{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,28]],"date-time":"2026-09-28T13:31:57Z","timestamp":1790602317699,"version":"4.1.0"},"reference-count":110,"publisher":"MDPI AG","issue":"3","license":[{"start":{"date-parts":[[2014,7,1]],"date-time":"2014-07-01T00:00:00Z","timestamp":1404172800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/3.0\/"}],"funder":[{"DOI":"10.13039\/100000002","name":"NIH","doi-asserted-by":"publisher","award":["1R01AI100560"],"award-info":[{"award-number":["1R01AI100560"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000875","name":"The Pew Charitable Trusts","doi-asserted-by":"publisher","id":[{"id":"10.13039\/100000875","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Antibiotics"],"abstract":"<jats:p>The production of \u03b2-lactamase enzymes is one of the most distributed resistance mechanisms towards \u03b2-lactam antibiotics. Metallo-\u03b2-lactamases constitute a worrisome group of these kinds of enzymes, since they present a broad spectrum profile, being  able to hydrolyze not only penicillins, but also the latest generation of cephalosporins and carbapenems, which constitute at present the last resource antibiotics. The VIM, IMP, and NDM enzymes comprise the main groups of clinically relevant metallo-\u03b2-lactamases. Here we present an update of the features of the natural variants that have emerged and of the ones that have been engineered in the laboratory, in an effort to find sequence and structural determinants of substrate preferences. This knowledge is of upmost importance in novel drug design efforts. We also discuss the advances in knowledge achieved by means of in vitro directed evolution experiments, and the potential of this approach to predict natural evolution of metallo-\u03b2-lactamases.The production of \u03b2-lactamase enzymes is one of the most distributed resistance mechanisms towards \u03b2-lactam antibiotics. Metallo-\u03b2-lactamases constitute a worrisome group of these kinds of enzymes, since they present a broad spectrum profile, being  able to hydrolyze not only penicillins, but also the latest generation of cephalosporins and carbapenems, which constitute at present the last resource antibiotics. The VIM, IMP, and NDM enzymes comprise the main groups of clinically relevant metallo-\u03b2-lactamases. Here we present an update of the features of the natural variants that have emerged and of the ones that have been engineered in the laboratory, in an effort to find sequence and structural determinants of substrate preferences. This knowledge is of upmost importance in novel drug design efforts. We also discuss the advances in knowledge achieved by means of in vitro directed evolution experiments, and the potential of this approach to predict natural evolution of metallo-\u03b2-lactamases.<\/jats:p>","DOI":"10.3390\/antibiotics3030285","type":"journal-article","created":{"date-parts":[[2014,7,1]],"date-time":"2014-07-01T12:08:07Z","timestamp":1404216487000},"page":"285-316","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":82,"title":["Evolution of Metallo-\u03b2-lactamases: Trends Revealed by Natural Diversity and in vitro Evolution"],"prefix":"10.3390","volume":"3","author":[{"given":"Mar\u00eda-Roc\u00edo","family":"Meini","sequence":"first","affiliation":[{"name":"Institute of Molecular and Cellular Biology of Rosario (IBR, CONICET-UNR) and Biophysics Area, Faculty of Biochemistry and Pharmaceutical Sciences, National University of Rosario,  Ocampo y Esmeralda, CONICET-Rosario, Rosario 2000, Argentina"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Leticia","family":"Llarrull","sequence":"additional","affiliation":[{"name":"Institute of Molecular and Cellular Biology of Rosario (IBR, CONICET-UNR) and Biophysics Area, Faculty of Biochemistry and Pharmaceutical Sciences, National University of Rosario,  Ocampo y Esmeralda, CONICET-Rosario, Rosario 2000, Argentina"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alejandro","family":"Vila","sequence":"additional","affiliation":[{"name":"Institute of Molecular and Cellular Biology of Rosario (IBR, CONICET-UNR) and Biophysics Area, Faculty of Biochemistry and Pharmaceutical Sciences, National University of Rosario,  Ocampo y Esmeralda, CONICET-Rosario, Rosario 2000, Argentina"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2014,7,1]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"395","DOI":"10.1021\/cr030102i","article-title":"Bacterial resistance to beta-lactam antibiotics: Compelling opportunism, compelling opportunity","volume":"105","author":"Fisher","year":"2005","journal-title":"Chem. Rev."