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==CIRV p19 mutant T111S in complex with siRNA==
==CIRV p19 mutant T111S in complex with siRNA==
<StructureSection load='6bjh' size='340' side='right' caption='[[6bjh]], [[Resolution|resolution]] 2.58&Aring;' scene=''>
<StructureSection load='6bjh' size='340' side='right'caption='[[6bjh]], [[Resolution|resolution]] 2.58&Aring;' scene=''>
== Structural highlights ==
== Structural highlights ==
<table><tr><td colspan='2'>[[6bjh]] is a 4 chain structure. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=6BJH OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=6BJH FirstGlance]. <br>
<table><tr><td colspan='2'>[[6bjh]] is a 4 chain structure with sequence from [http://en.wikipedia.org/wiki/Cirv Cirv]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=6BJH OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=6BJH FirstGlance]. <br>
</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=6bjh FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=6bjh OCA], [http://pdbe.org/6bjh PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=6bjh RCSB], [http://www.ebi.ac.uk/pdbsum/6bjh PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=6bjh ProSAT]</span></td></tr>
</td></tr><tr id='gene'><td class="sblockLbl"><b>[[Gene|Gene:]]</b></td><td class="sblockDat">ORF4 ([http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&srchmode=5&id=39443 CIRV])</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=6bjh FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=6bjh OCA], [http://pdbe.org/6bjh PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=6bjh RCSB], [http://www.ebi.ac.uk/pdbsum/6bjh PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=6bjh ProSAT]</span></td></tr>
</table>
</table>
== Function ==
== Function ==
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<div style="background-color:#fffaf0;">
<div style="background-color:#fffaf0;">
== Publication Abstract from PubMed ==
== Publication Abstract from PubMed ==
Tombusviruses express a 19 kDa protein (p19) that, as a dimeric protein, suppresses the RNAs silencing pathway during infection by binding short-interfering RNA (siRNA) and preventing their association with the RNA-induced silencing complex (RISC). The p19 protein can bind to both endogenous and synthetic siRNAs with a high degree of size selectivity but with little sequence dependence. It also binds to other endogenous small RNAs such as microRNAs (miRNAs) but with lower affinity than to canonical siRNAs. It has become apparent, however, that miRNAs play a large role in gene regulation; their influence extends to expression and processing that affects virtually all eukaryotic processes. In order to develop new tools to study endogenous small RNAs, proteins that suppress specific miRNAs are required. Herein we describe mutational analysis of the p19 binding surface with the aim of creating p19 mutants with increased affinity for miR-122. By site-directed mutagenesis of a single residue, we describe p19 mutants with a nearly 50-fold increased affinity for miR-122 without altering the affinity for siRNA. Upon further mutational analysis of this site, we postulate that the higher affinity relies on hydrogen-bonding interactions but can be sterically hindered by residues with bulky side chains. Finally, we demonstrate the effectiveness of a mutant p19, p19-T111S, at sequestering miR-122 in human hepatoma cell lines, as compared to wild-type p19. Overall, our results suggest that p19 can be engineered to enhance its affinity toward specific small RNA molecules, particularly noncanonical miRNAs that are distinguishable based on locations of base-pair mismatches. The p19-T111S mutant also represents a new tool for the study of the function of miR-122 in post-transcriptional silencing in the human liver.
Viral suppressors of RNA silencing (VSRSs) are a diverse group of viral proteins that have evolved to disrupt eukaryotic RNA silencing pathways, thereby contributing to viral pathogenicity. The p19 protein is a VSRS that selectively binds to short interfering RNAs (siRNAs) over microRNAs (miRNAs). Mutational analysis has identified single amino acid substitutions that reverse this selectivity through new high-affinity interactions with human miR-122. Herein, we report crystal structures of complexed p19-T111S (2.6 A), p19-T111H (2.3 A) and wild-type p19 protein (2.2 A) from the Carnation Italian ringspot virus with small interfering RNA (siRNA) ligands. Structural comparisons reveal that these mutations do not lead to major changes in p19 architecture, but instead promote subtle rearrangement of residues and solvent molecules along the p19 midline. These observations suggest p19 uses many small interactions to distinguish siRNAs from miRNAs and perturbing these interactions can create p19 variants with novel RNA-recognition properties. DATABASE: Model data are deposited in the PDB database under the accession numbers 6BJG, 6BJH and 6BJV.


