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[[Image:1ejt.gif|left|200px]]<br /><applet load="1ejt" size="350" color="white" frame="true" align="right" spinBox="true"
caption="1ejt, resolution 2.0&Aring;" />
'''CRYSTAL STRUCTURE OF THE H219Q VARIANT OF KLEBSIELLA AEROGENES UREASE'''<br />


==Overview==
==CRYSTAL STRUCTURE OF THE H219Q VARIANT OF KLEBSIELLA AEROGENES UREASE==
Klebsiella aerogenes urease uses a dinuclear nickel active site to catalyze urea hydrolysis at &gt;10(14)-fold the spontaneous rate. To better define the enzyme mechanism, we examined the kinetics and structures for a suite of site-directed variants involving four residues at the active site: His320, His219, Asp221, and Arg336. Compared to wild-type urease, the H320A, H320N, and H320Q variants exhibit similar approximately 10(-)(5)-fold deficiencies in rates, modest K(m) changes, and disorders in the peptide flap covering their active sites. The pH profiles for these mutant enzymes are anomalous with optima near 6 and shoulders that extend to pH 9. H219A urease exhibits 10(3)-fold increased K(m) over that of native enzyme, whereas the increase is less marked ( approximately 10(2)-fold) in the H219N and H219Q variants that retain hydrogen bonding capability. Structures for these variants show clearly resolved active site water molecules covered by well-ordered peptide flaps. Whereas the D221N variant is only moderately affected compared to wild-type enzyme, D221A urease possesses low activity ( approximately 10(-)(3) that of native enzyme), a small increase in K(m), and a pH 5 optimum. The crystal structure for D221A urease is reminiscent of the His320 variants. The R336Q enzyme has a approximately 10(-)(4)-fold decreased catalytic rate with near-normal pH dependence and an unaffected K(m). Phenylglyoxal inactivates the R336Q variant at over half the rate observed for native enzyme, demonstrating that modification of non-active-site arginines can eliminate activity, perhaps by affecting the peptide flap. Our data favor a mechanism in which His219 helps to polarize the substrate carbonyl group, a metal-bound terminal hydroxide or bridging oxo-dianion attacks urea to form a tetrahedral intermediate, and protonation occurs via the general acid His320 with Asp221 and Arg336 orienting and influencing the acidity of this residue. Furthermore, we conclude that the simple bell-shaped pH dependence of k(cat) and k(cat)/K(m) for the native enzyme masks a more complex underlying pH dependence involving at least four pK(a)s.
<StructureSection load='1ejt' size='340' side='right'caption='[[1ejt]], [[Resolution|resolution]] 2.00&Aring;' scene=''>
== Structural highlights ==
<table><tr><td colspan='2'>[[1ejt]] is a 3 chain structure with sequence from [https://en.wikipedia.org/wiki/Klebsiella_aerogenes Klebsiella aerogenes]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1EJT OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1EJT 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&#8491;</td></tr>
<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=KCX:LYSINE+NZ-CARBOXYLIC+ACID'>KCX</scene>, <scene name='pdbligand=NI:NICKEL+(II)+ION'>NI</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=1ejt FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1ejt OCA], [https://pdbe.org/1ejt PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1ejt RCSB], [https://www.ebi.ac.uk/pdbsum/1ejt PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1ejt ProSAT]</span></td></tr>
</table>
== Function ==
[https://www.uniprot.org/uniprot/URE1_KLEAE URE1_KLEAE]
== 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/ej/1ejt_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=1ejt ConSurf].
<div style="clear:both"></div>


==About this Structure==
==See Also==
1EJT is a [http://en.wikipedia.org/wiki/Protein_complex Protein complex] structure of sequences from [http://en.wikipedia.org/wiki/Klebsiella_aerogenes Klebsiella aerogenes] with <scene name='pdbligand=NI:'>NI</scene> as [http://en.wikipedia.org/wiki/ligand ligand]. Active as [http://en.wikipedia.org/wiki/Urease Urease], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.5.1.5 3.5.1.5] Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1EJT OCA].
*[[Urease 3D structures|Urease 3D structures]]
 
__TOC__
==Reference==
</StructureSection>
Kinetic and structural characterization of urease active site variants., Pearson MA, Park IS, Schaller RA, Michel LO, Karplus PA, Hausinger RP, Biochemistry. 2000 Jul 25;39(29):8575-84. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=10913264 10913264]
[[Category: Klebsiella aerogenes]]
[[Category: Klebsiella aerogenes]]
[[Category: Protein complex]]
[[Category: Large Structures]]
[[Category: Urease]]
[[Category: Hausinger RP]]
[[Category: Hausinger, R P.]]
[[Category: Karplus PA]]
[[Category: Karplus, P A.]]
[[Category: Michel LO]]
[[Category: Michel, L O.]]
[[Category: Park IS]]
[[Category: Park, I S.]]
[[Category: Pearson MA]]
[[Category: Pearson, M A.]]
[[Category: Schaller RA]]
[[Category: Schaller, R A.]]
[[Category: NI]]
[[Category: alpha-beta barrel]]
[[Category: nickel metalloenzyme]]
 
''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 12:28:30 2008''

Latest revision as of 13:03, 20 March 2024

CRYSTAL STRUCTURE OF THE H219Q VARIANT OF KLEBSIELLA AEROGENES UREASECRYSTAL STRUCTURE OF THE H219Q VARIANT OF KLEBSIELLA AEROGENES UREASE

Structural highlights

1ejt is a 3 chain structure with sequence from Klebsiella aerogenes. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance.
Method:X-ray diffraction, Resolution 2Å
Ligands:,
Resources:FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT

Function

URE1_KLEAE

Evolutionary Conservation

Check, as determined by ConSurfDB. You may read the explanation of the method and the full data available from ConSurf.

See Also

1ejt, resolution 2.00Å

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