1ejs: Difference between revisions

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~~[[Image:1ejs.gif|left|200px]]~~

{{Structure

==Crystal Structure of the H219N Variant of Klebsiella Aerogenes Urease==

~~|PDB~~= 1ejs |~~SIZE~~=~~350|CAPTION~~= <~~scene name~~='~~initialview01~~'>1ejs</~~scene~~>, ~~resolution~~ 2.0&~~Aring~~;

|~~SITE~~=

== Structural highlights ==

~~|LIGAND~~= <scene name='pdbligand=KCX:LYSINE+NZ-CARBOXYLIC+ACID'>KCX</scene>, <scene name='pdbligand=NI:NICKEL+(II)+ION'>NI</scene>

<table><tr><td colspan='2'>[[1ejs]] 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=1EJS OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1EJS FirstGlance]. <br>

~~|ACTIVITY~~= <~~span~~ class=~~'plainlinks'~~>~~[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]~~ </~~span~~>

</td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 2Å</td></tr>

~~|GENE=~~

<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>

~~|DOMAIN=~~

<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1ejs FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1ejs OCA], [https://pdbe.org/1ejs PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1ejs RCSB], [https://www.ebi.ac.uk/pdbsum/1ejs PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1ejs ProSAT]</span></td></tr>

</table>

~~|RESOURCES~~=<span class='plainlinks'>[~~http~~://~~oca~~.~~weizmann.ac.il/oca-docs~~/fgij/fg.htm?mol=1ejs FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1ejs OCA], [~~http~~://www.ebi.ac.uk/pdbsum/1ejs PDBsum], [~~http~~://www.~~rcsb~~.org/~~pdb~~/~~explore~~.do?~~structureId~~=1ejs ~~RCSB~~]</~~span~~>

== Function ==

}}

[https://www.uniprot.org/uniprot/URE1_KLEAE URE1_KLEAE]

== Evolutionary Conservation ==

[[Image:Consurf_key_small.gif|200px|right]]

Check<jmol>

<scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/ej/1ejs_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=1ejs ConSurf].

~~'''CRYSTAL STRUCTURE OF THE H219N VARIANT OF KLEBSIELLA AEROGENES UREASE'''~~

==See Also==

*[[Urease 3D structures|Urease 3D structures]]

__TOC__

==~~Overview~~==

</StructureSection>

Klebsiella aerogenes urease uses a dinuclear nickel active site to catalyze urea hydrolysis at >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.

~~==About this Structure==~~

~~1EJS is a~~ [[~~Protein complex~~]] structure of sequences from [http://en.wikipedia.org/wiki/Klebsiella_aerogenes Klebsiella aerogenes]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1EJS OCA].

~~==Reference==~~

~~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:~~/~~/www.ncbi.nlm.nih.gov/pubmed/10913264 10913264]~~

[[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: alpha-beta barrel]]~~

~~[[Category: nickel metalloenzyme~~]]

~~''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sun Mar 30 20:04:02 2008''~~

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@@ Line 1: / Line 1: @@
-[[Image:1ejs.gif|left|200px]]
- {{Structure
+==Crystal Structure of the H219N Variant of Klebsiella Aerogenes Urease==
-|PDB= 1ejs |SIZE=350|CAPTION= <scene name='initialview01'>1ejs</scene>, resolution 2.0&Aring;
+<StructureSection load='1ejs' size='340' side='right'caption='[[1ejs]], [[Resolution|resolution]] 2.00&Aring;' scene=''>
-|SITE=
+== Structural highlights ==
-|LIGAND= <scene name='pdbligand=KCX:LYSINE+NZ-CARBOXYLIC+ACID'>KCX</scene>, <scene name='pdbligand=NI:NICKEL+(II)+ION'>NI</scene>
+<table><tr><td colspan='2'>[[1ejs]] 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=1EJS OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1EJS FirstGlance]. <br>
-|ACTIVITY= <span class='plainlinks'>[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] </span>
+</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>
-|GENE=
+<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>
-|DOMAIN=
+<tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=1ejs FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1ejs OCA], [https://pdbe.org/1ejs PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1ejs RCSB], [https://www.ebi.ac.uk/pdbsum/1ejs PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1ejs ProSAT]</span></td></tr>
-|RELATEDENTRY=[[1ejr|1EJR]], [[1ejt|1EJT]], [[1eju|1EJU]], [[1ejv|1EJV]], [[1ejw|1EJW]], [[1ejx|1EJX]]
+</table>
-|RESOURCES=<span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=1ejs FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1ejs OCA], [http://www.ebi.ac.uk/pdbsum/1ejs PDBsum], [http://www.rcsb.org/pdb/explore.do?structureId=1ejs RCSB]</span>
+== 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/1ejs_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=1ejs ConSurf].
+<div style="clear:both"></div>
-'''CRYSTAL STRUCTURE OF THE H219N VARIANT OF KLEBSIELLA AEROGENES UREASE'''
+==See Also==
+*[[Urease 3D structures|Urease 3D structures]]
+__TOC__
-==Overview==
+</StructureSection>
-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.
-==About this Structure==
-EJS is a [[Protein complex]] structure of sequences from [http://en.wikipedia.org/wiki/Klebsiella_aerogenes Klebsiella aerogenes]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1EJS OCA].
-==Reference==
-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://www.ncbi.nlm.nih.gov/pubmed/10913264 10913264]
 [[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: alpha-beta barrel]]
-[[Category: nickel metalloenzyme]]
-''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sun Mar 30 20:04:02 2008''