1krj

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(New page: 200px<br /><applet load="1krj" size="450" color="white" frame="true" align="right" spinBox="true" caption="1krj, resolution 2.00&Aring;" /> '''Engineering Calcium-...)
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[[Image:1krj.gif|left|200px]]<br /><applet load="1krj" size="450" color="white" frame="true" align="right" spinBox="true"
 
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caption="1krj, resolution 2.00&Aring;" />
 
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'''Engineering Calcium-binding site into Cytochrome c Peroxidase (CcP)'''<br />
 
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==Overview==
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==Engineering Calcium-binding site into Cytochrome c Peroxidase (CcP)==
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We have previously shown that the K(+) site found in ascorbate peroxidase, can be successfully engineered into the closely homologous peroxidase, cytochrome c peroxidase (CCP) (Bonagura, C. A. , Sundaramoorthy, M., Pappa, H. S., Patterson, W. R., and Poulos, T. L. (1996) Biochemistry 35, 6107-6115; Bonagura, C. A., Sundaramoorthy, M., Bhaskar, B., and Poulos, T. L. (1999) Biochemistry 38, 5538-5545). All other peroxidases bind, Ca(2+) rather than K(+). Using the K(+)-binding CCP mutant (CCPK2) as a, template protein, together with observations from structural modeling, mutants were designed that should bind Ca(2+) selectively. The crystal, structure of the first generation mutant, CCPCA1, showed that a smaller, cation, perhaps Na(+), is bound instead of Ca(2+). This is probably, because the full eight-ligand coordination sphere did not form owing to a, local disordering of one of the essential cation ligands. Based on these, observations, a second mutant, CCPCA2, was designed. The crystal structure, showed Ca(2+) binding in the CCPCA2 mutant and a well ordered, cation-binding loop with the full complement of eight protein to cation, ligands. Because cation binding to the engineered loop results in, diminished CCP activity and destabilization of the essential Trp(191), radical as measured by EPR spectroscopy, these measurements can be used as, sensitive methods for determining cation-binding selectivity. Both, activity and EPR titration studies show that CCPCA2 binds Ca(2+) more, effectively than K(+), demonstrating that an iterative protein, engineering-based approach is important in switching protein cation, selectivity.
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<StructureSection load='1krj' size='340' side='right'caption='[[1krj]], [[Resolution|resolution]] 2.00&Aring;' scene=''>
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== Structural highlights ==
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<table><tr><td colspan='2'>[[1krj]] is a 1 chain structure with sequence from [https://en.wikipedia.org/wiki/Saccharomyces_cerevisiae Saccharomyces cerevisiae]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1KRJ OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1KRJ FirstGlance]. <br>
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</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&#8491;</td></tr>
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<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=HEM:PROTOPORPHYRIN+IX+CONTAINING+FE'>HEM</scene>, <scene name='pdbligand=K:POTASSIUM+ION'>K</scene></td></tr>
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<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1krj FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1krj OCA], [https://pdbe.org/1krj PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1krj RCSB], [https://www.ebi.ac.uk/pdbsum/1krj PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1krj ProSAT]</span></td></tr>
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</table>
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== Function ==
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[https://www.uniprot.org/uniprot/CCPR_YEAST CCPR_YEAST] Destroys radicals which are normally produced within the cells and which are toxic to biological systems.
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== Evolutionary Conservation ==
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[[Image:Consurf_key_small.gif|200px|right]]
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Check<jmol>
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<jmolCheckbox>
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<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/kr/1krj_consurf.spt"</scriptWhenChecked>
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<scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview01.spt</scriptWhenUnchecked>
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<text>to colour the structure by Evolutionary Conservation</text>
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</jmolCheckbox>
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</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=1krj ConSurf].
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<div style="clear:both"></div>
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==About this Structure==
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==See Also==
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1KRJ is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Saccharomyces_cerevisiae Saccharomyces cerevisiae] with K and HEM as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Cytochrome-c_peroxidase Cytochrome-c peroxidase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.11.1.5 1.11.1.5] Full crystallographic information is available from [http://ispc.weizmann.ac.il/oca-bin/ocashort?id=1KRJ OCA].
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*[[Cytochrome c peroxidase 3D structures|Cytochrome c peroxidase 3D structures]]
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__TOC__
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==Reference==
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</StructureSection>
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Conversion of an engineered potassium-binding site into a calcium-selective site in cytochrome c peroxidase., Bonagura CA, Bhaskar B, Sundaramoorthy M, Poulos TL, J Biol Chem. 1999 Dec 31;274(53):37827-33. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=10608846 10608846]
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[[Category: Large Structures]]
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[[Category: Cytochrome-c peroxidase]]
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[[Category: Saccharomyces cerevisiae]]
[[Category: Saccharomyces cerevisiae]]
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[[Category: Single protein]]
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[[Category: Bhaskar B]]
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[[Category: Bhaskar, B.]]
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[[Category: Bonagura CA]]
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[[Category: Bonagura, C.A.]]
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[[Category: Poulos TL]]
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[[Category: Poulos, T.L.]]
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[[Category: Sundaramoorthy M]]
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[[Category: Sundaramoorthy, M.]]
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[[Category: HEM]]
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[[Category: K]]
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[[Category: bidentate bond]]
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[[Category: calcium selectivity]]
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[[Category: cation-binding loop]]
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[[Category: lignin peroxidase]]
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[[Category: open/closed conformer]]
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[[Category: pentagonal bipyrimidal geometry]]
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[[Category: trp191 cationic-radical]]
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''Page seeded by [http://ispc.weizmann.ac.il/oca OCA ] on Tue Nov 20 19:45:06 2007''
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Current revision

Engineering Calcium-binding site into Cytochrome c Peroxidase (CcP)

PDB ID 1krj

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