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"551","DOI":"10.1016\/j.mib.2010.09.008","article-title":"The future of the \u03b2-lactams","volume":"13","author":"Llarrull","year":"2010","journal-title":"Curr. Opin. Microbiol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1686","DOI":"10.1016\/j.bcp.2007.05.021","article-title":"Metallo-beta-lactamases (classification, activity, genetic organization, structure, zinc coordination) and their superfamily","volume":"74","author":"Bebrone","year":"2007","journal-title":"Biochem. Pharmacol."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"657","DOI":"10.1128\/CMR.18.4.657-686.2005","article-title":"Extended-spectrum beta-lactamases: A clinical update","volume":"18","author":"Paterson","year":"2005","journal-title":"Clin. Microbiol. Rev."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"1791","DOI":"10.3201\/eid1710.110655","article-title":"Global Spread of Carbapenemase-producing Enterobacteriaceae","volume":"17","author":"Nordmann","year":"2011","journal-title":"Emerg. Infect. Dis."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"721","DOI":"10.1021\/ar0400241","article-title":"Metallo-beta-lactamases: Novel weaponry for antibiotic resistance in bacteria","volume":"39","author":"Crowder","year":"2006","journal-title":"Acc. Chem. Res."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"15842","DOI":"10.1021\/ja801168r","article-title":"Metal content and localization during turnover in B. cereus metallo-beta-lactamase","volume":"130","author":"Llarrull","year":"2008","journal-title":"J. Am. Chem. Soc."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"15852","DOI":"10.1021\/ja801169j","article-title":"Trapping and characterization of a reaction intermediate in carbapenem hydrolysis by B. cereus metallo-beta-lactamase","volume":"130","author":"Tioni","year":"2008","journal-title":"J. Am. Chem. Soc."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"2873","DOI":"10.1128\/jb.170.6.2873-2878.1988","article-title":"Cloning, nucleotide sequence, and expression of the Bacillus cereus 5\/B\/6 beta-lactamase II structural gene","volume":"170","author":"Lim","year":"1988","journal-title":"J. Bacteriol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"1878","DOI":"10.1128\/AAC.44.7.1878-1886.2000","article-title":"Molecular and biochemical heterogeneity of class B carbapenem-hydrolyzing beta-lactamases in Chryseobacterium meningosepticum","volume":"44","author":"Bellais","year":"2000","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"199","DOI":"10.1016\/0167-4781(94)90011-6","article-title":"Sequence analysis of the L1 metallo-beta-lactamase from Xanthomonas maltophilia","volume":"1218","author":"Walsh","year":"1994","journal-title":"Biochim. Biophys. Acta"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1584","DOI":"10.1128\/AAC.43.7.1584","article-title":"Cloning and characterization of blaVIM, a new integron-borne metallo-beta-lactamase gene from a Pseudomonas aeruginosa clinical isolate","volume":"43","author":"Lauretti","year":"1999","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"902","DOI":"10.1128\/AAC.43.4.902","article-title":"Biochemical characterization of the Pseudomonas aeruginosa 101\/1477 metallo-\u03b2-lactamase IMP-1 produced by Escherichia coli","volume":"43","author":"Laraki","year":"1999","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"5046","DOI":"10.1128\/AAC.00774-09","article-title":"Characterization of a new metallo-beta-lactamase gene, bla(NDM-1), and a novel erythromycin esterase gene carried on a unique genetic structure in Klebsiella pneumoniae sequence type 14 from India","volume":"53","author":"Yong","year":"2009","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"1243","DOI":"10.4155\/fmc.13.55","article-title":"Targeting metallo-\u03b2-lactamase enzymes in antibiotic resistance","volume":"5","author":"King","year":"2013","journal-title":"Future Med. Chem."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"5171","DOI":"10.1016\/j.bmcl.2007.06.089","article-title":"A minimalistic approach to identify substrate binding features in B1 metallo-beta-lactamases","volume":"17","author":"Tomatis","year":"2007","journal-title":"Bioorg. Med. Chem. Lett."},{"key":"ref_17","first-page":"e22","article-title":"New Delhi Metallo-beta lactamase-1 containing enterobacteriaceae: Origin, diagnosis, treatment and public health concern","volume":"11","author":"Charan","year":"2012","journal-title":"Pan Afr. Med. J."