Sodium carbonate versus borate buffer for lactase quenching, laboratory work

A laboratory exercise for undergraduate advanced students of enzymology and biocatalysis is presented. Since enzyme assays can be quenched or continuous, this experiment compares the performance of two quenching agents for lactase, in a continuous setup. Enzymatic activity of β‐galactosidase (Asperg...

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Published in:Biochemistry and molecular biology education 2021-11, Vol.49 (6), p.935-941
Main Authors: Ramos‐de‐la‐Peña, Ana Mayela, Mercado‐Valenzo, Oscar Mario, Clorio‐Carrillo, Jorge Abraham, López‐Incio, Jazmin Dessiret, Monroy‐Borrego, Andrea Gabriela, Marrero‐Bretado, Mariana Monserrat, González‐Valdez, José, Aguilar, Oscar
Format: Article
Language:English
Subjects:
Advanced Courses
analytical chemistry
beta-Galactosidase
Biochemistry
Borates
Carbonates
Catalysis
College Science
Enzymatic activity
Enzymes
Humans
Laboratories
Lactase
R&D
Research & development
Science Activities
Science education
Science Instruction
Science Laboratories
Sodium carbonate
Undergraduate Students
upper‐division undergraduate
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Summary:A laboratory exercise for undergraduate advanced students of enzymology and biocatalysis is presented. Since enzyme assays can be quenched or continuous, this experiment compares the performance of two quenching agents for lactase, in a continuous setup. Enzymatic activity of β‐galactosidase (Aspergillus oryzae) was determined based on the release of 4‐nitrophenol from 4‐nitrophenyl β‐D‐galactopyranoside using a microplate reader. Sodium carbonate and borate buffer were tested as quenching agents, and experimental control was the unstopped assay. Based on released 4‐nitrophenol, enzyme activity, and rate constant k, the students could assess the performance of each termination agent. The experiment promotes disciplinary and transversal competencies, including research‐based learning, critical thinking, and introduce the students to high‐throughput techniques that are common in the research and development environment.
ISSN:1470-8175
1539-3429
DOI:10.1002/bmb.21567