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| [[Image:1mku.gif|left|200px]]<br /><applet load="1mku" size="350" color="white" frame="true" align="right" spinBox="true"
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| caption="1mku, resolution 1.8Å" />
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| '''CARBOXYLIC ESTER HYDROLASE, ORTHORHOMBIC FORM OF THE TRIPLE MUTANT'''<br />
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| ==Overview== | | ==CARBOXYLIC ESTER HYDROLASE, ORTHORHOMBIC FORM OF THE TRIPLE MUTANT== |
| The aspartate-99 of secreted phospholipase A2 (PLA2) has been proposed to be critical for the catalytic mechanism and interfacial activation of PLA2. Aspartate-99 connects the catalytic machinery (including the catalytic diad, the putative catalytic waters W5 and W6, and the calcium cofactor) to the hydrogen-bonding network. The latter involves Y52, Y73, the structural water, and the N-terminal region putatively required for the interfacial activation. A triple mutant of bovine pancreatic PLA2 with substitutions aspartate plus adjacent tyrosine residues (Y52,73F/D99N) was constructed, its X-ray structure was determined, and kinetic characteristics were analyzed. The kinetic properties of the D99N mutant constructed previously were also further analyzed. The X-ray structure of the Y52,73F/D99N mutant indicated a substantial disruption of the hydrogen-bonding network including the loss of the structural water similar to that seen in the structure of the D99N mutant published previously [Kumar, A., Sekharudu, Y. C., Ramakrishnan, B., Dupureur, C. M., Zhu, H., Tsai, M.-D., & Sundaralingam, M. (1994) Protein Sci. 3, 2082-2088]. Kinetic analysis demonstrated that these mutants possessed considerable catalytic activity with a k(cat) value of about 5% compared to WT. The values of the interfacial Michaelis constant were also little perturbed (ca. 4-fold lower for D99N and marginally higher for Y52,73F/D99N). The results taken together suggest that the hydrogen-bonding network is not critically important for interfacial activation. Instead, it is the chemical step that is perturbed, though only modestly, in the mutants.
| | <StructureSection load='1mku' size='340' side='right'caption='[[1mku]], [[Resolution|resolution]] 1.80Å' scene=''> |
| | == Structural highlights == |
| | <table><tr><td colspan='2'>[[1mku]] is a 1 chain structure with sequence from [https://en.wikipedia.org/wiki/Bos_taurus Bos taurus]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1MKU OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1MKU FirstGlance]. <br> |
| | </td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 1.8Å</td></tr> |
| | <tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CA:CALCIUM+ION'>CA</scene></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=1mku FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1mku OCA], [https://pdbe.org/1mku PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1mku RCSB], [https://www.ebi.ac.uk/pdbsum/1mku PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1mku ProSAT]</span></td></tr> |
| | </table> |
| | == Function == |
| | [https://www.uniprot.org/uniprot/PA21B_BOVIN PA21B_BOVIN] PA2 catalyzes the calcium-dependent hydrolysis of the 2-acyl groups in 3-sn-phosphoglycerides. |
| | == 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/mk/1mku_consurf.spt"</scriptWhenChecked> |
| | <scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.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=1mku ConSurf]. |
| | <div style="clear:both"></div> |
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| ==About this Structure== | | ==See Also== |
| 1MKU is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Bos_taurus Bos taurus] with <scene name='pdbligand=CA:'>CA</scene> as [http://en.wikipedia.org/wiki/ligand ligand]. Active as [http://en.wikipedia.org/wiki/Phospholipase_A(2) Phospholipase A(2)], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.1.1.4 3.1.1.4] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1MKU OCA].
| | *[[Phospholipase A2 3D structures|Phospholipase A2 3D structures]] |
| | | __TOC__ |
| ==Reference==
| | </StructureSection> |
| Phospholipase A2 engineering. Structural and functional roles of the highly conserved active site residue aspartate-99., Sekar K, Yu BZ, Rogers J, Lutton J, Liu X, Chen X, Tsai MD, Jain MK, Sundaralingam M, Biochemistry. 1997 Mar 18;36(11):3104-14. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=9115986 9115986]
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| [[Category: Bos taurus]] | | [[Category: Bos taurus]] |
| [[Category: Phospholipase A(2)]] | | [[Category: Large Structures]] |
| [[Category: Single protein]]
| | [[Category: Sundaralingam M]] |
| [[Category: Sundaralingam, M.]] | |
| [[Category: CA]]
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| [[Category: carboxylic ester hydrolase]]
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| [[Category: enzyme]]
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| [[Category: hydrolase]]
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| [[Category: orthorhombic form]]
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| ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 13:56:13 2008''
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