In order to add a biochemistry component to a junior-level integrative studio laboratory for chemistry majors, a project on enzyme kinetics using multiple assay methods was developed.  The project focuses on the proteolytic activity of caspase-3 and uses both colorimetric assays on a plate reader and calorimetric assays using isotherma titration calorimetry.  The project is broken into four phases: 1) recombinant protein expression and purification from a bacterial system, 2) enzyme kinetics characterization using a colorimetric assay, 3) enzyme kinetics comparison using a calorimetric assay, and 4) investigation of enzyme kinetics on a potential natural substrate.  As the project progresses, the students compare the two assay methods, evaluate the Michaelis constant and the substrate-binding constant results, and explore the enzyme kinetics of proteolysis on potential cellular substrates.  The project design and implementation will be discussed along with ideas for incorporation in other courses and implementing a similar project with limited instrumentation.

Design and implementation of a multi-method enzyme kinetics project for a junior-level studio laboratory

249th National Meeting (2015)

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