|
|
| (2 intermediate revisions not shown.) |
| Line 1: |
Line 1: |
| | + | |
| | ==Structure of the N-terminal domain of RPA70, E7R mutant== | | ==Structure of the N-terminal domain of RPA70, E7R mutant== |
| - | <StructureSection load='4ipc' size='340' side='right' caption='[[4ipc]], [[Resolution|resolution]] 1.22Å' scene=''> | + | <StructureSection load='4ipc' size='340' side='right'caption='[[4ipc]], [[Resolution|resolution]] 1.22Å' scene=''> |
| | == Structural highlights == | | == Structural highlights == |
| - | <table><tr><td colspan='2'>[[4ipc]] is a 1 chain structure with sequence from [http://en.wikipedia.org/wiki/Homo_sapiens Homo sapiens]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4IPC OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=4IPC FirstGlance]. <br> | + | <table><tr><td colspan='2'>[[4ipc]] is a 1 chain structure with sequence from [https://en.wikipedia.org/wiki/Homo_sapiens Homo sapiens]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4IPC OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=4IPC FirstGlance]. <br> |
| - | </td></tr><tr id='related'><td class="sblockLbl"><b>[[Related_structure|Related:]]</b></td><td class="sblockDat">[[4ipd|4ipd]], [[4ipg|4ipg]], [[4iph|4iph]]</td></tr> | + | </td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 1.22Å</td></tr> |
| - | <tr id='gene'><td class="sblockLbl"><b>[[Gene|Gene:]]</b></td><td class="sblockDat">RPA1, REPA1, RPA70 ([http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&srchmode=5&id=9606 Homo sapiens])</td></tr>
| + | <tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=4ipc FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=4ipc OCA], [https://pdbe.org/4ipc PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=4ipc RCSB], [https://www.ebi.ac.uk/pdbsum/4ipc PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=4ipc ProSAT]</span></td></tr> |
| - | <tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=4ipc FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=4ipc OCA], [http://www.rcsb.org/pdb/explore.do?structureId=4ipc RCSB], [http://www.ebi.ac.uk/pdbsum/4ipc PDBsum]</span></td></tr> | + | |
| | </table> | | </table> |
| | == Function == | | == Function == |
| - | [[http://www.uniprot.org/uniprot/RFA1_HUMAN RFA1_HUMAN]] Plays an essential role in several cellular processes in DNA metabolism including replication, recombination and DNA repair. Binds and subsequently stabilizes single-stranded DNA intermediates and thus prevents complementary DNA from reannealing.<ref>PMID:19116208</ref> <ref>PMID:19996105</ref> Functions as component of the alternative replication protein A complex (aRPA). aRPA binds single-stranded DNA and probably plays a role in DNA repair; it does not support chromosomal DNA replication and cell cycle progression through S-phase. In vitro, aRPA cannot promote efficient priming by DNA polymerase alpha but supports DNA polymerase delta synthesis in the presence of PCNA and replication factor C (RFC), the dual incision/excision reaction of nucleotide excision repair and RAD51-dependent strand exchange.<ref>PMID:19116208</ref> <ref>PMID:19996105</ref> | + | [https://www.uniprot.org/uniprot/RFA1_HUMAN RFA1_HUMAN] Plays an essential role in several cellular processes in DNA metabolism including replication, recombination and DNA repair. Binds and subsequently stabilizes single-stranded DNA intermediates and thus prevents complementary DNA from reannealing.<ref>PMID:19116208</ref> <ref>PMID:19996105</ref> Functions as component of the alternative replication protein A complex (aRPA). aRPA binds single-stranded DNA and probably plays a role in DNA repair; it does not support chromosomal DNA replication and cell cycle progression through S-phase. In vitro, aRPA cannot promote efficient priming by DNA polymerase alpha but supports DNA polymerase delta synthesis in the presence of PCNA and replication factor C (RFC), the dual incision/excision reaction of nucleotide excision repair and RAD51-dependent strand exchange.<ref>PMID:19116208</ref> <ref>PMID:19996105</ref> |
| | <div style="background-color:#fffaf0;"> | | <div style="background-color:#fffaf0;"> |
| | == Publication Abstract from PubMed == | | == Publication Abstract from PubMed == |
| Line 17: |
Line 17: |
| | From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | | From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> |
| | </div> | | </div> |
| | + | <div class="pdbe-citations 4ipc" style="background-color:#fffaf0;"></div> |
| | | | |
| | ==See Also== | | ==See Also== |
| - | *[[Single-stranded DNA-binding protein|Single-stranded DNA-binding protein]] | + | *[[Single-stranded DNA-binding protein 3D structures|Single-stranded DNA-binding protein 3D structures]] |
| | == References == | | == References == |
| | <references/> | | <references/> |
| Line 25: |
Line 26: |
| | </StructureSection> | | </StructureSection> |
| | [[Category: Homo sapiens]] | | [[Category: Homo sapiens]] |
| - | [[Category: Chazin, W J]] | + | [[Category: Large Structures]] |
| - | [[Category: Feldkamp, M D]] | + | [[Category: Chazin WJ]] |
| - | [[Category: Fesik, S W]] | + | [[Category: Feldkamp MD]] |
| - | [[Category: Frank, A O]] | + | [[Category: Fesik SW]] |
| - | [[Category: Vangamudi, B]] | + | [[Category: Frank AO]] |
| - | [[Category: Ob-fold]] | + | [[Category: Vangamudi B]] |
| - | [[Category: Protein binding]]
| + | |
| Structural highlights
Function
RFA1_HUMAN Plays an essential role in several cellular processes in DNA metabolism including replication, recombination and DNA repair. Binds and subsequently stabilizes single-stranded DNA intermediates and thus prevents complementary DNA from reannealing.[1] [2] Functions as component of the alternative replication protein A complex (aRPA). aRPA binds single-stranded DNA and probably plays a role in DNA repair; it does not support chromosomal DNA replication and cell cycle progression through S-phase. In vitro, aRPA cannot promote efficient priming by DNA polymerase alpha but supports DNA polymerase delta synthesis in the presence of PCNA and replication factor C (RFC), the dual incision/excision reaction of nucleotide excision repair and RAD51-dependent strand exchange.[3] [4]
Publication Abstract from PubMed
Replication protein A (RPA) is the primary single-stranded DNA (ssDNA) binding protein in eukaryotes. The N-terminal domain of the RPA70 subunit (RPA70N) interacts via a basic cleft with a wide range of DNA processing proteins, including several that regulate DNA damage response and repair. Small molecule inhibitors that disrupt these protein-protein interactions are therefore of interest as chemical probes of these critical DNA processing pathways and as inhibitors to counter the upregulation of DNA damage response and repair associated with treatment of cancer patients with radiation or DNA-damaging agents. Determination of three-dimensional structures of protein-ligand complexes is an important step for elaboration of small molecule inhibitors. However, although crystal structures of free RPA70N and an RPA70N-peptide fusion construct have been reported, RPA70N-inhibitor complexes have been recalcitrant to crystallization. Analysis of the P61 lattice of RPA70N crystals led us to hypothesize that the ligand-binding surface was occluded. Surface reengineering to alter key crystal lattice contacts led to the design of RPA70N E7R, E100R, and E7R/E100R mutants. These mutants crystallized in a P212121 lattice that clearly had significant solvent channels open to the critical basic cleft. Analysis of X-ray crystal structures, target peptide binding affinities, and 15N-1H heteronuclear single-quantum coherence nuclear magnetic resonance spectra showed that the mutations do not result in perturbations of the RPA70N ligand-binding surface. The success of the design was demonstrated by determining the structure of RPA70N E7R soaked with a ligand discovered in a previously reported molecular fragment screen. A fluorescence anisotropy competition binding assay revealed this compound can inhibit the interaction of RPA70N with the peptide binding motif from the DNA damage response protein ATRIP. The implications of the results are discussed in the context of ongoing efforts to design RPA70N inhibitors.
Surface Reengineering of RPA70N Enables Cocrystallization with an Inhibitor of the Replication Protein A Interaction Motif of ATR Interacting Protein.,Feldkamp MD, Frank AO, Kennedy JP, Patrone JD, Vangamudi B, Waterson AG, Fesik SW, Chazin WJ Biochemistry. 2013 Sep 6. PMID:23962067[5]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
See Also
References
- ↑ Mason AC, Haring SJ, Pryor JM, Staloch CA, Gan TF, Wold MS. An alternative form of replication protein a prevents viral replication in vitro. J Biol Chem. 2009 Feb 20;284(8):5324-31. doi: 10.1074/jbc.M808963200. Epub 2008, Dec 29. PMID:19116208 doi:10.1074/jbc.M808963200
- ↑ Kemp MG, Mason AC, Carreira A, Reardon JT, Haring SJ, Borgstahl GE, Kowalczykowski SC, Sancar A, Wold MS. An alternative form of replication protein a expressed in normal human tissues supports DNA repair. J Biol Chem. 2010 Feb 12;285(7):4788-97. doi: 10.1074/jbc.M109.079418. Epub 2009 , Dec 7. PMID:19996105 doi:10.1074/jbc.M109.079418
- ↑ Mason AC, Haring SJ, Pryor JM, Staloch CA, Gan TF, Wold MS. An alternative form of replication protein a prevents viral replication in vitro. J Biol Chem. 2009 Feb 20;284(8):5324-31. doi: 10.1074/jbc.M808963200. Epub 2008, Dec 29. PMID:19116208 doi:10.1074/jbc.M808963200
- ↑ Kemp MG, Mason AC, Carreira A, Reardon JT, Haring SJ, Borgstahl GE, Kowalczykowski SC, Sancar A, Wold MS. An alternative form of replication protein a expressed in normal human tissues supports DNA repair. J Biol Chem. 2010 Feb 12;285(7):4788-97. doi: 10.1074/jbc.M109.079418. Epub 2009 , Dec 7. PMID:19996105 doi:10.1074/jbc.M109.079418
- ↑ Feldkamp MD, Frank AO, Kennedy JP, Patrone JD, Vangamudi B, Waterson AG, Fesik SW, Chazin WJ. Surface Reengineering of RPA70N Enables Cocrystallization with an Inhibitor of the Replication Protein A Interaction Motif of ATR Interacting Protein. Biochemistry. 2013 Sep 6. PMID:23962067 doi:10.1021/bi400542z
|
