is affected by response to cellular stress, cell cycle exit, DNA damage, etc<ref name=Hirschi>PMID: 20694007</ref>. This is because the competition further controls cell overgrowth besides phosphorylation regulation. Studies determined a biochemical mechanism where directly competing kinases and phosphatase activity regulates Rb phosphorylation and activity, but there has not been an established study that defined a mechanism that controlled the outcome of the competition between each enzyme<ref name=Hirschi>PMID: 20694007</ref>. | + | It has been found that PP1c '''directly inhibits phosphorylation''' of RbC from Cdk2-CycA[http://www.neb.com/nebecomm/products/productp6025.asp]; it is not affected by phosphatase activity but by the presence of the '''KLRF docking site'''[http://www.nature.com/nsmb/journal/v17/n9/fig_tab/nsmb.1868_F3.html], which binds the catalytic subunit of PP1c to a specific site of Rb<ref name=Hirschi>PMID: 20694007</ref>. In addition, it has been observed that PP1c makes complexes with RbC when there are many phosphatases produced, thus overthrowing Cdk activity and inhibiting cell progression from the G1 phase. This competition is important in terms of '''cell signaling''', which is affected by response to cellular stress, cell cycle exit, DNA damage, etc<ref name=Hirschi>PMID: 20694007</ref>. This is because the competition further controls cell overgrowth besides phosphorylation regulation. Studies determined a biochemical mechanism where directly competing kinases and phosphatase activity regulates Rb phosphorylation and activity, but there has not been an established study that defined a mechanism that controlled the outcome of the competition between each enzyme<ref name=Hirschi>PMID: 20694007</ref>. |
==Relevance== | ==Relevance== | ||
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<StructureSection load='3n5u' size='500' side='right' caption='Structure of Rb-PP1c(PDB entry [[3n5u]])' scene=''> | <StructureSection load='3n5u' size='500' side='right' caption='Structure of Rb-PP1c(PDB entry [[3n5u]])' scene=''> | ||
==Structure and Function== | ==Structure and Function== | ||
| - | PP1[http://www.signaling-gateway.org/update/images/su-0406-3-i1.jpg] is known as protein serine/threonine phosphatase, and has an N-terminal, α/β domain, and a C-terminal β domain. The β domains come together as a <scene name='User:Iris_To/Retinoblastoma_Protein_Regulation/Rb_cterm_binding/7'>β-sandwich</scene>. The catalytic site is inside a a large Y-shaped cleft made by the three domains. Grooves are made by the domains: a hydrophobic, acidic, and C-terminal groove<ref>Terrak, Mohammed, Kerff, Frederic, Langsetmo, Knut, Tao, Terence, & Dominguez, Roberto Structural basis of protein phosphatase 1 regulation. Nature, 429, 780–784 (17 June 2004); 10.1038/nature02582</ref>. Rb866-889 is required for binding with PP1c; Rb870-882 is required for PP1c association with all of the significant interacting residues. Rb 870-882 also contains the Cyclin A docking site and KLRF sequence(like RVxF motif) where kinase and phosphatase compete. | + | PP1[http://www.signaling-gateway.org/update/images/su-0406-3-i1.jpg] is known as protein serine/threonine phosphatase, and has an N-terminal, α/β domain, and a C-terminal β domain. The β domains come together as a <scene name='User:Iris_To/Retinoblastoma_Protein_Regulation/Rb_cterm_binding/7'>β-sandwich</scene>. The catalytic site is inside a a large Y-shaped cleft made by the three domains. Grooves are made by the domains: a hydrophobic, acidic, and C-terminal groove<ref>Terrak, Mohammed, Kerff, Frederic, Langsetmo, Knut, Tao, Terence, & Dominguez, Roberto Structural basis of protein phosphatase 1 regulation. Nature, 429, 780–784 (17 June 2004); 10.1038/nature02582</ref>. Rb866-889 is required for binding with PP1c; Rb870-882 is required for PP1c association with all of the significant interacting residues. Rb 870-882 also contains the Cyclin A docking site and KLRF sequence(like RVxF motif of PP1 in that it mediates binding<ref>Wakula, P., Beullens, M., Ceulemans, H., Stalmans, W., and Bollen, M. (2003) J. Biol. Chem. 278, 18817–18823</ref>) where kinase and phosphatase compete. |
<scene name='User:Iris_To/Retinoblastoma_Protein_Regulation/Rb_cterm_binding/2'>Rb binds PP1c with an extended conformation at hydrophobic interface</scene> of the β-sandwich subdomain, which is <scene name='User:Iris_To/Retinoblastoma_Protein_Regulation/Rb_cterm_binding/5'>away from phosphatase active site</scene><ref name=Hirschi>PMID: 20694007</ref>. <scene name='User:Iris_To/Retinoblastoma_Protein_Regulation/Rb_cterm_binding/3'>Arg876,Phe877,Asp877 form</scene> a <scene name='User:Iris_To/Retinoblastoma_Protein_Regulation/Rb_cterm_binding/6'>short β-strand</scene> that becomes part of sheet 1 of the PP1c β-sandwich subdomain, with hydrogen bonding interactions with the parallel adjacent strand. | <scene name='User:Iris_To/Retinoblastoma_Protein_Regulation/Rb_cterm_binding/2'>Rb binds PP1c with an extended conformation at hydrophobic interface</scene> of the β-sandwich subdomain, which is <scene name='User:Iris_To/Retinoblastoma_Protein_Regulation/Rb_cterm_binding/5'>away from phosphatase active site</scene><ref name=Hirschi>PMID: 20694007</ref>. <scene name='User:Iris_To/Retinoblastoma_Protein_Regulation/Rb_cterm_binding/3'>Arg876,Phe877,Asp877 form</scene> a <scene name='User:Iris_To/Retinoblastoma_Protein_Regulation/Rb_cterm_binding/6'>short β-strand</scene> that becomes part of sheet 1 of the PP1c β-sandwich subdomain, with hydrogen bonding interactions with the parallel adjacent strand. | ||