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"394","DOI":"10.1038\/499394a","article-title":"Antibiotic resistance: The last resort","volume":"499","author":"McKenna","year":"2013","journal-title":"Nature"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"f1493","DOI":"10.1136\/bmj.f1493","article-title":"The true cost of antimicrobial resistance","volume":"346","author":"Smith","year":"2013","journal-title":"Br. Med. J."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"775","DOI":"10.1038\/35021219","article-title":"Molecular mechanisms that confer antibacterial drug resistance","volume":"406","author":"Walsh","year":"2000","journal-title":"Nature"},{"key":"ref_21","unstructured":"\u03b2-Lactamase classification and amino acid sequences for TEM, SHV and OXA extended-spectrum and inhibitor resistant enzymes. Available online:http:\/\/www.lahey.org\/Studies\/."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"969","DOI":"10.1128\/AAC.01009-09","article-title":"Updated functional classification of beta-lactamases","volume":"54","author":"Bush","year":"2010","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"238","DOI":"10.1038\/84981","article-title":"Predicting the emergence of antibiotic resistance by directed evolution and structural analysis","volume":"8","author":"Orencia","year":"2001","journal-title":"Nat. Struct. Biol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"389","DOI":"10.1038\/370389a0","article-title":"Rapid evolution of a protein in vitro by DNA shuffling","volume":"370","author":"Stemmer","year":"1994","journal-title":"Nature"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"775","DOI":"10.1006\/jmbi.1998.2262","article-title":"The effect of high-frequency random mutagenesis on in vitro protein evolution: A study on TEM-1 beta-lactamase","volume":"285","author":"Zaccolo","year":"1999","journal-title":"J. Mol. Biol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"1015","DOI":"10.1111\/j.1574-6976.2010.00222.x","article-title":"Natural evolution of TEM-1 \u03b2-lactamase: Experimental reconstruction and clinical relevance","volume":"34","author":"Salverda","year":"2010","journal-title":"FEMS Microbiol. Rev."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"2023","DOI":"10.1128\/AAC.44.8.2023-2027.2000","article-title":"Amino acid substitutions in a variant of IMP-1 metallo-beta-lactamase","volume":"44","author":"Iyobe","year":"2000","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"13761","DOI":"10.1073\/pnas.0503495102","article-title":"Mimicking natural evolution in metallo-beta-lactamases through second-shell ligand mutations","volume":"102","author":"Tomatis","year":"2005","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"20605","DOI":"10.1073\/pnas.0807989106","article-title":"Adaptive protein evolution grants organismal fitness by improving catalysis and flexibility","volume":"105","author":"Tomatis","year":"2008","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"1343","DOI":"10.1128\/AAC.45.5.1343-1348.2001","article-title":"Plasmid-encoded metallo-beta-lactamase (IMP-6) conferring resistance to carbapenems, especially meropenem","volume":"45","author":"Yano","year":"2001","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"2908","DOI":"10.1128\/AAC.01637-08","article-title":"Secretion of GOB metallo-beta-lactamase in Escherichia coli depends strictly on the cooperation between the cytoplasmic DnaK chaperone system and the Sec machinery: Completion of folding and Zn(II) ion acquisition occur in the bacterial periplasm","volume":"53","author":"Limansky","year":"2009","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"698","DOI":"10.1038\/nchembio.1005","article-title":"Metallo-\u03b2-lactamases withstand low Zn(II) conditions by tuning metal-ligand interactions","volume":"8","author":"Meini","year":"2012","journal-title":"Nat. Chem. Biol."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"947","DOI":"10.2217\/fmb.13.34","article-title":"Antibiotic resistance in Zn(II)-deficient environments: Metallo-\u03b2-lactamase activation in the periplasm","volume":"8","author":"Meini","year":"2013","journal-title":"Future Microbiol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"2043","DOI":"10.1016\/j.jinorgbio.2008.05.007","article-title":"Outsmarting metallo-beta-lactamases by mimicking their natural evolution","volume":"102","author":"Oelschlaeger","year":"2008","journal-title":"J. Inorg. Biochem."