{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,29]],"date-time":"2026-09-29T10:55:08Z","timestamp":1790679308928,"version":"4.1.0"},"reference-count":31,"publisher":"American Chemical Society (ACS)","issue":"2","license":[{"start":{"date-parts":[[2015,12,9]],"date-time":"2015-12-09T00:00:00Z","timestamp":1449619200000},"content-version":"vor","delay-in-days":365,"URL":"https:\/\/pubs.acs.org\/page\/policy\/authorchoice_termsofuse.html"}],"funder":[{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["AI32956"],"award-info":[{"award-number":["AI32956"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000928","name":"Welch Foundation","doi-asserted-by":"publisher","award":["Q1279"],"award-info":[{"award-number":["Q1279"]}],"id":[{"id":"10.13039\/100000928","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2015,1,20]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>Extended-spectrum \u03b2-lactamases (ESBLs) pose a threat to public health because of their ability to confer resistance to extended-spectrum cephalosporins such as cefotaxime. The CTX-M \u03b2-lactamases are the most widespread ESBL enzymes among antibiotic resistant bacteria. Many of the active site residues are conserved between the CTX-M family and non-ESBL \u03b2-lactamases such as TEM-1, but the residues Ser237 and Arg276 are specific to the CTX-M family, suggesting that they may help to define the increased specificity for cefotaxime hydrolysis. To test this hypothesis, site-directed mutagenesis of these positions was performed in the CTX-M-14 \u03b2-lactamase. Substitutions of Ser237 and Arg276 with their TEM-1 counterparts, Ala237 and Asn276, had a modest effect on cefotaxime hydrolysis, as did removal of the Arg276 side chain in an R276A mutant. The S237A:R276N and S237A:R276A double mutants, however, exhibited 29- and 14-fold losses in catalytic efficiency for cefotaxime hydrolysis, respectively, while the catalytic efficiency for benzylpenicillin hydrolysis was unchanged. Therefore, together, the Ser237 and Arg276 residues are important contributors to the cefotaximase substrate profile of the enzyme. High-resolution crystal structures of the CTX-M-14 S70G, S70G:S237A, and S70G:S237A:R276A variants alone and in complex with cefotaxime show that residues Ser237 and Arg276 in the wild-type enzyme promote the expansion of the active site to accommodate cefotaxime and favor a conformation of cefotaxime that allows optimal contacts between the enzyme and substrate. The conservation of these residues, linked to their effects on structure and catalysis, imply that their coevolution is an important specificity determinant in the CTX-M family.<\/jats:p>","DOI":"10.1021\/bi501195g","type":"journal-article","created":{"date-parts":[[2014,12,9]],"date-time":"2014-12-09T11:07:25Z","timestamp":1418123245000},"page":"447-457","source":"Crossref","is-referenced-by-count":58,"title":["Molecular Basis for the Catalytic Specificity of the\nCTX-M Extended-Spectrum \u03b2-Lactamases"],"prefix":"10.1021","volume":"54","author":[{"given":"Carolyn J.","family":"Adamski","sequence":"first","affiliation":[{"name":"Baylor College of Medicine , , , One Baylor Plaza , , ,","place":["Houston, Texas, United States, 77030"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ana Maria","family":"Cardenas","sequence":"additional","affiliation":[{"name":"Baylor College of Medicine , , , One Baylor Plaza , , ,","place":["Houston, Texas, United States, 77030"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nicholas G.","family":"Brown","sequence":"additional","affiliation":[{"name":"Baylor College of Medicine , , , One Baylor Plaza , , ,","place":["Houston, Texas, United States, 77030"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lori