Revision as of 20:51, 30 November 2011
Contents |
Retinoblastoma Protein
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| 3n5u, resolution 3.20Å () | |||||||
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| Ligands: | , | ||||||
| Gene: | PPP1A, PPP1CA (Homo sapiens) | ||||||
| Activity: | Phosphoprotein phosphatase, with EC number 3.1.3.16 | ||||||
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| Resources: | FirstGlance, OCA, RCSB, PDBsum | ||||||
| Coordinates: | save as pdb, mmCIF, xml | ||||||
The retinoblastoma protein (Rb) is a suppressor protein, also known as a tumor suppressor, involved in negative regulation of the cell cycle[1] due to its ability to bind to the transcription factor E2F[2]. It acts as a cell cycle checkpoint during the G1 phase, determining if the cell cycle should continue or stop. In its normal state, Rb is activated, which prevents the cell cycle to continue because it can recruit transcriptional co-repressors, blocking transcription[1]. Rb is deactivated after being phosphorylated by Cyclin-dependent kinases (Cdks)[3], and thus cannot associate factors that inhibit transcription factors that allow the cell cycle to continue. Normally, once the cell cycle is complete, Rb is activated once more by being dephosphorylated by phosphoprotein phosphatases (PP1)[4] to remove the cell from mitosis. Unregulated deactivation of Rb can lead to uncontrolled growth of cells, which is why studying this protein is important in cancer research.
Regulation of Rb
From previous studies, it has been determined that the phosphorylation state and activity of Rb are controlled by a balance of kinase and phosphatase activity, in which Cdks phosphorylate Rb from late G1 to mitosis and PP1 dephosphorylates Rb for mitotic exit[2]. An enzyme-docking site for PP1c (the catalytic subunit of PP1) was determined to overlap with the docking site for Cdks, more specifically the RxL Cyclin binding site[3] that brings Cdk-Cyclins to Rb, through a crystal structure; this crystal structure initiated studies on phosphatase and kinase competition for the docking site.
It has been found that PP1c directly inhibits phosphorylation of RbC from Cdk2-CycA[5]; it is not affected by phosphatase activity but by the presence of the KLRF docking site[6], which binds the catalytic subunit of PP1c to a specific site of Rb[2]. In addition, it has been observed that PP1c makes complexes with RbC when there are many phosphatases produced, thus overthrowing Cdk activity and inhibiting cell progression from the G1 phase. This competition is important in terms of cell signaling, which is affected by response to cellular stress, cell cycle exit, DNA damage, etc[2]. This is because the competition further controls cell overgrowth besides phosphorylation regulation. Studies determined a biochemical mechanism where directly competing kinases and phosphatase activity regulates Rb phosphorylation and activity, but there has not been an established study that defined a mechanism that controlled the outcome of the competition between each enzyme[2].
Relevance
While there are other characteristics of cancer cells, such as cell dispersal and apoptosis resistance, the foundation of cell proliferation is at the cell cycle[4]. Studying this competition in overlapping docking sites is important in cancer research because it can give a better understanding of how Rb can be manipulated if there is an error in regulation, leading to unwanted cell growth.
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3D Structures
1o9k with E2F
1h25 with Cdk2/Cyclin A
References
- ↑ Lee C, Chang JH, Lee HS, Cho Y. Structural basis for the recognition of the E2F transactivation domain by the retinoblastoma tumor suppressor. Genes Dev. 2002 Dec 15;16(24):3199-212. PMID:12502741 doi:10.1101/gad.1046102
- ↑ 2.0 2.1 2.2 2.3 2.4 2.5 Hirschi A, Cecchini M, Steinhardt RC, Schamber MR, Dick FA, Rubin SM. An overlapping kinase and phosphatase docking site regulates activity of the retinoblastoma protein. Nat Struct Mol Biol. 2010 Sep;17(9):1051-7. Epub 2010 Aug 8. PMID:20694007 doi:10.1038/nsmb.1868
- ↑ Lowe ED, Tews I, Cheng KY, Brown NR, Gul S, Noble ME, Gamblin SJ, Johnson LN. Specificity determinants of recruitment peptides bound to phospho-CDK2/cyclin A. Biochemistry. 2002 Dec 31;41(52):15625-34. PMID:12501191
- ↑ http://www.ias.ac.in/currsci/sep102001/515.pdf
- ↑ Terrak, Mohammed, Kerff, Frederic, Langsetmo, Knut, Tao, Terence, & Dominguez, Roberto Structural basis of protein phosphatase 1 regulation. Nature, 429, 780–784 (17 June 2004); 10.1038/nature02582
- ↑ Wakula, P., Beullens, M., Ceulemans, H., Stalmans, W., and Bollen, M. (2003) J. Biol. Chem. 278, 18817–18823
![[1]](http://www.signaling-gateway.org/update/images/su-0406-3-i1.jpg){kind=link}