Biochemical Laboratory Experiments on Polyphenol Oxidase

• Mica Cabrera ^[1] ; Sameerah Muhammad ^[1] ; Esiason Rodriguez ^[1] ; Monika Sommerhalter ^[1]
  1. [1] California State University East Bay, United States
• Localización: Journal of chemical education, ISSN 0021-9584, Vol. 101, Nº 8, 2024, págs. 3500-3505
• Idioma: inglés
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• Resumen

  • A common goal of biochemical laboratory courses is to give students hands-on experience in enzyme purification and protein characterization techniques. We present a well-tested module of five linked experiments centered on the enzyme polyphenol oxidase (PPO) from banana. This module includes buffer and plant extract preparation, protein and enzyme activity assays, column chromatography, and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Our electrophoresis experiment exploits the fact that PPO is activated by SDS for rapid and enzyme sensitive gel staining. PPO activity can be measured with various phenolic substrates. We tested different assay procedures and recommend dopamine as a substrate in a classroom setting. All five experiments, including the chromatography experiment for which we used gravity flow and syringe operated columns, provide hands-on learning experiences in essential laboratory techniques for undergraduate Biochemistry students. Our assessment of students’ learning challenges revealed that most students require assistance in interpreting and converting enzyme activity units. The use of nonstandard units in the scientific literature makes this a topic worthy of discussion.