B.","family":"Horton","sequence":"additional","affiliation":[{"name":"Baylor College of Medicine , , , One Baylor Plaza , , ,","place":["Houston, Texas, United States, 77030"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Banumathi","family":"Sankaran","sequence":"additional","affiliation":[{"name":"Lawrence Berkeley National Laboratory , , , ,","place":["Berkeley, California, United States, 94720"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"B. V. Venkataram","family":"Prasad","sequence":"additional","affiliation":[{"name":"Baylor College of Medicine , , , One Baylor Plaza , , ,","place":["Houston, Texas, United States, 77030"]},{"name":"Baylor College of Medicine , , , One Baylor Plaza , , ,","place":["Houston, Texas, United States, 77030"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hiram F.","family":"Gilbert","sequence":"additional","affiliation":[{"name":"Baylor College of Medicine , , , One Baylor Plaza , , ,","place":["Houston, Texas, United States, 77030"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Timothy","family":"Palzkill","sequence":"additional","affiliation":[{"name":"Baylor College of Medicine , , , One Baylor Plaza , , ,","place":["Houston, Texas, United States, 77030"]},{"name":"Baylor College of Medicine , , , One Baylor Plaza , , ,","place":["Houston, Texas, United States, 77030"]},{"name":"Baylor College of Medicine , , , One Baylor Plaza , , ,","place":["Houston, Texas, United States, 77030"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"316","published-online":{"date-parts":[[2014,12,9]]},"reference":[{"key":"2026090205570037800_cit1","doi-asserted-by":"crossref","first-page":"395","DOI":"10.1021\/cr030102i","article-title":"Bacterial resistance to beta-lactam\nantibiotics: compelling opportunism, compelling opportunity","volume":"105","author":"Fisher","year":"2005","journal-title":"Chem. Rev."},{"key":"2026090205570037800_cit2","doi-asserted-by":"crossref","first-page":"269","DOI":"10.1042\/bj2760269","article-title":"A standard\nnumbering scheme for the class A \u03b2-lactamases","volume":"276","author":"Ambler","year":"1991","journal-title":"Biochem. J."},{"key":"2026090205570037800_cit3","doi-asserted-by":"crossref","first-page":"455","DOI":"10.1146\/annurev-micro-090110-102911","article-title":"Epidemiological\nexpansion, structural studies, and\nclinical challenges of new \u03b2-lactamases from Gram-negative bacteria","volume":"65","author":"Bush","year":"2011","journal-title":"Annu. Rev. Microbiol."},{"key":"2026090205570037800_cit4","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1128\/AAC.48.1.1-14.2004","article-title":"Growing group\nof extended-spectrum beta-lactamases: the CTX-M enzymes","volume":"48","author":"Bonnet","year":"2004","journal-title":"Antimicrob. Agents Chemother."},{"key":"2026090205570037800_cit5","doi-asserted-by":"crossref","first-page":"349","DOI":"10.1016\/j.jmb.2005.02.010","article-title":"Atomic resolution\nstructures of CTX-M beta-lactamases: extended spectrum activities\nfrom increased mobility and decreased stability","volume":"348","author":"Chen","year":"2005","journal-title":"J. Mol. Biol."},{"key":"2026090205570037800_cit6","doi-asserted-by":"crossref","first-page":"1","DOI":"10.3389\/fmicb.2012.00110","article-title":"CTX-M enzymes: origin and diffusion","volume":"3","author":"Canton","year":"2012","journal-title":"Front. Microbiol"},{"key":"2026090205570037800_cit7","doi-asserted-by":"crossref","first-page":"238","DOI":"10.1038\/84981","article-title":"Predicting\nthe emergence of antibiotic resistance by directed evolution and structural\nanalysis","volume":"8","author":"Orencia","year":"2001","journal-title":"Nat. Struct. Biol."