Protein thermal stability in the undergraduate biochemistry laboratory: Exploring protein thermal stability with...

We created a novel laboratory experience where undergraduate students explore the techniques used to study protein misfolding, unfolding, and aggregation. Despite the importance of protein misfolding and aggregation diseases, protein unfolding is not typically explored in undergraduate biochemistry laboratory classes. Yeast alcohol dehydrogenase (YADH) is used in the undergraduate biochemistry laboratory course at Miami University as the model system to explore protein overexpression and purification, bioinformatics, and enzyme characterization. Using one model protein across the entire semester allows the students to independently link topics introduced in the individual experiments; for example, students might draw connections between the thermal denaturation experiment and the requirement to keep the enzyme cold during a kinetics experiment. Students quantitated changes in secondary structure resulting from thermal denaturation by analyzing circular dichroism data. Monitoring the turbidity of a YADH solution with a temperature-controlled UV-Vis spectrometer was a reliable and easy method for undergraduate students to observe the thermally-induced aggregation of YADH. Together these experiments provide undergraduate students with first-hand experience in techniques to study protein unfolding and aggregation.