Resumen de Preparative Protein Production from Inclusion Bodies and Crystallization: A Seven-Week Biochemistry Sequence

Megan J. Peterson, W. Kalani Snyder, Shelley Westerman, Benjamin J. McFarland

• We describe how to produce and purify proteins from Escherichia coli inclusion bodies by adapting versatile, preparative-scale techniques to the undergraduate laboratory schedule. This 7-week sequence of experiments fits into an annual cycle of research activity in biochemistry courses. Recombinant proteins are expressed as inclusion bodies, which are collected, washed, then solubilized in urea. Stepwise dialysis to dilute urea over the course of a week produces refolded protein. Column chromatography is used to purify protein into fractions, which are then analyzed with gel electrophoresis and concentration assays. Students culminate the project by designing crystallization trials in sitting-drop trays. Student evaluation of the experience has been positive, listing 5−12 new techniques learned, which are transferable to graduate research in academia and industry.