BADAN-conjugated ?-lactamases as biosensors for ?-lactam antibiotic detection.

ABSTRACT: ?-Lactam antibiotic detection has significant implications in food safety control, environmental monitoring and pharmacokinetics study. Here, we report the development of two BADAN-conjugated ?-lactamases, E166Cb and E166Cb/N170Q, as sensitive biosensors for ?-lactam antibiotic detection. These biosensors were constructed by coupling an environment-sensitive BADAN probe onto location 166 at the active site of the PenP ?-lactamase E166C and E166C/N170Q mutants. They gave fluorescence turn-on signals in response to ?-lactam antibiotics. Molecular dynamics simulation of E166Cb suggested that the turn-on signal might be attributed to a polarity change of the microenvironment of BADAN and the removal of the fluorescence quenching effect on BADAN exerted by a nearby Tyr-105 upon the antibiotic binding. In the detection of four ?-lactams (penicillin G, penicillin V, cefotaxime and moxalactam), both E166Cb and E166Cb/N170Q delivered signal outputs in an antibiotic-concentration dependent manner with a dynamic range spanning from 10 nM to 1 ?M. Compared to E166Cb, E166Cb/N170Q generally exhibited more stable signals owing to its higher deficiency in hydrolyzing the antibiotic analyte. The overall biosensor performance of E166Cb and E166Cb/N170Q was comparable to that of their respective fluorescein-modified counterparts, E166Cf and E166Cf/N170Q. But comparatively, the BADAN-conjugated enzymes showed a higher sensitivity, displayed a faster response in detecting moxalactam and a more stable fluorescence signals towards penicillin G. This study illustrates the potential of BADAN-conjugated ?-lactamases as biosensing devices for ?-lactam antibiotics.

SUBMITTER: Au HW 

PROVIDER: S-EPMC7598492 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature