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| | ==TURNIP YELLOW MOSAIC VIRUS PROTEASE/DEUBIQUITINASE DOMAIN, I847A MUTANT== | | ==TURNIP YELLOW MOSAIC VIRUS PROTEASE/DEUBIQUITINASE DOMAIN, I847A MUTANT== |
| - | <StructureSection load='5lwa' size='340' side='right' caption='[[5lwa]], [[Resolution|resolution]] 1.65Å' scene=''> | + | <StructureSection load='5lwa' size='340' side='right'caption='[[5lwa]], [[Resolution|resolution]] 1.65Å' scene=''> |
| | == Structural highlights == | | == Structural highlights == |
| - | <table><tr><td colspan='2'>[[5lwa]] is a 2 chain structure. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=5LWA OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=5LWA FirstGlance]. <br> | + | <table><tr><td colspan='2'>[[5lwa]] is a 2 chain structure with sequence from [https://en.wikipedia.org/wiki/Turnip_yellow_mosaic_virus Turnip yellow mosaic virus]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=5LWA OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=5LWA FirstGlance]. <br> |
| - | </td></tr><tr id='related'><td class="sblockLbl"><b>[[Related_structure|Related:]]</b></td><td class="sblockDat">[[4a5u|4a5u]], [[5lw5|5lw5]]</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]] 1.653Å</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=5lwa FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=5lwa OCA], [http://pdbe.org/5lwa PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=5lwa RCSB], [http://www.ebi.ac.uk/pdbsum/5lwa PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=5lwa ProSAT]</span></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=5lwa FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=5lwa OCA], [https://pdbe.org/5lwa PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=5lwa RCSB], [https://www.ebi.ac.uk/pdbsum/5lwa PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=5lwa ProSAT]</span></td></tr> |
| | </table> | | </table> |
| | == Function == | | == Function == |
| - | [[http://www.uniprot.org/uniprot/POLR_TYMV POLR_TYMV]] Acts as a cysteine protease, methyltransferase and deubiquitinase. The cyteine protease activity cleaves the polyprotein giving rise to mature proteins. The methyltransferase domain is probably involved in viral RNA capping. The deubiquitylating activity counteracts the degradation of the viral polymerase mediated by the host ubiquitin-proteasome system. The polymerase is thus stabilized and infectivity is increased.<ref>PMID:22117220</ref> RNA-directed RNA polymerase is responsible for the replication and transcription of the genome.<ref>PMID:22117220</ref> | + | [https://www.uniprot.org/uniprot/POLR_TYMV POLR_TYMV] Acts as a cysteine protease, methyltransferase and deubiquitinase. The cyteine protease activity cleaves the polyprotein giving rise to mature proteins. The methyltransferase domain is probably involved in viral RNA capping. The deubiquitylating activity counteracts the degradation of the viral polymerase mediated by the host ubiquitin-proteasome system. The polymerase is thus stabilized and infectivity is increased.<ref>PMID:22117220</ref> RNA-directed RNA polymerase is responsible for the replication and transcription of the genome.<ref>PMID:22117220</ref> |
| | <div style="background-color:#fffaf0;"> | | <div style="background-color:#fffaf0;"> |
| | == Publication Abstract from PubMed == | | == Publication Abstract from PubMed == |
| - | Processing of the polyprotein of Turnip yellow mosaic virus is mediated by the protease PRO. PRO cleaves at two places, one of which is at the C-terminus of the PRO domain of another polyprotein molecule. In addition to this processing activity, PRO possesses an ubiquitin hydrolase (DUB) activity. The crystal structure of PRO has previously been reported in its polyprotein-processing mode with the C-terminus of one PRO inserted into the catalytic site of the next PRO, generating PRO polymers in the crystal packing of the trigonal space group. Here, two mutants designed to disrupt specific PRO-PRO interactions were generated, produced and purified. Crystalline plates were obtained by seeding and cross-seeding from initial `sea urchin'-like microcrystals of one mutant. The plates diffracted to beyond 2 A resolution at a synchrotron source and complete data sets were collected for the two mutants. Data processing and analysis indicated that both mutant crystals belonged to the same monoclinic space group, with two molecules of PRO in the asymmetric unit.
| + | The positive-strand RNA virus Turnip yellow mosaic virus (TYMV) encodes an ovarian tumor (OTU)-like protease/deubiquitinase (PRO/DUB) protein domain involved both in proteolytic processing of the viral polyprotein through its PRO activity, and in removal of ubiquitin chains from ubiquitylated substrates through its DUB activity. Here, the crystal structures of TYMV PRO/DUB mutants and molecular dynamics simulations reveal that an idiosyncratic mobile loop participates in reversibly constricting its unusual catalytic site by adopting "open", "intermediate" or "closed" conformations. The two cis-prolines of the loop form a rigid flap that in the most closed conformation zips up against the other side of the catalytic cleft. The intermediate and closed conformations also correlate with a reordering of the TYMV PRO/DUB catalytic dyad, that then assumes a classical, yet still unusually mobile, OTU DUB alignment. Further structure-based mutants designed to interfere with the loop's mobility were assessed for enzymatic activity in vitro and in vivo, and were shown to display reduced DUB activity while retaining PRO activity. This indicates that control of the switching between the dual PRO/DUB activities resides prominently within this loop next to the active site. Introduction of mutations into the viral genome revealed that the DUB activity contributes to the extent of viral RNA accumulation both in single cells and in whole plants. In addition, the conformation of the mobile flap was also found to influence symptoms severity in planta. Such mutants now provide powerful tools with which to study the specific roles of reversible ubiquitylation in viral infection. |
| | | | |
| - | Crystallization of mutants of Turnip yellow mosaic virus protease/ubiquitin hydrolase designed to prevent protease self-recognition.,Ayach M, Bressanelli S Acta Crystallogr F Struct Biol Commun. 2015 Apr;71(Pt 4):405-8. doi:, 10.1107/S2053230X15003945. Epub 2015 Mar 20. PMID:25849500<ref>PMID:25849500</ref>
| + | A mobile loop near the active site acts as a switch between the dual activities of a viral protease/deubiquitinase.,Jupin I, Ayach M, Jomat L, Fieulaine S, Bressanelli S PLoS Pathog. 2017 Nov 8;13(11):e1006714. doi: 10.1371/journal.ppat.1006714., eCollection 2017 Nov. PMID:29117247<ref>PMID:29117247</ref> |
| | | | |
| | 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> |
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| | __TOC__ | | __TOC__ |
| | </StructureSection> | | </StructureSection> |
| - | [[Category: Ayach, M]] | + | [[Category: Large Structures]] |
| - | [[Category: Bressanelli, S]] | + | [[Category: Turnip yellow mosaic virus]] |
| - | [[Category: Cysteine protease]] | + | [[Category: Ayach M]] |
| - | [[Category: Deubiquitinase]] | + | [[Category: Bressanelli S]] |
| - | [[Category: Hydrolase]]
| + | |
| - | [[Category: Virus replicase polyprotein]]
| + | |
| Structural highlights
Function
POLR_TYMV Acts as a cysteine protease, methyltransferase and deubiquitinase. The cyteine protease activity cleaves the polyprotein giving rise to mature proteins. The methyltransferase domain is probably involved in viral RNA capping. The deubiquitylating activity counteracts the degradation of the viral polymerase mediated by the host ubiquitin-proteasome system. The polymerase is thus stabilized and infectivity is increased.[1] RNA-directed RNA polymerase is responsible for the replication and transcription of the genome.[2]
Publication Abstract from PubMed
The positive-strand RNA virus Turnip yellow mosaic virus (TYMV) encodes an ovarian tumor (OTU)-like protease/deubiquitinase (PRO/DUB) protein domain involved both in proteolytic processing of the viral polyprotein through its PRO activity, and in removal of ubiquitin chains from ubiquitylated substrates through its DUB activity. Here, the crystal structures of TYMV PRO/DUB mutants and molecular dynamics simulations reveal that an idiosyncratic mobile loop participates in reversibly constricting its unusual catalytic site by adopting "open", "intermediate" or "closed" conformations. The two cis-prolines of the loop form a rigid flap that in the most closed conformation zips up against the other side of the catalytic cleft. The intermediate and closed conformations also correlate with a reordering of the TYMV PRO/DUB catalytic dyad, that then assumes a classical, yet still unusually mobile, OTU DUB alignment. Further structure-based mutants designed to interfere with the loop's mobility were assessed for enzymatic activity in vitro and in vivo, and were shown to display reduced DUB activity while retaining PRO activity. This indicates that control of the switching between the dual PRO/DUB activities resides prominently within this loop next to the active site. Introduction of mutations into the viral genome revealed that the DUB activity contributes to the extent of viral RNA accumulation both in single cells and in whole plants. In addition, the conformation of the mobile flap was also found to influence symptoms severity in planta. Such mutants now provide powerful tools with which to study the specific roles of reversible ubiquitylation in viral infection.
A mobile loop near the active site acts as a switch between the dual activities of a viral protease/deubiquitinase.,Jupin I, Ayach M, Jomat L, Fieulaine S, Bressanelli S PLoS Pathog. 2017 Nov 8;13(11):e1006714. doi: 10.1371/journal.ppat.1006714., eCollection 2017 Nov. PMID:29117247[3]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
References
- ↑ Chenon M, Camborde L, Cheminant S, Jupin I. A viral deubiquitylating enzyme targets viral RNA-dependent RNA polymerase and affects viral infectivity. EMBO J. 2012 Feb 1;31(3):741-53. doi: 10.1038/emboj.2011.424. Epub 2011 Nov 25. PMID:22117220 doi:http://dx.doi.org/10.1038/emboj.2011.424
- ↑ Chenon M, Camborde L, Cheminant S, Jupin I. A viral deubiquitylating enzyme targets viral RNA-dependent RNA polymerase and affects viral infectivity. EMBO J. 2012 Feb 1;31(3):741-53. doi: 10.1038/emboj.2011.424. Epub 2011 Nov 25. PMID:22117220 doi:http://dx.doi.org/10.1038/emboj.2011.424
- ↑ Jupin I, Ayach M, Jomat L, Fieulaine S, Bressanelli S. A mobile loop near the active site acts as a switch between the dual activities of a viral protease/deubiquitinase. PLoS Pathog. 2017 Nov 8;13(11):e1006714. doi: 10.1371/journal.ppat.1006714., eCollection 2017 Nov. PMID:29117247 doi:http://dx.doi.org/10.1371/journal.ppat.1006714
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