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"39","DOI":"10.1016\/j.meegid.2013.07.028","article-title":"An in silico approach for understanding the molecular evolution of clinically important metallo-beta-lactamases","volume":"20","author":"Pal","year":"2013","journal-title":"Infect. Genet. Evol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"3481","DOI":"10.1128\/AAC.00255-12","article-title":"Systematic analysis of metallo-\u03b2-lactamases using an automated database","volume":"56","author":"Widmann","year":"2012","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"358","DOI":"10.1016\/j.jhin.2009.10.015","article-title":"Epidemiology and clonality of multidrug-resistant Acinetobacter baumannii from a healthcare region in Hong Kong","volume":"74","author":"Ho","year":"2010","journal-title":"J. Hosp. Infect."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"3512","DOI":"10.1128\/AAC.01646-09","article-title":"Nationwide investigation of extended-spectrum \u03b2-lactamases, metallo-\u03b2-lactamases, and extended-spectrum oxacillinases produced by ceftazidime-resistant Pseudomonas aeruginosa strains in France","volume":"54","author":"Hocquet","year":"2010","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"660","DOI":"10.1128\/AAC.45.3.660-663.2001","article-title":"Metallo-beta-lactamases Working Group Standard numbering scheme for class B beta-lactamases","volume":"45","author":"Galleni","year":"2001","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"2347","DOI":"10.1128\/AAC.48.7.2347-2349.2004","article-title":"Update of the Standard Numbering Scheme for Class B \u03b2-Lactamases","volume":"48","author":"Garau","year":"2004","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1016\/S0014-5793(01)02686-2","article-title":"Expansion of the zinc metallo-hydrolase family of the beta-lactamase fold","volume":"503","author":"Daiyasu","year":"2001","journal-title":"FEBS Lett."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"687","DOI":"10.1042\/bj3630687","article-title":"Mutational analysis of the two zinc-binding sites of the Bacillus cereus 569\/H\/9 metallo-beta-lactamase","volume":"363","author":"Bebrone","year":"2002","journal-title":"Biochem. J."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"8590","DOI":"10.1021\/bi8006912","article-title":"Engineered mononuclear variants in Bacillus cereus metallo-beta-lactamase BcII are inactive","volume":"47","author":"Abriata","year":"2008","journal-title":"Biochemistry"},{"key":"ref_44","first-page":"2556","article-title":"Identification of residues critical for metallo-beta-lactamase function by codon randomization and selection","volume":"10","author":"Materon","year":"2001","journal-title":"Protein Sci."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"12404","DOI":"10.1021\/bi980506i","article-title":"Crystal structure of the zinc-dependent beta-lactamase from Bacillus cereus at 1.9 A resolution: Binuclear active site with features of a mononuclear enzyme","volume":"37","author":"Fabiane","year":"1998","journal-title":"Biochemistry"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"604","DOI":"10.1016\/j.jmb.2007.11.012","article-title":"The three-dimensional structure of VIM-2, a Zn-beta-lactamase from Pseudomonas aeruginosa in its reduced and oxidised form","volume":"375","author":"Docquier","year":"2008","journal-title":"J. Mol. Biol."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"4288","DOI":"10.1021\/bi992569m","article-title":"Crystal structure of the IMP-1 metallo beta-lactamase from Pseudomonas aeruginosa and its complex with a mercaptocarboxylate inhibitor: Binding determinants of a potent, broad-spectrum inhibitor","volume":"39","author":"Concha","year":"2000","journal-title":"Biochemistry"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"4343","DOI":"10.1128\/AAC.00409-10","article-title":"High-resolution crystal structure of the subclass B3 metallo-beta-lactamase BJP-1: Rational basis for substrate specificity and interaction with sulfonamides","volume":"54","author":"Docquier","year":"2010","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"4464","DOI":"10.1128\/AAC.00288-09","article-title":"The Structure of the dizinc subclass B2 metallo-\u03b2-lactamase CphA reveals that the second inhibitory zinc ion binds in the histidine site","volume":"53","author":"Bebrone","year":"2009","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"341","DOI":"10.1093\/protein\/gzg049","article-title":"Insight into the mechanism of the IMP-1 metallo-beta-lactamase by molecular dynamics simulations","volume":"16","author":"Oelschlaeger","year":"2003","journal-title":"Protein Eng."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"730","DOI":"10.1021\/ct300712j","article-title":"The role of the flexible L43-S54 protein loop in the CcrA metallo-\u03b2-lactamase in binding structurally dissimilar \u03b2-lactam antibiotics","volume":"9","author":"Valdez","year":"2013","journal-title":"J. Chem. Theory Comput."