biostudies-literaturePatel MPU.S. Department of EnergyHHS | NIH | National Institute of General Medical SciencesNIAID NIH HHSHHS | NIH | National Institute of Allergy and Infectious DiseasesWelch FoundationNIGMS NIH HHSNIH HHS17971-17984https://www.ebi.ac.uk/biostudies/studies/S-EPMC6240883biostudies-literatureUnknown293(46)The CTX-M β-lactamases have emerged as the most widespread extended-spectrum β-lactamases (ESBLs) in Gram-negative bacteria. These enzymes rapidly hydrolyze cefotaxime, but not the related cephalosporin, ceftazidime. ESBL variants have evolved, however, that provide enhanced ceftazidime resistance. We show here that a natural variant at a nonactive site, <i>i.e.</i> second-shell residue N106S, enhances enzyme stability but reduces catalytic efficiency for cefotaxime and ceftazidime and decreases resistance levels. However, when the N106S variant was combined with an active-site variant, D240G, that enhances enzyme catalytic efficiency, but decreases stability, the resultant double mutant exhibited higher resistance levels than predicted on the basis of the phenotypes of each variant. We foThe Journal of biological chemistrySynergistic effects of functionally distinct substitutions in β-lactamase variants shed light on the evolution of bacterial drug resistance.PMC6240883T32 AI055449R01 AI032956Q1279P30 GM124169DE-AC02–05CH11231S10 OD021832AI32956GM124169Patel MPBrown CAStojanoski VPalzkill THu LPrasad BVVSun ZAdamski CJSankaran BfalseSynergistic effects of functionally distinct substitutions in β-lactamase variants shed light on the evolution of bacterial drug resistance.The CTX-M β-lactamases have emerged as the most widespread extended-spectrum β-lactamases (ESBLs) in Gram-negative bacteria. These enzymes rapidly hydrolyze cefotaxime, but not the related cephalosporin, ceftazidime. ESBL variants have evolved, however, that provide enhanced ceftazidime resistance. We show here that a natural variant at a nonactive site, <i>i.e.</i> second-shell residue N106S, enhances enzyme stability but reduces catalytic efficiency for cefotaxime and ceftazidime and decreases resistance levels. However, when the N106S variant was combined with an active-site variant, D240G, that enhances enzyme catalytic efficiency, but decreases stability, the resultant double mutant exhibited higher resistance levels than predicted on the basis of the phenotypes of each variant. We fo2018-01-01T00:00:00Z2018 Nov2026-04-29T03:32:18.068Z2024-10-18T12:26:17.583ZS-EPMC62408833027501310.1074/jbc.ra118.00379210.1074/jbc.RA118.003792