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| | <StructureSection load='4bwg' size='340' side='right'caption='[[4bwg]], [[Resolution|resolution]] 2.60Å' scene=''> | | <StructureSection load='4bwg' size='340' side='right'caption='[[4bwg]], [[Resolution|resolution]] 2.60Å' scene=''> |
| | == Structural highlights == | | == Structural highlights == |
| - | <table><tr><td colspan='2'>[[4bwg]] is a 12 chain structure with sequence from [http://en.wikipedia.org/wiki/Escherichia_coli_98nk2 Escherichia coli 98nk2]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4BWG OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=4BWG FirstGlance]. <br> | + | <table><tr><td colspan='2'>[[4bwg]] is a 12 chain structure with sequence from [https://en.wikipedia.org/wiki/Escherichia_coli_98NK2 Escherichia coli 98NK2]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4BWG OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=4BWG FirstGlance]. <br> |
| - | </td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat"><scene name='pdbligand=GOL:GLYCEROL'>GOL</scene>, <scene name='pdbligand=SO4:SULFATE+ION'>SO4</scene></td></tr> | + | </td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=GOL:GLYCEROL'>GOL</scene>, <scene name='pdbligand=SO4:SULFATE+ION'>SO4</scene></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=4bwg FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=4bwg OCA], [http://pdbe.org/4bwg PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=4bwg RCSB], [http://www.ebi.ac.uk/pdbsum/4bwg PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=4bwg 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=4bwg FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=4bwg OCA], [https://pdbe.org/4bwg PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=4bwg RCSB], [https://www.ebi.ac.uk/pdbsum/4bwg PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=4bwg ProSAT]</span></td></tr> |
| | </table> | | </table> |
| - | <div style="background-color:#fffaf0;">
| + | == Function == |
| - | == Publication Abstract from PubMed == | + | [[https://www.uniprot.org/uniprot/SUBA_ECOLX SUBA_ECOLX]] Protease subunit of subtilase cytotoxin SubAB5 (PubMed:17024087). An endoprotease specific for host endoplasmic reticulum (ER) chaperone BiP/HSPA5, has no activity on human HSP70 or HSPA8 (PubMed:17024087). Cleaves between 'Leu-416' and 'Leu-417' of BiP/HSPA5 in the hinge between BiP's ATPase and protein-binding domains (PubMed:17024087). This induces host ER stress response and eventual cell death (PubMed:18005237, PubMed:18433465). Culture supernatant of E.coli expressing both subA and subB are toxic for Vero cells (African green monkey kidney cell line), Chinese hamster ovary cells and Hct-8 cells (human colonic epithelial cell line); the subunits are not toxic individually (PubMed:15226357). Purified SubAB5 is highly toxic, <0.1 pg is able to kill at least 50% of 30'000 Vero cells in a microtiter plate assay after 3 days; no cytotoxicity is seen at 24 hours (PubMed:15226357). Preabsorption with cells expressing a ganglioside GM2 mimic reduced cytotoxicity of SubAB5 by 93% in the Vero cytotoxicity assay (PubMed:15226357). Intraperitoneal injection of 200 ng of purified SubAB5 kills mice; the higher the dose the faster the mice die. Animals injected intraperitoneally with purified SubAB5 have microvascular thrombi in the brain and other organs, including the renal tubules and glomeruli (PubMed:15226357). Injection induces an unfolded response in mice (PubMed:17024087). Mice fed E.coli cells expressing cloned SubAB5 experience drastic weight loss and appear ill and lethargic (PubMed:15226357). Protein synthesis in Vero cells is transiently inhibited by SubAB5; both subunits are required for this effect (PubMed:17101670, PubMed:18005237, PubMed:18433465). Inhibition of protein synthesis is prevented by brefeldin A; cells are arrested in the G1 phase (PubMed:18005237). SubAB5 at 100 ng/ml induced caspase-dependent apoptosis in Vero cells through mitochondrial membrane damage (PubMed:19380466).<ref>PMID:15226357</ref> <ref>PMID:17024087</ref> <ref>PMID:17101670</ref> <ref>PMID:18005237</ref> <ref>PMID:18433465</ref> <ref>PMID:19380466</ref> |