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"1917","DOI":"10.1096\/fj.12-224014","article-title":"NDM-1, the ultimate promiscuous enzyme: Substrate recognition and catalytic mechanism","volume":"27","author":"Kim","year":"2013","journal-title":"FASEB J."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"227","DOI":"10.1007\/978-1-62703-658-0_12","article-title":"A case study comparing quantitative stability-flexibility relationships across five metallo-\u03b2-lactamases highlighting differences within NDM-1","volume":"1084","author":"Brown","year":"2014","journal-title":"Methods Mol. Biol."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"14507","DOI":"10.1021\/bi990986t","article-title":"NMR characterization of the metallo-beta-lactamase from Bacteroides fragilis and its interaction with a tight-binding inhibitor: Role of an active-site loop","volume":"38","author":"Scrofani","year":"1999","journal-title":"Biochemistry"},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"3129","DOI":"10.1002\/anie.201310866","article-title":"Monitoring conformational changes in the NDM-1 metallo-\u03b2-lactamase by 19F-NMR spectroscopy","volume":"53","author":"Rydzik","year":"2014","journal-title":"Angew. Chem. Int. Ed."},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"890","DOI":"10.1016\/j.jmb.2006.01.003","article-title":"Crystal structure of Pseudomonas aeruginosa SPM-1 provides insights into variable zinc affinity of metallo-beta-lactamases","volume":"357","author":"Murphy","year":"2006","journal-title":"J. Mol. Biol."},{"key":"ref_57","doi-asserted-by":"crossref","first-page":"891","DOI":"10.1128\/AAC.44.4.891-897.2000","article-title":"Characterization of VIM-2, a carbapenem-hydrolyzing metallo-beta-lactamase and its plasmid- and integron-borne gene from a Pseudomonas aeruginosa clinical isolate in France","volume":"44","author":"Poirel","year":"2000","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_58","doi-asserted-by":"crossref","first-page":"455","DOI":"10.1146\/annurev-micro-090110-102911","article-title":"Epidemiological expansion, structural studies, and clinical challenges of new \u03b2-lactamases from gram-negative bacteria","volume":"65","author":"Bush","year":"2011","journal-title":"Annu. Rev. Microbiol."},{"key":"ref_59","doi-asserted-by":"crossref","first-page":"257","DOI":"10.1093\/jac\/dkg067","article-title":"On functional and structural heterogeneity of VIM-type metallo-\u03b2-lactamases","volume":"51","author":"Docquier","year":"2003","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_60","doi-asserted-by":"crossref","first-page":"1248","DOI":"10.1128\/AAC.01486-09","article-title":"Biochemical and structural characterization of the subclass B1 metallo-\u03b2-lactamase VIM-4","volume":"55","author":"Lassaux","year":"2011","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_61","doi-asserted-by":"crossref","first-page":"2905","DOI":"10.1128\/AAC.00166-08","article-title":"Kinetic characterization of VIM-7, a divergent member of the VIM metallo-beta-lactamase family","volume":"52","author":"Samuelsen","year":"2008","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_62","doi-asserted-by":"crossref","first-page":"2250","DOI":"10.1128\/AAC.01025-07","article-title":"Biochemical characterization of metallo-beta-lactamase VIM-11 from a Pseudomonas aeruginosa clinical strain","volume":"52","author":"Marchiaro","year":"2008","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_63","doi-asserted-by":"crossref","first-page":"1950","DOI":"10.1093\/jac\/dkq259","article-title":"Role of changes in the L3 loop of the active site in the evolution of enzymatic activity of VIM-type metallo-\u03b2-lactamases","volume":"65","author":"Merino","year":"2010","journal-title":"J. Antimicrob. Chemother."},{"key":"ref_64","doi-asserted-by":"crossref","first-page":"471","DOI":"10.1128\/AAC.00458-09","article-title":"VIM-19, a metallo-\u03b2-lactamase with increased carbapenemase activity from Escherichia coli and Klebsiella pneumoniae","volume":"54","author":"Nordmann","year":"2010","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_65","doi-asserted-by":"crossref","first-page":"3283","DOI":"10.1128\/AAC.06249-11","article-title":"Detection and characterization of VIM-31, a new variant of VIM-2 with Tyr224His and His252Arg mutations, in a clinical isolate of Enterobacter cloacae","volume":"56","author":"Bogaerts","year":"2012","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_66","doi-asserted-by":"crossref","first-page":"6647","DOI":"10.1021\/jm701031n","article-title":"Crystallographic investigation of the inhibition mode of a VIM-2 metallo-beta-lactamase from Pseudomonas aeruginosa by a mercaptocarboxylate inhibitor","volume":"50","author":"Yamaguchi","year":"2007","journal-title":"J. Med. Chem."