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| - | [[Image:1wuo.gif|left|200px]] | |
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| - | {{Structure
| + | ==Crystal structure of metallo-beta-lactamase IMP-1 mutant (D81A)== |
| - | |PDB= 1wuo |SIZE=350|CAPTION= <scene name='initialview01'>1wuo</scene>, resolution 2.01Å
| + | <StructureSection load='1wuo' size='340' side='right'caption='[[1wuo]], [[Resolution|resolution]] 2.01Å' scene=''> |
| - | |SITE=
| + | == Structural highlights == |
| - | |LIGAND= <scene name='pdbligand=ACY:ACETIC+ACID'>ACY</scene>, <scene name='pdbligand=CSW:CYSTEINE-S-DIOXIDE'>CSW</scene>, <scene name='pdbligand=ZN:ZINC+ION'>ZN</scene> | + | <table><tr><td colspan='2'>[[1wuo]] is a 4 chain structure with sequence from [https://en.wikipedia.org/wiki/Serratia_marcescens Serratia marcescens]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1WUO OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1WUO FirstGlance]. <br> |
| - | |ACTIVITY= <span class='plainlinks'>[http://en.wikipedia.org/wiki/Beta-lactamase Beta-lactamase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.5.2.6 3.5.2.6] </span>
| + | </td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 2.01Å</td></tr> |
| - | |GENE= BLAIMP-1 ([http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&srchmode=5&id=615 Serratia marcescens])
| + | <tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=ACY:ACETIC+ACID'>ACY</scene>, <scene name='pdbligand=CSD:3-SULFINOALANINE'>CSD</scene>, <scene name='pdbligand=ZN:ZINC+ION'>ZN</scene></td></tr> |
| - | |DOMAIN=
| + | <tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1wuo FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1wuo OCA], [https://pdbe.org/1wuo PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1wuo RCSB], [https://www.ebi.ac.uk/pdbsum/1wuo PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1wuo ProSAT]</span></td></tr> |
| - | |RELATEDENTRY=[[1wup|1WUP]]
| + | </table> |
| - | |RESOURCES=<span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=1wuo FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1wuo OCA], [http://www.ebi.ac.uk/pdbsum/1wuo PDBsum], [http://www.rcsb.org/pdb/explore.do?structureId=1wuo RCSB]</span>
| + | == Function == |
| - | }}
| + | [https://www.uniprot.org/uniprot/BLAB_SERMA BLAB_SERMA] Confers resistance to imipenem and broad-spectrum beta-lactams. Also hydrolyzes carbapenems. |
| | + | == Evolutionary Conservation == |
| | + | [[Image:Consurf_key_small.gif|200px|right]] |
| | + | Check<jmol> |
| | + | <jmolCheckbox> |
| | + | <scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/wu/1wuo_consurf.spt"</scriptWhenChecked> |
| | + | <scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview03.spt</scriptWhenUnchecked> |
| | + | <text>to colour the structure by Evolutionary Conservation</text> |
| | + | </jmolCheckbox> |
| | + | </jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1wuo ConSurf]. |
| | + | <div style="clear:both"></div> |
| | + | <div style="background-color:#fffaf0;"> |
| | + | == Publication Abstract from PubMed == |
| | + | Metallo-beta-lactamase IMP-1 is a di-Zn(II) metalloenzyme that efficiently hydrolyzes beta-lactam antibiotics. Wild-type (WT) IMP-1 has a conserved Asp-120(81) in the active site, which plays an important role in catalysis. To probe the catalytic role of Asp-120(81) in IMP-1, the IMP-1 mutants, D120(81)A and D120(81)E, were prepared by site-directed mutagenesis, and various kinetics studies were conducted. The IMP-1 mutants exhibited 10(2)-10(4)-fold drops in k(cat) values compared with WT despite the fact that they contained two Zn(II) ions in the active site. To