View source for 8akj


==Acyl-enzyme complex of cephalothin bound to deacylation mutant KPC-2 (E166Q)==
<StructureSection load='8akj' size='340' side='right'caption='[[8akj]], [[Resolution|resolution]] 1.35&Aring;' scene=''>
== Structural highlights ==
<table><tr><td colspan='2'>[[8akj]] is a 1 chain structure with sequence from [https://en.wikipedia.org/wiki/Klebsiella_pneumoniae Klebsiella pneumoniae]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=8AKJ OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=8AKJ 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]] 1.35&#8491;</td></tr>
<tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=CEO:5-METHYLENE-2-[2-OXO-1-(2-THIOPHEN-2-YL-ACETYLAMINO)-ETHYL]-5,6-DIHYDRO-2H-[1,3]THIAZINE-4-CARBOXYLIC+ACID'>CEO</scene>, <scene name='pdbligand=GOL:GLYCEROL'>GOL</scene>, <scene name='pdbligand=SO4:SULFATE+ION'>SO4</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=8akj FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=8akj OCA], [https://pdbe.org/8akj PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=8akj RCSB], [https://www.ebi.ac.uk/pdbsum/8akj PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=8akj ProSAT]</span></td></tr>
</table>
== Function ==
[https://www.uniprot.org/uniprot/BLKPC_KLEPN BLKPC_KLEPN] Hydrolyzes carbapenems, penicillins, cephalosporins and monobactams with varying efficiency.
<div style="background-color:#fffaf0;">
== Publication Abstract from PubMed ==
KPC-2 (Klebsiella pneumoniae carbapenemase-2) is a globally disseminated serine-beta-lactamase (SBL) responsible for extensive beta-lactam antibiotic resistance in Gram-negative pathogens. SBLs inactivate beta-lactams via a mechanism involving a hydrolytically labile covalent acyl-enzyme intermediate. Carbapenems, the most potent beta-lactams, evade the activity of many SBLs by forming long-lived inhibitory acyl-enzymes; however, carbapenemases such as KPC-2 efficiently deacylate carbapenem acyl-enzymes. We present high-resolution (1.25-1.4 A) crystal structures of KPC-2 acyl-enzymes with representative penicillins (ampicillin), cephalosporins (cefalothin), and carbapenems (imipenem, meropenem, and ertapenem) obtained utilizing an isosteric deacylation-deficient mutant (E166Q). The mobility of the Omega-loop (residues 165-170) negatively correlates with antibiotic turnover rates (k(cat)), highlighting the role of this region in positioning catalytic residues for efficient hydrolysis of different beta-lactams. Carbapenem-derived acyl-enzyme structures reveal the predominance of the Delta1-(2R) imine rather than the Delta2 enamine tautomer. Quantum mechanics/molecular mechanics molecular dynamics simulations of KPC-2:meropenem acyl-enzyme deacylation used an adaptive string method to differentiate the reactivity of the two isomers. These identify the Delta1-(2R) isomer as having a significantly (7 kcal/mol) higher barrier than the Delta2 tautomer for the (rate-determining) formation of the tetrahedral deacylation intermediate. Deacylation is therefore likely to proceed predominantly from the Delta2, rather than the Delta1-(2R) acyl-enzyme, facilitated by tautomer-specific differences in hydrogen-bonding networks involving the carbapenem C-3 carboxylate and the deacylating water and stabilization by protonated N-4, accumulating a negative charge on the Delta2 enamine-derived oxyanion. Taken together, our data show how the flexible Omega-loop helps confer broad-spectrum activity upon KPC-2, while carbapenemase activity stems from efficient deacylation of the Delta2-enamine acyl-enzyme tautomer.

Tautomer-Specific Deacylation and Omega-Loop Flexibility Explain the Carbapenem-Hydrolyzing Broad-Spectrum Activity of the KPC-2 beta-Lactamase.,Tooke CL, Hinchliffe P, Beer M, Zinovjev K, Colenso CK, Schofield CJ, Mulholland AJ, Spencer J J Am Chem Soc. 2023 Apr 5;145(13):7166-7180. doi: 10.1021/jacs.2c12123. Epub 2023 , Mar 27. PMID:36972204<ref>PMID:36972204</ref>

From MEDLINE&reg;/PubMed&reg;, a database of the U.S. National Library of Medicine.<br>
</div>
<div class="pdbe-citations 8akj" style="background-color:#fffaf0;"></div>
== References ==
<references/>
__TOC__
</StructureSection>
[[Category: Klebsiella pneumoniae]]
[[Category: Large Structures]]
[[Category: Hinchliffe P]]
[[Category: Spencer J]]
[[Category: Tooke CL]]