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| [[Image:1mdl.gif|left|200px]]<br /><applet load="1mdl" size="350" color="white" frame="true" align="right" spinBox="true"
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| caption="1mdl, resolution 1.85Å" />
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| '''MANDELATE RACEMASE MUTANT K166R CO-CRYSTALLIZED WITH (R)-MANDELATE'''<br />
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| ==Overview== | | ==MANDELATE RACEMASE MUTANT K166R CO-CRYSTALLIZED WITH (R)-MANDELATE== |
| On the basis of the available high-resolution structures of mandelate, racemase (MR) from Pseudomonas putida [Landro, J. A., Gerlt, J. A., Kozarich, J. W., Koo, C. W., Shah, V. J., Kenyon, G. L., Neidhart, D. J., Fujita, J., & Petsko, G. A. (1994) Biochemistry 33, 635-643], Lys 166 and, His 297 are positioned appropriately to participate in catalysis as, acid/base catalysts that either abstract the alpha-proton from the, enantiomers of mandelate to form an enolic intermediate or protonate the, enolic intermediate to form the enantiomers of mandelate, with Lys 166, participating as the (S)-specific acid/base catalyst and His 297, participating as the (R)-specific acid/base catalyst. In this paper we, report the structural and mechanistic properties of the mutant in which, Lys 166 has been replaced with arginine (K166R). The structure of K166R, has been determined at 1.85 A resolution with the substrate (S)-mandelate, bound in the active site. The structure of this complex reveals no, geometric alterations in the active site, with the exception that the, longer side chain of Arg 166 is necessarily displaced upward from the, position occupied by Lys 166 by steric interactions with the bound, substrate. In contrast to the H297N mutant of MR [Landro, J. A., Kallarakal, A. T., Ransom, S. C., Gerlt, J. A., Kozarich, J. W., Neidhart, D. J., & Kenyon, G. L. (1991) Biochemistry 30, 9275-9281], the K166R, exhibits low levels of racemase activity [kcat is reduced 5 x 10(3)-fold, in the (R)- to (S)-direction and 1 x 10(3)-fold in the (S)- to, (R)-direction]. The substrate and solvent deuterium isotope effects, support a reaction coordinate for the K166R-catalyzed reaction in which, the transition state for interconversion of bound (S)-mandelate and the, stabilized enolic intermediate is higher in energy that the transition, state for interconversion of bound (R)-mandelate and the stabilized enolic, intermediate. The solvent deuterium isotope effect when (S)-mandelate is, substrate (2.2 +/- 0.3) supports the proposal that the formation of the, enolic intermediate involves partial transfer of a solvent-derived proton, from Glu 317 to the substrate as the alpha-proton is abstracted [Mitra, B., Kallarakal, A. T., Kozarich, J. W., Gerlt, J. A., Clifton, J. G., Petsko, G. A., & Kenyon, G. L. (1995) Biochemistry 34, 2777-2787].(ABSTRACT TRUNCATED AT 400 WORDS)
| | <StructureSection load='1mdl' size='340' side='right'caption='[[1mdl]], [[Resolution|resolution]] 1.85Å' scene=''> |
| | == Structural highlights == |
| | <table><tr><td colspan='2'>[[1mdl]] is a 1 chain structure with sequence from [https://en.wikipedia.org/wiki/Pseudomonas_aeruginosa Pseudomonas aeruginosa]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1MDL OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1MDL 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]] 1.85Å</td></tr> |
| | <tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=MG:MAGNESIUM+ION'>MG</scene>, <scene name='pdbligand=RMN:(R)-MANDELIC+ACID'>RMN</scene>, <scene name='pdbligand=SMN:(S)-MANDELIC+ACID'>SMN</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=1mdl FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1mdl OCA], [https://pdbe.org/1mdl PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1mdl RCSB], [https://www.ebi.ac.uk/pdbsum/1mdl PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1mdl ProSAT]</span></td></tr> |
| | </table> |
| | == Function == |
| | [https://www.uniprot.org/uniprot/MANR_PSEPU MANR_PSEPU] |
| | == 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/md/1mdl_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=1mdl ConSurf]. |
| | <div style="clear:both"></div> |
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| ==About this Structure== | | ==See Also== |
| 1MDL is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Pseudomonas_aeruginosa Pseudomonas aeruginosa] with <scene name='pdbligand=MG:'>MG</scene>, <scene name='pdbligand=RMN:'>RMN</scene> and <scene name='pdbligand=SMN:'>SMN</scene> as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Mandelate_racemase Mandelate racemase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=5.1.2.2 5.1.2.2] Known structural/functional Sites: <scene name='pdbsite=ACT:Site+Act+Constitutes+The+Acid/Base+Catalysts+Responsible+...'>ACT</scene>, <scene name='pdbsite=CAR:Site+Car+Constitutes+The+Binding+Site+For+The+Carboxyl+G+...'>CAR</scene> and <scene name='pdbsite=MTL:Site+Mtl+Constitutes+The+Direct+Metal+Ion+Ligands'>MTL</scene>. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1MDL OCA].
| | *[[Mandelate racemase|Mandelate racemase]] |
| | | __TOC__ |
| ==Reference==
| | </StructureSection> |
| Mechanism of the reaction catalyzed by mandelate racemase: structure and mechanistic properties of the K166R mutant., Kallarakal AT, Mitra B, Kozarich JW, Gerlt JA, Clifton JG, Petsko GA, Kenyon GL, Biochemistry. 1995 Mar 7;34(9):2788-97. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=7893690 7893690]
| | [[Category: Large Structures]] |
| [[Category: Mandelate racemase]] | |
| [[Category: Pseudomonas aeruginosa]] | | [[Category: Pseudomonas aeruginosa]] |
| [[Category: Single protein]]
| | [[Category: Clifton JG]] |
| [[Category: Clifton, J.G.]] | | [[Category: Petsko GA]] |
| [[Category: Petsko, G.A.]] | |
| [[Category: MG]]
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| [[Category: RMN]]
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| [[Category: SMN]]
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| [[Category: isomerase]]
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| [[Category: magnesium]]
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| [[Category: mandelate pathway]]
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| ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sun Feb 3 09:53:28 2008''
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