Exploring the pH Dependence of L-leucine-p-nitroanilide Cleavage by AminopeptidaseAeromonasProteolytica: A Combined Buffer-Enzyme Kinetics Experiment for the General Chemistry Laboratory
Laboratories and demonstrations, general chemistry
The concepts of weak acids and bases, and buffers are often difficult ones for general chemistry students to grasp and appreciate. In order to assist our students in learning these concepts and understanding their significance in nature and research, we have developed a two-lab period experiment that combines the preparation and characterization of buffer solutions followed by their use in conducting a simple study of the effect of pH on the activity of an enzyme. Through this experiment, students gain experience with pH titrations, analyzing acid/base titration curves, preparing buffer solutions, and conducting colorimetric assays of an enzyme using a spectrophotometer. Though the experiment described here is geared for general chemistry students, it is readily expanded for use in an upper-level biochemistry class by examining other factors that affect the activity of the enzyme including the presence of inhibitors.
The Chemical Educator
Frey, M. W.; Frey, S. T.; Soltau, S. R., Exploring the pH dependence of L-leucine-p-nitroanilide cleavage by aminopeptidase aeromonas proteolytica: a combined buffer-enzyme kinetics experiment for the general chemistry laboratory. Chem. Educ. 2010, 15, 117-120.