evaluate the acid-base characteristics of Asp-120(81), the pH dependence for hydrolysis was examined by stopped-flow studies. No observable pK(a) values between pH 5 and 9 were found for WT and D120(81)A. The rapid mixing of equimolar amounts of nitrocefin and all enzymes failed to result in the detection of an anion intermediate of nitrocefin at 650 nm. These results suggest that Asp-120(81) of IMP-1 is not a factor in decreasing the pK(a) for the water bridging two Zn(II) ions and is not a proton donor to the anionic intermediate. In the case of D120(81)E, the nitrocefin hydrolysis product, which shows a maximum absorption at 460 nm, was bound to D120(81)E in the protonated form. The three-dimensional structures of D120(81)A and D120(81)E were also determined at 2.0 and 3.0 A resolutions, respectively. In the case of D120(81)E, the Zn-Zn distance was increased by 0.3 A compared with WT, due to the change in the coordination mode of Glu-120(81)OE1 and the positional shift in the conserved His-263(197) at the active site. |
| | | | |
| - | '''Crystal structure of metallo-beta-lactamase IMP-1 mutant (D81A)'''
| + | Probing the role of Asp-120(81) of metallo-beta-lactamase (IMP-1) by site-directed mutagenesis, kinetic studies, and X-ray crystallography.,Yamaguchi Y, Kuroki T, Yasuzawa H, Higashi T, Jin W, Kawanami A, Yamagata Y, Arakawa Y, Goto M, Kurosaki H J Biol Chem. 2005 May 27;280(21):20824-32. Epub 2005 Mar 23. PMID:15788415<ref>PMID:15788415</ref> |
| | | | |
| | + | From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> |
| | + | </div> |
| | + | <div class="pdbe-citations 1wuo" style="background-color:#fffaf0;"></div> |
| | | | |
| - | ==Overview== | + | ==See Also== |
| - | Metallo-beta-lactamase IMP-1 is a di-Zn(II) metalloenzyme that efficiently hydrolyzes beta-lactam antibiotics. Wild-type (WT) IMP-1 has a conserved Asp-120(81) in the active site, which plays an important role in catalysis. To probe the catalytic role of Asp-120(81) in IMP-1, the IMP-1 mutants, D120(81)A and D120(81)E, were prepared by site-directed mutagenesis, and various kinetics studies were conducted. The IMP-1 mutants exhibited 10(2)-10(4)-fold drops in k(cat) values compared with WT despite the fact that they contained two Zn(II) ions in the active site. To evaluate the acid-base characteristics of Asp-120(81), the pH dependence for hydrolysis was examined by stopped-flow studies. No observable pK(a) values between pH 5 and 9 were found for WT and D120(81)A. The rapid mixing of equimolar amounts of nitrocefin and all enzymes failed to result in the detection of an anion intermediate of nitrocefin at 650 nm. These results suggest that Asp-120(81) of IMP-1 is not a factor in decreasing the pK(a) for the water bridging two Zn(II) ions and is not a proton donor to the anionic intermediate. In the case of D120(81)E, the nitrocefin hydrolysis product, which shows a maximum absorption at 460 nm, was bound to D120(81)E in the protonated form. The three-dimensional structures of D120(81)A and D120(81)E were also determined at 2.0 and 3.0 A resolutions, respectively. In the case of D120(81)E, the Zn-Zn distance was increased by 0.3 A compared with WT, due to the change in the coordination mode of Glu-120(81)OE1 and the positional shift in the conserved His-263(197) at the active site.
| + | *[[Beta-lactamase 3D structures|Beta-lactamase 3D structures]] |
| - | | + | == References == |
| - | ==About this Structure== | + | <references/> |
| - | 1WUO is a [[Single protein]] structure of sequence from [http://en.wikipedia.org/wiki/Serratia_marcescens Serratia marcescens]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1WUO OCA].
