This course is for scientists, including graduate students, postdoctoral scholars, staff scientists, and principal investigators, who want a rigorous introduction to expression and purification of proteins as well as analysis of protein structure and function.
Through hands-on experience in the lab as well as extensive lecture and discussion, each student will become familiar with key approaches in expression, purification, and analysis of soluble and membrane proteins and protein complexes from both natural sources and overexpression systems. The emphasis of the course is on the following:
1) Approaches in protein expression: Choosing the best bacterial or eukaryotic expression system tailored for the particular protein and experimental problem; determining how to optimize expression; understanding protein tagging: the advantages and pitfalls of various affinity and solubility tags.
2) Approaches in protein purification: Choosing the best strategy for a given protein including solubilization; bulk fractionation; liquid chromatography: including conventional methods (ion exchange, size exclusion, reverse phase, etc.) and affinity methods (e.g., MAC, DNA affinity, immunoaffinity, etc.), as well as FPLC/HPLC.
3) Approaches in protein analysis: introduction to common approaches for characterization of proteins including binding assays; activity assays; mass spectroscopy to identify protein interaction partners and post-translational modifications.
In addition to purification, students will also gain exposure to fundamental analytical approaches such as mass spectroscopy and protein structure determination (e.g., X-ray crystallography, cryo-EM, etc.).