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| | ==Crystal structure of the CFTR inhibitory factor Cif with the E153Q mutation adducted with the epibromohydrin hydrolysis intermediate== | | ==Crystal structure of the CFTR inhibitory factor Cif with the E153Q mutation adducted with the epibromohydrin hydrolysis intermediate== |
| - | <StructureSection load='4dnf' size='340' side='right' caption='[[4dnf]], [[Resolution|resolution]] 1.80Å' scene=''> | + | <StructureSection load='4dnf' size='340' side='right'caption='[[4dnf]], [[Resolution|resolution]] 1.80Å' scene=''> |
| | == Structural highlights == | | == Structural highlights == |
| - | <table><tr><td colspan='2'>[[4dnf]] is a 4 chain structure with sequence from [http://en.wikipedia.org/wiki/Pseab Pseab]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4DNF OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=4DNF FirstGlance]. <br> | + | <table><tr><td colspan='2'>[[4dnf]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Pseudomonas_aeruginosa_UCBPP-PA14 Pseudomonas aeruginosa UCBPP-PA14]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=4DNF OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=4DNF FirstGlance]. <br> |
| - | </td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat"><scene name='pdbligand=EBH:(2S)-1-BROMOPROPAN-2-OL'>EBH</scene></td></tr> | + | </td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=EBH:(2S)-1-BROMOPROPAN-2-OL'>EBH</scene></td></tr> |
| - | <tr id='related'><td class="sblockLbl"><b>[[Related_structure|Related:]]</b></td><td class="sblockDat">[[3kd2|3kd2]], [[3kda|3kda]], [[3pi6|3pi6]], [[4dln|4dln]], [[4dm7|4dm7]], [[4dmc|4dmc]], [[4dmf|4dmf]], [[4dmh|4dmh]], [[4dmk|4dmk]], [[4dno|4dno]]</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=4dnf FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=4dnf OCA], [https://pdbe.org/4dnf PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=4dnf RCSB], [https://www.ebi.ac.uk/pdbsum/4dnf PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=4dnf ProSAT]</span></td></tr> |
| - | <tr id='gene'><td class="sblockLbl"><b>[[Gene|Gene:]]</b></td><td class="sblockDat">PA14_26090 ([http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&srchmode=5&id=208963 PSEAB])</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=4dnf FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=4dnf OCA], [http://pdbe.org/4dnf PDBe], [http://www.rcsb.org/pdb/explore.do?structureId=4dnf RCSB], [http://www.ebi.ac.uk/pdbsum/4dnf PDBsum], [http://prosat.h-its.org/prosat/prosatexe?pdbcode=4dnf ProSAT]</span></td></tr> | + | |
| | </table> | | </table> |
| | + | == Function == |
| | + | [[https://www.uniprot.org/uniprot/A0A0H2ZD27_PSEAB A0A0H2ZD27_PSEAB]] |
| | <div style="background-color:#fffaf0;"> | | <div style="background-color:#fffaf0;"> |
| | == Publication Abstract from PubMed == | | == Publication Abstract from PubMed == |
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| | </div> | | </div> |
| | <div class="pdbe-citations 4dnf" style="background-color:#fffaf0;"></div> | | <div class="pdbe-citations 4dnf" style="background-color:#fffaf0;"></div> |
| | + | |
| | + | ==See Also== |
| | + | *[[CFTR inhibitory factor|CFTR inhibitory factor]] |
| | == References == | | == References == |
| | <references/> | | <references/> |
| | __TOC__ | | __TOC__ |
| | </StructureSection> | | </StructureSection> |
| - | [[Category: Pseab]] | + | [[Category: Large Structures]] |
| - | [[Category: Bahl, C D]] | + | [[Category: Pseudomonas aeruginosa UCBPP-PA14]] |
| - | [[Category: Madden, D R]] | + | [[Category: Bahl CD]] |
| - | [[Category: Alpha beta hydrolase]] | + | [[Category: Madden DR]] |
| - | [[Category: Covalently adducted with the epibromohydrin hydrolysis intermediate]]
| + | |
| - | [[Category: Epoxide hydrolase]]
| + | |
| - | [[Category: Hydrolase]]
| + | |
| - | [[Category: Secreted]]
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| Structural highlights
Function
[A0A0H2ZD27_PSEAB]
Publication Abstract from PubMed
The CFTR inhibitory factor (Cif) is an epoxide hydrolase (EH) virulence factor secreted by the bacterium Pseudomonas aeruginosa. Sequence alignments reveal a pattern of Cif-like substitutions that proved to be characteristic of a new subfamily of bacterial EHs. At the same time, crystallographic and mutagenetic data suggest that EH activity is required for virulence and that Cif's active site remains generally compatible with a canonical two-step EH mechanism. A hallmark of this mechanism is the formation of a covalent hydroxyalkyl-enzyme intermediate by nucleophilic attack. In several well-studied EHs, this intermediate has been captured at near stoichiometric levels, presumably reflecting rate-limiting hydrolysis. Here we show by mass spectrometry that only minimal levels of the expected intermediate can be trapped with WT Cif. In contrast, substantial amounts of intermediate are recovered from an active-site mutant (Cif-E153Q) that selectively targets the second, hydrolytic release step. Utilizing Cif-E153Q and a previously reported nucleophile mutant (Cif-D129S), we then captured Cif in the substrate-bound, hydroxyalkyl-intermediate, and product-bound states for 1,2-epoxyhexane, yielding the first crystallographic snapshots of an EH at these key stages along the reaction coordinate. Taken together, our data illuminate the proposed two-step hydrolytic mechanism of a new class of bacterial virulence factor. They also suggest that the failure of WT Cif to accumulate a covalent hydroxyalkyl-enzyme intermediate reflects an active-site chemistry in which hydrolysis is no longer the rate-limiting step, a noncanonical kinetic regime that may explain similar observations with a number of other EHs.
Visualizing the Mechanism of Epoxide Hydrolysis by the Bacterial Virulence Enzyme Cif.,Bahl CD, Hvorecny KL, Morisseau C, Gerber SA, Madden DR Biochemistry. 2016 Feb 9;55(5):788-97. doi: 10.1021/acs.biochem.5b01229. Epub, 2016 Jan 22. PMID:26752215[1]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
See Also
References
- ↑ Bahl CD, Hvorecny KL, Morisseau C, Gerber SA, Madden DR. Visualizing the Mechanism of Epoxide Hydrolysis by the Bacterial Virulence Enzyme Cif. Biochemistry. 2016 Feb 9;55(5):788-97. doi: 10.1021/acs.biochem.5b01229. Epub, 2016 Jan 22. PMID:26752215 doi:http://dx.doi.org/10.1021/acs.biochem.5b01229
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