Enzyme recognition element-based biosensors are very attractive for biosensor application due to a variety of measurable reaction products arising from a catalytic process. In this study, biosensor recognition elements have been developed via engineer bacterial enzymes (carboxylesterases (CEs)) which will used for narcotic detection because of their role in narcotics metabolism. The modification (insertion of cys-tag) allows the enzyme to bind into a transducer surface of a biosensor which will translate the reaction product into the detection system. The results demonstrate the successful isolation, cloning, expression, and purification of recombinant (pnbA1 and pnbA2), and engineered (pnbA1-cys and pnbA2-cys) bacterial carboxylesterases. Enzyme capability to hydrolyse cocaine into benzoylecgonine and methanol was quantified using HPLC. Both enzymes showed broad maximal activity between pH (8.0, 8.5, and 9.0), PnbA1 temperature stability ranging between (25 and 45 °C); however, PnbA2 stability range was (25-40 °C). Insertion of cys-tag at the N-terminal of the enzyme did not limit entrance to the active site which is located at the base of a cavity with dimensions 20 by 13 by 18 Å, and did not prevent substrate hydrolysis. Bacterial carboxylesterases pnbA1 and pnbA2 mimic hCE1 and not hCE2 in its reaction pathways hydrolysing cocaine into benzoylecgonine and methanol rather than ecgonine methyl ester and benzoic acid. These results are the first experimental evidence confirming the capability of bacterial carboxylesterase to hydrolyse cocaine into its main metabolites, therefore opening up the possibility to use these enzymes in numerous biotechnological applications in addition to a cocaine biosensor.
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PLoS One
December 2024
School of Biological Sciences, Universiti Sains Malaysia, Gelugor, Penang, Malaysia.
This study focuses on a novel lipase from Bacillus licheniformis IBRL-CHS2. The lipase gene was cloned into the pGEM-T Easy vector, and its sequences were registered in GenBank (KU984433 and AOT80658). It was identified as a member of the bacterial lipase subfamily 1.
View Article and Find Full Text PDFFood Res Int
December 2024
Department of Food Science and Nutrition, Faculty of Food Engineering, University of Campinas, Campinas, SP, Brazil. Electronic address:
Ensuring microbiological safety in fruit juices while maintaining their nutritional and sensory qualities remains a significant challenge in food processing. Traditional thermal methods, although effective against vegetative pathogens, can degrade important nutrients and are less effective at inactivating bacterial spores. High-pressure carbon dioxide (HPCD) technology has emerged as a promising non-thermal alternative, using CO under high pressure to inactivate spores and enzymes.
View Article and Find Full Text PDFFood Chem
February 2025
College of Landscape Architecture and Arts, Northwest A&F University, Yangling 712100, China. Electronic address:
Although the health benefits of chrysanthemums have been widely studied, there is a paucity of knowledge regarding Taihang chrysanthemum (Opisthopappus taihangensis). This study compared indoor shade drying, heat drying and freeze drying on the chemical profile and health-related activities of O. taihangensis.
View Article and Find Full Text PDFInt J Biol Macromol
January 2025
Department of Agricultural Chemistry, National Taiwan University, Taipei 10617, Taiwan; Institute of Biochemical Sciences, National Taiwan University, Taipei 10617, Taiwan; Genome and Systems Biology Degree Program, National Taiwan University and Academia Sinica, Taipei 10617, Taiwan; Center for Computational and Systems Biology, National Taiwan University, Taipei 10617, Taiwan. Electronic address:
Anal Chem
December 2024
Collaborative Innovation Centre of Henan Province for Green Manufacturing of Fine Chemicals; Key Laboratory of Green Chemical Media and Reactions, Ministry of Education; Henan International Joint Laboratory of Smart Molecules and Identification and Diagnostic Functions; School of Chemistry and Chemical Engineering, Henan Normal University, Xinxiang, Henan 453007, P. R. China.
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