1l7r: Difference between revisions
New page: left|200px<br /><applet load="1l7r" size="450" color="white" frame="true" align="right" spinBox="true" caption="1l7r, resolution 1.64Å" /> '''Tyr44Phe Mutant of B... |
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== | ==Tyr44Phe Mutant of Bacterial Cocaine Esterase cocE== | ||
The bacterial cocaine esterase, cocE, hydrolyzes cocaine faster than any | <StructureSection load='1l7r' size='340' side='right'caption='[[1l7r]], [[Resolution|resolution]] 1.64Å' scene=''> | ||
== Structural highlights == | |||
<table><tr><td colspan='2'>[[1l7r]] is a 1 chain structure with sequence from [https://en.wikipedia.org/wiki/Rhodococcus_sp._MB1 Rhodococcus sp. MB1]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1L7R OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1L7R 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]] 1.64Å</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=1l7r FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1l7r OCA], [https://pdbe.org/1l7r PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1l7r RCSB], [https://www.ebi.ac.uk/pdbsum/1l7r PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1l7r ProSAT]</span></td></tr> | |||
</table> | |||
== Function == | |||
[https://www.uniprot.org/uniprot/COCE_RHOSM COCE_RHOSM] Hydrolyzes cocaine to benzoate and ecgonine methyl ester, endowing the bacteria with the ability to utilize cocaine as a sole source of carbon and energy for growth, as this bacterium lives in the rhizosphere of coca plants. Also efficiently hydrolyzes cocaethylene, a more potent cocaine metabolite that has been observed in patients who concurrently abuse cocaine and alcohol. Is able to prevent cocaine-induced convulsions and lethality in rat.<ref>PMID:10698749</ref> <ref>PMID:16968810</ref> <ref>PMID:12369817</ref> | |||
== 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/l7/1l7r_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=1l7r ConSurf]. | |||
<div style="clear:both"></div> | |||
<div style="background-color:#fffaf0;"> | |||
== Publication Abstract from PubMed == | |||
The bacterial cocaine esterase, cocE, hydrolyzes cocaine faster than any other reported cocaine esterase. Hydrolysis of the cocaine benzoyl ester follows Michaelis-Menten kinetics with k(cat) = 7.8 s(-1) and K(M) = 640 nM. A similar rate is observed for hydrolysis of cocaethylene, a more potent cocaine metabolite that has been observed in patients who concurrently abuse cocaine and alcohol. The high catalytic proficiency, lack of observable product inhibition, and ability to hydrolyze both cocaine and cocaethylene make cocE an attractive candidate for rapid cocaine detoxification in an emergency setting. Recently, we determined the crystal structure of this enzyme, and showed that it is a serine carboxylesterase, with a catalytic triad formed by S117, H287, and D259 within a hydrophobic active site, and an oxyanion hole formed by the backbone amide of Y118 and the Y44 hydroxyl. The only enzyme previously known to use a Tyr side chain to form the oxyanion hole is prolyl oligopeptidase, but the Y44F mutation of cocE has a more deleterious effect on the specificity rate constant (k(cat)/K(M)) than the analogous Y473F mutation of prolyl oligopeptidase. Kinetic studies on a series of cocE mutants both validate the proposed mechanism, and reveal the relative contributions of active site residues toward substrate recognition and catalysis. Inspired by the anionic binding pocket of the cocaine binding antibody GNC92H2, we found that a Q55E mutation within the active site of cocE results in a modest (2-fold) improvement in K(M), but a 14-fold loss of k(cat). The pH rate profile of cocE was fit to the ionization of two groups (pK(a1) = 7.7; pK(a2) = 10.4) that likely represent titration of H287 and Y44, respectively. We also describe the crystal structures of both S117A and Y44F mutants of cocE. Finally, urea denaturation studies of cocE by fluorescence and circular dichroism show two unfolding transitions (0.5-0.6 M and 3.2-3.7 M urea), with the first transition likely representing pertubation of the active site. | |||
Biochemical characterization and structural analysis of a highly proficient cocaine esterase.,Turner JM, Larsen NA, Basran A, Barbas CF 3rd, Bruce NC, Wilson IA, Lerner RA Biochemistry. 2002 Oct 15;41(41):12297-307. PMID:12369817<ref>PMID:12369817</ref> | |||
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | |||
</div> | |||
<div class="pdbe-citations 1l7r" style="background-color:#fffaf0;"></div> | |||
==See Also== | |||
*[[Cocaine esterase|Cocaine esterase]] | |||
== References == | |||
<references/> | |||
__TOC__ | |||
</StructureSection> | |||
[[Category: Large Structures]] | |||
[[Category: Rhodococcus sp. MB1]] | |||
[[Category: Barbas III CF]] | |||
[[Category: Basran A]] | |||
[[Category: Bruce NC]] | |||
[[Category: Larsen NA]] | |||
[[Category: Lerner RA]] | |||
[[Category: Turner JM]] | |||
[[Category: Wilson IA]] | |||
