We apologize for Proteopedia being slow to respond. For the past two years, a new implementation of Proteopedia has been being built. Soon, it will replace this 18-year old system. All existing content will be moved to the new system at a date that will be announced here.

1f8w

From Proteopedia

(Difference between revisions)
Jump to: navigation, search
Current revision (10:11, 20 March 2024) (edit) (undo)
 
(16 intermediate revisions not shown.)
Line 1: Line 1:
-
[[Image:1f8w.jpg|left|200px]]<br /><applet load="1f8w" size="350" color="white" frame="true" align="right" spinBox="true"
 
-
caption="1f8w, resolution 2.45&Aring;" />
 
-
'''CRYSTAL STRUCTURE OF NADH PEROXIDASE MUTANT: R303M'''<br />
 
-
==Overview==
+
==CRYSTAL STRUCTURE OF NADH PEROXIDASE MUTANT: R303M==
-
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.
+
<StructureSection load='1f8w' size='340' side='right'caption='[[1f8w]], [[Resolution|resolution]] 2.45&Aring;' scene=''>
 +
== Structural highlights ==
 +
<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>
 +
</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>
 +
<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>
 +
<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>
 +
</table>
 +
== Function ==
 +
[https://www.uniprot.org/uniprot/NAPE_ENTFA NAPE_ENTFA] Peroxidase whose active site is a redox-active cysteine-sulfenic acid.
 +
== 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/f8/1f8w_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=1f8w ConSurf].
 +
<div style="clear:both"></div>
-
==About this Structure==
+
==See Also==
-
1F8W is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Enterococcus_faecalis Enterococcus faecalis] with <scene name='pdbligand=FAD:'>FAD</scene> as [http://en.wikipedia.org/wiki/ligand ligand]. Active as [http://en.wikipedia.org/wiki/NADH_peroxidase NADH peroxidase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=1.11.1.1 1.11.1.1] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1F8W OCA].
+
*[[NADH peroxidase|NADH peroxidase]]
-
 
+
__TOC__
-
==Reference==
+
</StructureSection>
-
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://ispc.weizmann.ac.il//pmbin/getpm?pmid=10956025 10956025]
+
[[Category: Enterococcus faecalis]]
[[Category: Enterococcus faecalis]]
-
[[Category: NADH peroxidase]]
+
[[Category: Large Structures]]
-
[[Category: Single protein]]
+
[[Category: Claiborne A]]
-
[[Category: Claiborne, A.]]
+
[[Category: Hol WGJ]]
-
[[Category: Hol, W G.J.]]
+
[[Category: Yeh JI]]
-
[[Category: Yeh, J I.]]
+
-
[[Category: FAD]]
+
-
[[Category: fad]]
+
-
[[Category: interface]]
+
-
[[Category: nad-binding domains]]
+
-
 
+
-
''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 12:36:06 2008''
+

Current revision

CRYSTAL STRUCTURE OF NADH PEROXIDASE MUTANT: R303M

PDB ID 1f8w

Drag the structure with the mouse to rotate

Proteopedia Page Contributors and Editors (what is this?)

OCA

Retrieved from "http://52.214.119.220/wiki/index.php/1f8w"
Personal tools