On the active site of mononuclear B1 metallo βlac

Authors: Jacopo Sgrignani Alessandra Magistrato Matteo Dal Peraro Alejandro J Vila Paolo Carloni Roberta Pierattelli

Publish Date: 2012/04/25

Volume: 26, Issue: 4, Pages: 425-435

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Abstract

Metalloβlactamases MβLs are ZnIIbased bacterial enzymes that hydrolyze βlactam antibiotics hampering their beneficial effects In the most relevant subclass B1 Xray crystallography studies on the enzyme from Bacillus Cereus point to either two zinc ions in two metal sites the socalled ‘3H’ and ‘DCH’ sites or a single ZnII ion in the 3H site where the ion is coordinated by Asp120 Cys221 and His263 residues However spectroscopic studies on the B1 enzyme from B Cereus in the monozinc form suggested the presence of the ZnII ion also in the DCH site where it is bound to an aspartate a cysteine a histidine and a water molecule A structural model of this enzyme in its DCH mononuclear form so far lacking is therefore required for inhibitor design and mechanistic studies By using force field based and mixed quantum–classical QM/MM molecular dynamics MD simulations of the protein in aqueous solution we constructed such structural model The geometry and the Hbond network at the catalytic site of this model in the free form and in complex with two common βlactam drugs is compared with experimental and theoretical findings of CphA and the recently solved crystal structure of new B2 MβL from Serratia fonticola SfhI These are MβLs from the B2 subclass which features an experimentally well established monozinc form in which the ZnII is located in the DCH site From our simulations the εεδ and δεδ protomers emerge as possible DCH monozinc reactive species giving a novel contribution to the discussion on the MβL reactivity and to the drug design processThis work has been supported in part by the FIRB project Contract RBLA032ZM7 by the EC contract SPINE II n° 031220 and by grants from HHMI Howard Hughes Medical Instituteand ANPCyT Agencia Nacional de Promoción Científica y Tecnológica to AJV AJV is a fellow of the John Simon Guggenheim Foundation and an HHMI International Research Scholar Access to the computational resources supplied by CASPUR and by CINECA is gratefully acknowledged

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