| - | Pathogenic strains of Escherichia coli produce a number of toxins that belong to the AB5 toxin family, which comprise a catalytic A-subunit that induces cellular dysfunction and a B-pentamer that recognizes host glycans. Whilst the molecular actions of many of the individual subunits of AB5 toxins are well understood, how they self-associate and the effect of this association on cytotoxicity is poorly understood. Here we have solved the structure of the holo SubAB toxin that, in contrast to other AB5 toxins whose molecular targets are located in the cytosol, cleaves the endoplasmic reticulum (ER) chaperone BiP. SubA interacts with SubB in a similar manner to other AB5 toxins via the A2 helix and a conserved disulfide bond that joins the A1 domain with the A2 helix. The structure revealed that the active site of SubA is not occluded by the B-pentamer, and the B-pentamer does not enhance or inhibit the activity of SubA. Structure-based sequence comparisons of other AB5 toxin family members, combined with extensive mutagenesis studies on SubB, shows how the hydrophobic patch on top of the B-pentamer plays a dominant role in binding the A subunit. The structure of SubAB and the accompanying functional characterisation of various mutants of SubAB provide a framework for understanding the important role of the B-pentamer in the assembly and the intracellular trafficking of this AB5 toxin.
| + | |
| - | | + | |
| - | Structural basis of subtilase cytotoxin SubAB assembly.,Le Nours J, Paton AW, Byres E, Troy S, Herdman BP, Johnson MD, Paton JC, Rossjohn J, Beddoe T J Biol Chem. 2013 Aug 6. PMID:23921389<ref>PMID:23921389</ref>
| + | |
| - | | + | |
| - | From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br>
| + | |
| - | </div> | + | |
| - | <div class="pdbe-citations 4bwg" style="background-color:#fffaf0;"></div> | + | |
| | == References == | | == References == |
| | <references/> | | <references/> |
| | __TOC__ | | __TOC__ |
| | </StructureSection> | | </StructureSection> |
| - | [[Category: Escherichia coli 98nk2]] | + | [[Category: Escherichia coli 98NK2]] |
| | [[Category: Large Structures]] | | [[Category: Large Structures]] |
| - | [[Category: Beddoe, T]] | + | [[Category: Beddoe T]] |
| - | [[Category: Byres, E]] | + | [[Category: Byres E]] |
| - | [[Category: Herdman, B P]] | + | [[Category: Herdman BP]] |
| - | [[Category: Johnson, M D]] | + | [[Category: Johnson MD]] |
| - | [[Category: Nours, J Le]] | + | [[Category: Le Nours J]] |
| - | [[Category: Paton, A W]] | + | [[Category: Paton AW]] |
| - | [[Category: Paton, J C]] | + | [[Category: Paton JC]] |
| - | [[Category: Rossjohn, J]] | + | [[Category: Rossjohn J]] |
| - | [[Category: Troy, S]] | + | [[Category: Troy S]] |
| - | [[Category: Ab5 toxin]]
| + | |
| - | [[Category: Cellular trafficking]]
| + | |
| - | [[Category: Disassembly]]
| + | |
| - | [[Category: Toxin]]
| + | |
| Structural highlights
Function