},{"key":"2026090205570037800_cit8","doi-asserted-by":"crossref","first-page":"85","DOI":"10.1016\/S0022-2836(02)00400-X","article-title":"Evolution of an\nantibiotic resistance\nenzyme constrained by stability and activity trade-offs","volume":"320","author":"Wang","year":"2002","journal-title":"J. Mol. Biol."},{"key":"2026090205570037800_cit9","doi-asserted-by":"crossref","first-page":"2079","DOI":"10.1006\/jmbi.1998.2432","article-title":"Crystal structure of the E166A mutant\nof extended-spectrum \u03b2-lactamase Toho-1 at 1.8 \u00c5 resolution","volume":"285","author":"Ibuka","year":"1999","journal-title":"J. Mol. Biol."},{"key":"2026090205570037800_cit10","doi-asserted-by":"crossref","first-page":"10634","DOI":"10.1021\/bi0342822","article-title":"Crystal structure of\nextended-spectrum beta-lactamase Toho-1: insights into the molecular\nmechanism for catalytic reaction and substrate specificity expansion","volume":"42","author":"Ibuka","year":"2003","journal-title":"Biochemistry"},{"key":"2026090205570037800_cit11","doi-asserted-by":"crossref","first-page":"108","DOI":"10.1016\/j.jmb.2010.04.062","article-title":"Structural insights into substrate\nrecognition and product expulsion in CTX-M enzymes","volume":"400","author":"Delmas","year":"2010","journal-title":"J. Mol. Biol."},{"key":"2026090205570037800_cit12","doi-asserted-by":"crossref","first-page":"192","DOI":"10.1016\/j.jmb.2007.10.026","article-title":"Structure and dynamics of CTX-M enzymes reveal insights\ninto substrate accommodation by extended-spectrum beta-lactamases","volume":"375","author":"Delmas","year":"2008","journal-title":"J. Mol. Biol."},{"key":"2026090205570037800_cit13","doi-asserted-by":"crossref","first-page":"792","DOI":"10.1093\/jac\/dkn031","article-title":"Structure-function\nstudies of arginine at position 276 in CTX-M \u03b2-lactamases","volume":"61","author":"Perez-Llarena","year":"2008","journal-title":"J. Antimicrob. Chemother."},{"key":"2026090205570037800_cit14","doi-asserted-by":"crossref","first-page":"2080","DOI":"10.1002\/pro.220","article-title":"Analysis of the plasticity of location\nof positive charge within the active site of the TEM-1 \u03b2-lactamase","volume":"18","author":"Marciano","year":"2009","journal-title":"Protein Sci."},{"key":"2026090205570037800_cit15","doi-asserted-by":"crossref","first-page":"700","DOI":"10.1038\/359700a0","article-title":"Molecular structure\nof the acyl-enzyme intermediate in \u03b2-lactam hydrolysis at 1.7\n\u00c5 resolution","volume":"359","author":"Strynadka","year":"1992","journal-title":"Nature"},{"key":"2026090205570037800_cit16","doi-asserted-by":"crossref","first-page":"32149","DOI":"10.1074\/jbc.M204212200","article-title":"The structural bases\nof antibiotic\nresistance in clinically derived mutant beta-lactamases TEM-30, TEM-32\nand TEM-34","volume":"277","author":"Wang","year":"2002","journal-title":"J. Biol. Chem."},{"key":"2026090205570037800_cit17","doi-asserted-by":"crossref","first-page":"3847","DOI":"10.1021\/bi00130a016","article-title":"Elucidation\nof the role of arginine-244 in the turnover processes of class A beta-lactamases","volume":"31","author":"Zafaralla","year":"1992","journal-title":"Biochemistry"},{"key":"2026090205570037800_cit18","first-page":"289","article-title":"Effect of substitution of Asn for\nArg-276 in the cefotaxime-hydrolyzing class A \u03b2-lactamase CTX-M-4","volume":"169","author":"Gazouli","year":"1998","journal-title":"FEMS Microbiol. Lett."},{"key":"2026090205570037800_cit19","doi-asserted-by":"crossref","first-page":"1259","DOI":"10.1128\/AAC.42.5.1259","article-title":"Sequence\nof the gene encoding a plasmid-mediated cefotaxime-hydrolyzing class\nA \u03b2-lactamase (CTX-M-4): involvement of serine 237 in cephalosporin\nhydrolysis","volume":"42","author":"Gazouli","year":"1998","journal-title":"Antimicrob. Agents Chemother."