},{"key":"ref_67","doi-asserted-by":"crossref","first-page":"174","DOI":"10.1016\/j.jmb.2011.05.035","article-title":"Structural and computational investigations of VIM-7: Insights into the substrate specificity of VIM metallo-\u03b2-lactamases","volume":"411","author":"Borra","year":"2011","journal-title":"J. Mol. Biol."},{"key":"ref_68","doi-asserted-by":"crossref","first-page":"26046","DOI":"10.1074\/jbc.M311373200","article-title":"Structural determinants of substrate binding to Bacillus cereus metallo-beta-lactamase","volume":"279","author":"Rasia","year":"2004","journal-title":"J. Biol. Chem."},{"key":"ref_69","doi-asserted-by":"crossref","first-page":"1188","DOI":"10.1101\/gr.849004","article-title":"WebLogo: A Sequence Logo Generator","volume":"14","author":"Crooks","year":"2004","journal-title":"Genome Res."},{"key":"ref_70","first-page":"512","article-title":"Estimation of the number of nucleotide substitutions in the control region of mitochondrial DNA in humans and chimpanzees","volume":"10","author":"Tamura","year":"1993","journal-title":"Mol. Biol. Evol."},{"key":"ref_71","doi-asserted-by":"crossref","first-page":"2725","DOI":"10.1093\/molbev\/mst197","article-title":"MEGA6: Molecular evolutionary genetics analysis version 6.0","volume":"30","author":"Tamura","year":"2013","journal-title":"Mol. Biol. Evol."},{"key":"ref_72","doi-asserted-by":"crossref","first-page":"11362","DOI":"10.1021\/ja303579d","article-title":"New Delhi metallo-\u03b2-lactamase: Structural insights into \u03b2-lactam recognition and inhibition","volume":"134","author":"King","year":"2012","journal-title":"J. Am. Chem. Soc."},{"key":"ref_73","doi-asserted-by":"crossref","first-page":"653","DOI":"10.1016\/j.jmb.2004.09.074","article-title":"Analysis of the context dependent sequence requirements of active site residues in the metallo-beta-lactamase IMP-1","volume":"344","author":"Materon","year":"2004","journal-title":"J. Mol. Biol."},{"key":"ref_74","doi-asserted-by":"crossref","first-page":"319","DOI":"10.1016\/S1074-5521(03)00070-X","article-title":"Analysis of the importance of the Metallo-\u03b2-lactamase active site loop in substrate binding and catalysis","volume":"10","author":"Moali","year":"2003","journal-title":"Chem. Biol."},{"key":"ref_75","doi-asserted-by":"crossref","first-page":"1368","DOI":"10.1110\/ps.0305303","article-title":"Role of a solvent-exposed tryptophan in the recognition and binding of antibiotic substrates for a metallo-beta-lactamase","volume":"12","author":"Huntley","year":"2003","journal-title":"Protein Sci."},{"key":"ref_76","doi-asserted-by":"crossref","first-page":"45015","DOI":"10.1074\/jbc.M107054200","article-title":"Thiomandelic acid, a broad spectrum inhibitor of Zinc \u03b2-Lactamases kinetic and spectroscopic studies","volume":"276","author":"Mollard","year":"2001","journal-title":"J. Biol. Chem."},{"key":"ref_77","doi-asserted-by":"crossref","first-page":"397","DOI":"10.1042\/BJ20131003","article-title":"Solution structures of the Bacillus cereus metallo-\u03b2-lactamase BcII and its complex with the broad spectrum inhibitor R-thiomandelic acid","volume":"456","author":"Karsisiotis","year":"2013","journal-title":"Biochem. J."},{"key":"ref_78","doi-asserted-by":"crossref","first-page":"7930","DOI":"10.1021\/bi100894r","article-title":"Evidence of adaptability in metal coordination geometry and active-site loop conformation among B1 metallo-beta-lactamases","volume":"49","author":"Buschiazzo","year":"2010","journal-title":"Biochemistry"},{"key":"ref_79","doi-asserted-by":"crossref","first-page":"3197","DOI":"10.1128\/AAC.01336-09","article-title":"Mutational analysis of VIM-2 reveals an essential determinant for Metallo-\u03b2-lactamase stability and folding","volume":"54","author":"Borgianni","year":"2010","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_80","doi-asserted-by":"crossref","first-page":"2080","DOI":"10.1002\/pro.220","article-title":"Analysis of the plasticity of location of the Arg244 positive charge within the active site of the TEM-1 beta-lactamase","volume":"18","author":"Marciano","year":"2009","journal-title":"Protein Sci."},{"key":"ref_81","doi-asserted-by":"crossref","first-page":"147","DOI":"10.1128\/AAC.35.1.147","article-title":"Transferable imipenem resistance in Pseudomonas aeruginosa","volume":"35","author":"Watanabe","year":"1991","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_82","doi-asserted-by":"crossref","first-page":"214","DOI":"10.3109\/1040841X.2011.559944","article-title":"IMP-type metallo-\u03b2-lactamases in Gram-negative bacilli: Distribution, phylogeny, and association with integrons","volume":"37","author":"Zhao","year":"2011","journal-title":"Crit. Rev. Microbiol."