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1f8w

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[[Image:1f8w.jpg|left|200px]]
 
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==CRYSTAL STRUCTURE OF NADH PEROXIDASE MUTANT: R303M==
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The line below this paragraph, containing "STRUCTURE_1f8w", creates the "Structure Box" on the page.
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<StructureSection load='1f8w' size='340' side='right'caption='[[1f8w]], [[Resolution|resolution]] 2.45&Aring;' scene=''>
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You may change the PDB parameter (which sets the PDB file loaded into the applet)
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== Structural highlights ==
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or the SCENE parameter (which sets the initial scene displayed when the page is loaded),
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<table><tr><td colspan='2'>[[1f8w]] is a 1 chain structure with sequence from [https://en.wikipedia.org/wiki/Enterococcus_faecalis Enterococcus faecalis]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1F8W OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1F8W FirstGlance]. <br>
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or leave the SCENE parameter empty for the default display.
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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]] 2.45&#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=CSX:S-OXY+CYSTEINE'>CSX</scene>, <scene name='pdbligand=FAD:FLAVIN-ADENINE+DINUCLEOTIDE'>FAD</scene></td></tr>
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{{STRUCTURE_1f8w| PDB=1f8w | SCENE= }}
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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=1f8w FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1f8w OCA], [https://pdbe.org/1f8w PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1f8w RCSB], [https://www.ebi.ac.uk/pdbsum/1f8w PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1f8w ProSAT]</span></td></tr>
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</table>
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== Function ==
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[https://www.uniprot.org/uniprot/NAPE_ENTFA NAPE_ENTFA] Peroxidase whose active site is a redox-active cysteine-sulfenic acid.
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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/f8/1f8w_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=1f8w ConSurf].
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<div style="clear:both"></div>
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'''CRYSTAL STRUCTURE OF NADH PEROXIDASE MUTANT: R303M'''
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==See Also==
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*[[NADH peroxidase|NADH peroxidase]]
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__TOC__
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==Overview==
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</StructureSection>
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The crystal structure of the flavoprotein NADH peroxidase shows that the Arg303 side chain forms a hydrogen bond with the active-site His10 imidazole and is therefore likely to influence the catalytic mechanism. Dithionite titration of an R303M mutant [E(FAD, Cys42-sulfenic acid)] yields a two-electron reduced intermediate (EH(2)) with enhanced flavin fluorescence and almost no charge-transfer absorbance at pH 7.0; the pK(a) for the nascent Cys42-SH is increased by over 3.5 units in comparison with the wild-type EH(2) pK(a) of &lt;/=4.5. NADH titration of the mutant peroxidase yields the same EH(2) intermediate, but in contrast to the behavior of wild-type enzyme, this species can be reduced directly to an EH(4).NAD(+) complex. Kinetic analyses demonstrate that the R303M mutant is severely compromised, although active, with k(cat) = 3 s(-)(1) at pH 7.0, 5 degrees C; enzyme-monitored turnover results indicate that the steady-state consists predominantly of an E-FADH(2).NAD(+) species. When the oxidized mutant is reacted anaerobically with 0.9 equiv of NADH/FAD, a clearly biphasic pattern is observed at 450 nm; relatively rapid flavin reduction is followed by reoxidation at 2.6-2.7 s(-)(1) ( approximately k(cat)). Thus replacement of Arg303 with Met leads to an altered peroxidase form in which the rate-limiting step in turnover is the intramolecular transfer of electrons from FADH(2) --&gt; Cys42-SOH. The crystal structure of the R303M peroxidase has been refined at 2.45 A resolution. In addition to eliminating the Arg303 interactions with His10 and Glu14, the mutant exhibits a significant change in the conformation of the Cys42-SOH side chain relative to FAD and His10 in particular. These and other results provide a detailed understanding of Arg303 and its role in the structure and mechanism of this unique flavoprotein peroxidase.
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==About this Structure==
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1F8W is a [[Single protein]] structure of sequence from [http://en.wikipedia.org/wiki/Enterococcus_faecalis Enterococcus faecalis]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1F8W OCA].
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==Reference==
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Analysis of the kinetic and redox properties of the NADH peroxidase R303M mutant: correlation with the crystal structure., Crane EJ 3rd, Yeh JI, Luba J, Claiborne A, Biochemistry. 2000 Aug 29;39(34):10353-64. PMID:[http://www.ncbi.nlm.nih.gov/pubmed/10956025 10956025]
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[[Category: Enterococcus faecalis]]
[[Category: Enterococcus faecalis]]
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[[Category: NADH peroxidase]]
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[[Category: Large Structures]]
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[[Category: Single protein]]
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[[Category: Claiborne A]]
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[[Category: Claiborne, A.]]
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[[Category: Hol WGJ]]
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[[Category: Hol, W G.J.]]
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[[Category: Yeh JI]]
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[[Category: Yeh, J I.]]
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[[Category: Fad]]
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[[Category: Interface]]
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[[Category: Nad-binding domain]]
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''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Fri May 2 16:02:41 2008''
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Current revision

CRYSTAL STRUCTURE OF NADH PEROXIDASE MUTANT: R303M

PDB ID 1f8w

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