Enhanced specificity of the viral suppressor of RNA silencing protein p19 toward sequestering of human microRNA-122.,Cheng J, Danielson DC, Nasheri N, Singaravelu R, Pezacki JP Biochemistry. 2011 Sep 13;50(36):7745-55. doi: 10.1021/bi2008273. Epub 2011 Aug, 17. PMID:21819044<ref>PMID:21819044</ref>
Structural insights into interactions between viral suppressor of RNA silencing protein p19 mutants and small RNAs.,Foss DV, Schirle NT, MacRae IJ, Pezacki JP FEBS Open Bio. 2019 Jun;9(6):1042-1051. doi: 10.1002/2211-5463.12644. Epub 2019, May 17. PMID:31021526<ref>PMID:31021526</ref>


From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br>
From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br>
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__TOC__
__TOC__
</StructureSection>
</StructureSection>
[[Category: Cirv]]
[[Category: Large Structures]]
[[Category: Foss, D V]]
[[Category: Foss, D V]]
[[Category: MacRae, I J]]
[[Category: MacRae, I J]]

Revision as of 09:36, 3 July 2019

CIRV p19 mutant T111S in complex with siRNACIRV p19 mutant T111S in complex with siRNA

Structural highlights

6bjh is a 4 chain structure with sequence from Cirv. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance.
Gene:ORF4 (CIRV)
Resources:FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT

Function

[P19_CIRV] Acts as a suppressor of RNA-mediated gene silencing, also known as post-transcriptional gene silencing (PTGS), a mechanism of plant viral defense that limits the accumulation of viral RNAs. Binds to short interfering RNAs (siRNAs) with high affinity. Acts as a molecular caliper to specifically select siRNAs based on the length of the duplex region of the RNA.[1] [2]

Publication Abstract from PubMed

Viral suppressors of RNA silencing (VSRSs) are a diverse group of viral proteins that have evolved to disrupt eukaryotic RNA silencing pathways, thereby contributing to viral pathogenicity. The p19 protein is a VSRS that selectively binds to short interfering RNAs (siRNAs) over microRNAs (miRNAs). Mutational analysis has identified single amino acid substitutions that reverse this selectivity through new high-affinity interactions with human miR-122. Herein, we report crystal structures of complexed p19-T111S (2.6 A), p19-T111H (2.3 A) and wild-type p19 protein (2.2 A) from the Carnation Italian ringspot virus with small interfering RNA (siRNA) ligands. Structural comparisons reveal that these mutations do not lead to major changes in p19 architecture, but instead promote subtle rearrangement of residues and solvent molecules along the p19 midline. These observations suggest p19 uses many small interactions to distinguish siRNAs from miRNAs and perturbing these interactions can create p19 variants with novel RNA-recognition properties. DATABASE: Model data are deposited in the PDB database under the accession numbers 6BJG, 6BJH and 6BJV.

Structural insights into interactions between viral suppressor of RNA silencing protein p19 mutants and small RNAs.,Foss DV, Schirle NT, MacRae IJ, Pezacki JP FEBS Open Bio. 2019 Jun;9(6):1042-1051. doi: 10.1002/2211-5463.12644. Epub 2019, May 17. PMID:31021526[3]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.

References

  1. ↑ Vargason JM, Szittya G, Burgyan J, Tanaka Hall TM. Size selective recognition of siRNA by an RNA silencing suppressor. Cell. 2003 Dec 26;115(7):799-811. PMID:14697199
  2. ↑ Xia Z, Zhu Z, Zhu J, Zhou R. Recognition mechanism of siRNA by viral p19 suppressor of RNA silencing: a molecular dynamics study. Biophys J. 2009 Mar 4;96(5):1761-9. doi: 10.1016/j.bpj.2008.11.047. PMID:19254536 doi:http://dx.doi.org/10.1016/j.bpj.2008.11.047
  3. ↑ Foss DV, Schirle NT, MacRae IJ, Pezacki JP. Structural insights into interactions between viral suppressor of RNA silencing protein p19 mutants and small RNAs. FEBS Open Bio. 2019 Jun;9(6):1042-1051. doi: 10.1002/2211-5463.12644. Epub 2019, May 17. PMID:31021526 doi:http://dx.doi.org/10.1002/2211-5463.12644

6bjh, resolution 2.58Å

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