},{"key":"ref_83","doi-asserted-by":"crossref","first-page":"6403","DOI":"10.1128\/AAC.01440-12","article-title":"The sequence-activity relationship between metallo-\u03b2-lactamases IMP-1, IMP-6, and IMP-25 suggests an evolutionary adaptation to meropenem exposure","volume":"56","author":"Liu","year":"2012","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_84","doi-asserted-by":"crossref","first-page":"765","DOI":"10.1110\/ps.041093405","article-title":"Impact of remote mutations on metallo-beta-lactamase substrate specificity: Implications for the evolution of antibiotic resistance","volume":"14","author":"Oelschlaeger","year":"2005","journal-title":"Protein Sci."},{"key":"ref_85","doi-asserted-by":"crossref","first-page":"8945","DOI":"10.1021\/bi0300332","article-title":"Modeling domino effects in enzymes: Molecular basis of the substrate specificity of the bacterial metallo-beta-lactamases IMP-1 and IMP-6","volume":"42","author":"Oelschlaeger","year":"2003","journal-title":"Biochemistry"},{"key":"ref_86","first-page":"85","article-title":"Characterization of the active-site residues asparagine 167 and lysine 161 of the IMP-1 metallo beta-lactamase","volume":"197","author":"Haruta","year":"2001","journal-title":"FEMS Microbiol. Lett."},{"key":"ref_87","doi-asserted-by":"crossref","first-page":"11330","DOI":"10.1021\/bi0010524","article-title":"Mutational analysis of metallo-beta-lactamase CcrA from Bacteroides fragilis","volume":"39","author":"Yanchak","year":"2000","journal-title":"Biochemistry"},{"key":"ref_88","doi-asserted-by":"crossref","first-page":"5696","DOI":"10.1128\/AAC.00340-11","article-title":"Analysis of the functional contributions of Asn233 in metallo-\u03b2-lactamase IMP-1","volume":"55","author":"Brown","year":"2011","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_89","doi-asserted-by":"crossref","first-page":"5122","DOI":"10.1128\/AAC.02341-12","article-title":"Biochemical characterization of IMP-30, a metallo-\u03b2-lactamase with enhanced activity toward ceftazidime","volume":"57","author":"Pegg","year":"2013","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_90","doi-asserted-by":"crossref","first-page":"582","DOI":"10.1128\/AAC.47.2.582-587.2003","article-title":"Biochemical characterization of the acquired metallo-beta-lactamase SPM-1 from Pseudomonas aeruginosa","volume":"47","author":"Murphy","year":"2003","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_91","doi-asserted-by":"crossref","first-page":"499","DOI":"10.1099\/jmm.0.052555-0","article-title":"Global spread of antibiotic resistance: The example of New Delhi metallo-\u03b2-lactamase (NDM)-mediated carbapenem resistance","volume":"62","author":"Johnson","year":"2013","journal-title":"J. Med. Microbiol."},{"key":"ref_92","doi-asserted-by":"crossref","first-page":"657","DOI":"10.1586\/14787210.2013.811924","article-title":"Current status of carbapenemases in Latin America","volume":"11","author":"Maya","year":"2013","journal-title":"Expert Rev. Anti Infect. Ther."},{"key":"ref_93","doi-asserted-by":"crossref","first-page":"597","DOI":"10.1016\/S1473-3099(10)70143-2","article-title":"Emergence of a new antibiotic resistance mechanism in India, Pakistan, and the UK: A molecular, biological, and epidemiological study","volume":"10","author":"Kumarasamy","year":"2010","journal-title":"Lancet Infect. Dis."},{"key":"ref_94","doi-asserted-by":"crossref","first-page":"588","DOI":"10.1016\/j.tim.2011.09.005","article-title":"The emerging NDM carbapenemases","volume":"19","author":"Nordmann","year":"2011","journal-title":"Trends Microbiol."},{"key":"ref_95","doi-asserted-by":"crossref","first-page":"355","DOI":"10.1016\/S1473-3099(11)70059-7","article-title":"Dissemination of NDM-1 positive bacteria in the New Delhi environment and its implications for human health: An environmental point prevalence study","volume":"11","author":"Walsh","year":"2011","journal-title":"Lancet Infect. Dis."},{"key":"ref_96","doi-asserted-by":"crossref","first-page":"164","DOI":"10.1016\/S1473-3099(11)70048-2","article-title":"Balkan NDM-1: Escape or transplant?","volume":"11","author":"Livermore","year":"2011","journal-title":"Lancet Infect. Dis."