| + | __TOC__ |
| - | | + | </StructureSection> |
| - | ==Reference==
| + | [[Category: Large Structures]] |
| - | Probing the role of Asp-120(81) of metallo-beta-lactamase (IMP-1) by site-directed mutagenesis, kinetic studies, and X-ray crystallography., Yamaguchi Y, Kuroki T, Yasuzawa H, Higashi T, Jin W, Kawanami A, Yamagata Y, Arakawa Y, Goto M, Kurosaki H, J Biol Chem. 2005 May 27;280(21):20824-32. Epub 2005 Mar 23. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/15788415 15788415]
| + | |
| - | [[Category: Beta-lactamase]] | + | |
| | [[Category: Serratia marcescens]] | | [[Category: Serratia marcescens]] |
| - | [[Category: Single protein]]
| + | [[Category: Goto M]] |
| - | [[Category: Goto, M.]] | + | [[Category: Yamagata Y]] |
| - | [[Category: Yamagata, Y.]] | + | [[Category: Yamaguchi Y]] |
| - | [[Category: Yamaguchi, Y.]] | + | |
| - | [[Category: aspartic acid]]
| + | |
| - | [[Category: hydrolysis]]
| + | |
| - | [[Category: metallo-beta-lactamase]]
| + | |
| - | | + | |
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Mon Mar 31 00:41:15 2008''
| + | |
| Structural highlights
Function
BLAB_SERMA Confers resistance to imipenem and broad-spectrum beta-lactams. Also hydrolyzes carbapenems.
Evolutionary Conservation
Check, as determined by ConSurfDB. You may read the explanation of the method and the full data available from ConSurf.
Publication Abstract from PubMed
Metallo-beta-lactamase IMP-1 is a di-Zn(II) metalloenzyme that efficiently hydrolyzes beta-lactam antibiotics. Wild-type (WT) IMP-1 has a conserved Asp-120(81) in the active site, which plays an important role in catalysis. To probe the catalytic role of Asp-120(81) in IMP-1, the IMP-1 mutants, D120(81)A and D120(81)E, were prepared by site-directed mutagenesis, and various kinetics studies were conducted. The IMP-1 mutants exhibited 10(2)-10(4)-fold drops in k(cat) values compared with WT despite the fact that they contained two Zn(II) ions in the active site. To evaluate the acid-base characteristics of Asp-120(81), the pH dependence for hydrolysis was examined by stopped-flow studies. No observable pK(a) values between pH 5 and 9 were found for WT and D120(81)A. The rapid mixing of equimolar amounts of nitrocefin and all enzymes failed to result in the detection of an anion intermediate of nitrocefin at 650 nm. These results suggest that Asp-120(81) of IMP-1 is not a factor in decreasing the pK(a) for the water bridging two Zn(II) ions and is not a proton donor to the anionic intermediate. In the case of D120(81)E, the nitrocefin hydrolysis product, which shows a maximum absorption at 460 nm, was bound to D120(81)E in the protonated form. The three-dimensional structures of D120(81)A and D120(81)E were also determined at 2.0 and 3.0 A resolutions, respectively. In the case of D120(81)E, the Zn-Zn distance was increased by 0.3 A compared with WT, due to the change in the coordination mode of Glu-120(81)OE1 and the positional shift in the conserved His-263(197) at the active site.
Probing the role of Asp-120(81) of metallo-beta-lactamase (IMP-1) by site-directed mutagenesis, kinetic studies, and X-ray crystallography.,Yamaguchi Y, Kuroki T, Yasuzawa H, Higashi T, Jin W, Kawanami A, Yamagata Y, Arakawa Y, Goto M, Kurosaki H J Biol Chem. 2005 May 27;280(21):20824-32. Epub 2005 Mar 23. PMID:15788415[1]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
See Also
References
- ↑ Yamaguchi Y, Kuroki T, Yasuzawa H, Higashi T, Jin W, Kawanami A, Yamagata Y, Arakawa Y, Goto M, Kurosaki H. Probing the role of Asp-120(81) of metallo-beta-lactamase (IMP-1) by site-directed mutagenesis, kinetic studies, and X-ray crystallography. J Biol Chem. 2005 May 27;280(21):20824-32. Epub 2005 Mar 23. PMID:15788415 doi:10.1074/jbc.M414314200
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