[SUBA_ECOLX] Protease subunit of subtilase cytotoxin SubAB5 (PubMed:17024087). An endoprotease specific for host endoplasmic reticulum (ER) chaperone BiP/HSPA5, has no activity on human HSP70 or HSPA8 (PubMed:17024087). Cleaves between 'Leu-416' and 'Leu-417' of BiP/HSPA5 in the hinge between BiP's ATPase and protein-binding domains (PubMed:17024087). This induces host ER stress response and eventual cell death (PubMed:18005237, PubMed:18433465). Culture supernatant of E.coli expressing both subA and subB are toxic for Vero cells (African green monkey kidney cell line), Chinese hamster ovary cells and Hct-8 cells (human colonic epithelial cell line); the subunits are not toxic individually (PubMed:15226357). Purified SubAB5 is highly toxic, <0.1 pg is able to kill at least 50% of 30'000 Vero cells in a microtiter plate assay after 3 days; no cytotoxicity is seen at 24 hours (PubMed:15226357). Preabsorption with cells expressing a ganglioside GM2 mimic reduced cytotoxicity of SubAB5 by 93% in the Vero cytotoxicity assay (PubMed:15226357). Intraperitoneal injection of 200 ng of purified SubAB5 kills mice; the higher the dose the faster the mice die. Animals injected intraperitoneally with purified SubAB5 have microvascular thrombi in the brain and other organs, including the renal tubules and glomeruli (PubMed:15226357). Injection induces an unfolded response in mice (PubMed:17024087). Mice fed E.coli cells expressing cloned SubAB5 experience drastic weight loss and appear ill and lethargic (PubMed:15226357). Protein synthesis in Vero cells is transiently inhibited by SubAB5; both subunits are required for this effect (PubMed:17101670, PubMed:18005237, PubMed:18433465). Inhibition of protein synthesis is prevented by brefeldin A; cells are arrested in the G1 phase (PubMed:18005237). SubAB5 at 100 ng/ml induced caspase-dependent apoptosis in Vero cells through mitochondrial membrane damage (PubMed:19380466).[1] [2] [3] [4] [5] [6]
References
- ↑ Paton AW, Srimanote P, Talbot UM, Wang H, Paton JC. A new family of potent AB(5) cytotoxins produced by Shiga toxigenic Escherichia coli. J Exp Med. 2004 Jul 5;200(1):35-46. doi: 10.1084/jem.20040392. Epub 2004 Jun 28. PMID:15226357 doi:http://dx.doi.org/10.1084/jem.20040392
- ↑ Paton AW, Beddoe T, Thorpe CM, Whisstock JC, Wilce MC, Rossjohn J, Talbot UM, Paton JC. AB5 subtilase cytotoxin inactivates the endoplasmic reticulum chaperone BiP. Nature. 2006 Oct 5;443(7111):548-52. PMID:17024087 doi:10.1038/nature05124
- ↑ Morinaga N, Yahiro K, Matsuura G, Watanabe M, Nomura F, Moss J, Noda M. Two distinct cytotoxic activities of subtilase cytotoxin produced by shiga-toxigenic Escherichia coli. Infect Immun. 2007 Jan;75(1):488-96. doi: 10.1128/IAI.01336-06. Epub 2006 Nov 13. PMID:17101670 doi:http://dx.doi.org/10.1128/IAI.01336-06
- ↑ Morinaga N, Yahiro K, Matsuura G, Moss J, Noda M. Subtilase cytotoxin, produced by Shiga-toxigenic Escherichia coli, transiently inhibits protein synthesis of Vero cells via degradation of BiP and induces cell cycle arrest at G1 by downregulation of cyclin D1. Cell Microbiol. 2008 Apr;10(4):921-9. doi: 10.1111/j.1462-5822.2007.01094.x. Epub, 2007 Nov 14. PMID:18005237 doi:http://dx.doi.org/10.1111/j.1462-5822.2007.01094.x
- ↑ Wolfson JJ, May KL, Thorpe CM, Jandhyala DM, Paton JC, Paton AW. Subtilase cytotoxin activates PERK, IRE1 and ATF6 endoplasmic reticulum stress-signalling pathways. Cell Microbiol. 2008 Sep;10(9):1775-86. doi: 10.1111/j.1462-5822.2008.01164.x., Epub 2008 Apr 21. PMID:18433465 doi:http://dx.doi.org/10.1111/j.1462-5822.2008.01164.x
- ↑ Matsuura G, Morinaga N, Yahiro K, Komine R, Moss J, Yoshida H, Noda M. Novel subtilase cytotoxin produced by Shiga-toxigenic Escherichia coli induces apoptosis in vero cells via mitochondrial membrane damage. Infect Immun. 2009 Jul;77(7):2919-24. doi: 10.1128/IAI.01510-08. Epub 2009 Apr, 20. PMID:19380466 doi:http://dx.doi.org/10.1128/IAI.01510-08
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