},{"key":"2026090205570037800_cit20","doi-asserted-by":"crossref","first-page":"17156","DOI":"10.1074\/jbc.M113.463521","article-title":"Identification of the\n\u03b2-lactamase inhibitor protein-II (BLIP-II) interface residues\nessential for binding affinity and specificity for class A \u03b2-lactamases","volume":"288","author":"Brown","year":"2013","journal-title":"J. Biol. Chem."},{"key":"2026090205570037800_cit21","doi-asserted-by":"crossref","DOI":"10.1002\/0471220639","volume-title":"Enzymes: A Practical Introduction to Structure, Mechanism, and Data\nAnalysis","author":"Copeland","year":"2000","edition":"2"},{"key":"2026090205570037800_cit22","doi-asserted-by":"crossref","first-page":"271","DOI":"10.1107\/S0907444910048675","article-title":"iMOSFLM:\na new graphical interface for diffraction-image processing with MOSFLM","volume":"67","author":"Battye","year":"2011","journal-title":"Acta Crystallogr., Sect. D.: Biol. Crystallogr."},{"key":"2026090205570037800_cit23","doi-asserted-by":"crossref","first-page":"760","DOI":"10.1107\/S0907444994003112","article-title":"The\nCCP4 suite: programs for protein crystallography","volume":"50","author":"Collaborative Computational Project.","year":"1994","journal-title":"Acta Crystallogr., Sect. D: Biol. Crystallogr."},{"key":"2026090205570037800_cit24","doi-asserted-by":"crossref","first-page":"1022","DOI":"10.1107\/S0021889897006766","article-title":"MOLREP: an automated program for\nmolecular replacement","volume":"30","author":"Vagin","year":"1997","journal-title":"J. Appl. Crystallogr."},{"key":"2026090205570037800_cit25","doi-asserted-by":"crossref","first-page":"658","DOI":"10.1107\/S0021889807021206","article-title":"Phaser\ncrystallographic software","volume":"40","author":"McCoy","year":"2007","journal-title":"J. Appl. 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Model."},{"key":"2026090205570037800_cit29","unstructured":"The PyMOL Molecular\nGraphics System, version 1.3, Shro\u0308dinger,\nLLC."},{"key":"2026090205570037800_cit30","doi-asserted-by":"crossref","first-page":"1605","DOI":"10.1002\/jcc.20084","article-title":"UCSF Chimera\u2500a visualization\nsystem for exploratory research and analysis","volume":"25","author":"Pettersen","year":"2004","journal-title":"J. Comput. Chem."},{"key":"2026090205570037800_cit31","doi-asserted-by":"crossref","first-page":"8509","DOI":"10.1021\/bi00489a001","article-title":"Additivity\nof mutational effects in proteins","volume":"29","author":"Wells","year":"1990","journal-title":"Biochemistry"}],"container-title":["Biochemistry"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/pubs.acs.org\/doi\/pdf\/10.1021\/bi501195g","content-type":"application\/pdf","content-version":"vor","intended-application":"unspecified"},{"URL":"https:\/\/pubs.acs.org\/bichaw\/article-pdf\/54\/2\/447\/15709786\/bi501195g.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/pubs.acs.org\/bichaw\/article-pdf\/54\/2\/447\/15709786\/bi501195g.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,9,2]],"date-time":"2026-09-02T09:57:15Z","timestamp":1788343035000},"score":1,"resource":{"primary":{"URL":"https:\/\/pubs.acs.org\/bichaw\/article\/54\/2\/447\/1478047\/Molecular-Basis-for-the-Catalytic-Specificity-of"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2014,12,9]]},"references-count":31,"journal-issue":{"issue":"2","published-online":{"date-parts":[[2014,12,9]]},"published-print":{"date-parts":[[2015,1,20]]}},"URL":"https:\/\/doi.org\/10.1021\/bi501195g","relation":{},"ISSN":["0006-2960","1520-4995"],"issn-type":[{"value":"0006-2960","type":"print"},{"value":"1520-4995","type":"electronic"}],"subject":[],"published":{"date-parts":[[2014,12,9]]}}}