},{"key":"ref_97","doi-asserted-by":"crossref","first-page":"3929","DOI":"10.1128\/AAC.00226-11","article-title":"Emergence of NDM-1 Metallo-\u03b2-Lactamase in Pseudomonas aeruginosa clinical isolates from Serbia","volume":"55","author":"Jovcic","year":"2011","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_98","doi-asserted-by":"crossref","first-page":"2184","DOI":"10.1128\/AAC.05961-11","article-title":"NDM-4 metallo-\u03b2-lactamase with increased carbapenemase activity from Escherichia coli","volume":"56","author":"Nordmann","year":"2012","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_99","doi-asserted-by":"crossref","first-page":"5952","DOI":"10.1128\/AAC.05108-11","article-title":"A novel variant, NDM-5, of the New Delhi metallo-\u03b2-lactamase in a multidrug-resistant Escherichia coli ST648 isolate recovered from a patient in the United Kingdom","volume":"55","author":"Hornsey","year":"2011","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_100","doi-asserted-by":"crossref","first-page":"e61322","DOI":"10.1371\/journal.pone.0061322","article-title":"First identification of novel NDM carbapenemase, NDM-7, in Escherichia coli in France","volume":"8","author":"Cuzon","year":"2013","journal-title":"PLoS One"},{"key":"ref_101","doi-asserted-by":"crossref","first-page":"2394","DOI":"10.1128\/AAC.02553-12","article-title":"NDM-8 metallo-\u03b2-lactamase in a multidrug-resistant Escherichia coli strain isolated in Nepal","volume":"57","author":"Tada","year":"2013","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_102","doi-asserted-by":"crossref","first-page":"1032","DOI":"10.1128\/AAC.48.3.1032-1033.2004","article-title":"In vitro evolution predicts that the IMP-1 metallo-beta-lactamase does not have the potential to evolve increased activity against imipenem","volume":"48","author":"Hall","year":"2004","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_103","doi-asserted-by":"crossref","first-page":"2084","DOI":"10.1128\/AAC.43.8.2084","article-title":"Role of permeability in the activities of \u03b2-Lactams against gram-negative bacteria which produce a group 3 \u03b2-Lactamase","volume":"43","author":"Matsumura","year":"1999","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_104","doi-asserted-by":"crossref","first-page":"184","DOI":"10.1016\/j.cbpa.2009.11.026","article-title":"Design strategies for the creation of artificial metalloenzymes","volume":"14","author":"Heinisch","year":"2010","journal-title":"Curr. Opin. Chem. Biol."},{"key":"ref_105","doi-asserted-by":"crossref","first-page":"201","DOI":"10.1016\/j.cbpa.2010.11.020","article-title":"Optimizing non-natural protein function with directed evolution","volume":"15","author":"Brustad","year":"2011","journal-title":"Curr. Opin. Chem. Biol."},{"key":"ref_106","doi-asserted-by":"crossref","first-page":"535","DOI":"10.1126\/science.1118953","article-title":"Design and evolution of new catalytic activity with an existing protein scaffold","volume":"311","author":"Park","year":"2006","journal-title":"Science"},{"key":"ref_107","doi-asserted-by":"crossref","first-page":"2314","DOI":"10.1074\/jbc.M112.430199","article-title":"Evolution of broad spectrum \u03b2-lactam resistance in an engineered metallo-\u03b2-lactamase","volume":"288","author":"Sun","year":"2013","journal-title":"J. Biol. Chem."},{"key":"ref_108","doi-asserted-by":"crossref","first-page":"e1003817","DOI":"10.1371\/journal.ppat.1003817","article-title":"Host-specific enzyme-substrate interactions in SPM-1 metallo-\u03b2-lactamase are modulated by second sphere residues","volume":"10","author":"Gonzalez","year":"2014","journal-title":"PLoS Pathog."},{"key":"ref_109","doi-asserted-by":"crossref","first-page":"227","DOI":"10.1016\/j.chom.2012.01.017","article-title":"Zinc sequestration by the neutrophil protein calprotectin enhances Salmonella growth in the inflamed gut","volume":"11","author":"Liu","year":"2012","journal-title":"Cell Host Microbe"},{"key":"ref_110","doi-asserted-by":"crossref","first-page":"e1003068","DOI":"10.1371\/journal.ppat.1003068","article-title":"Identification of an Acinetobacter baumannii zinc acquisition system that facilitates resistance to calprotectin-mediated zinc sequestration","volume":"8","author":"Hood","year":"2012","journal-title":"PLoS Pathog."}],"container-title":["Antibiotics"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2079-6382\/3\/3\/285\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T21:13:09Z","timestamp":1760217189000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2079-6382\/3\/3\/285"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2014,7,1]]},"references-count":110,"journal-issue":{"issue":"3","published-online":{"date-parts":[[2014,9]]}},"alternative-id":["antibiotics3030285"],"URL":"https:\/\/doi.org\/10.3390\/antibiotics3030285","relation":{},"ISSN":["2079-6382"],"issn-type":[{"value":"2079-6382","type":"electronic"}],"subject":[],"published":{"date-parts":[[2014,7,1]]}}}