Interactions among proteins are essential for any living entity. They are being studied for decades through various approaches. Over the years, many different techniques and instruments were employed in order to describe individual binding parameters, e.g. stoichiometry, equilibrium binding constants, kinetic constants, or enthalpy and entropy of binding. In order to use these techniques efficiently, it is crucial to understand their principles as well as be able to compare their outcomes, advantages and limitations. For this, the basic university lectures are frequently insufficient since they might lack a more practical approach. Here, various forms of more targeted education (workshops, specialized courses, etc.) based on hands-on exercises play their irreplaceable role.
In order to highlight differences and similarities of the techniques, it is highly desirable to use the same interacting system for all experiments. While there are various systems suggested for individual types of analysis (e.g. Ca2+-EDTA for isothermal titration calorimetry [1] or lysozyme-NAG3 for MST [2]), a more universal interacting couple is more rare to be found. In this talk, we would like to demonstrate an application of bovine pancreas α-chymotrypsin and soybean trypsin inhibitor in a specialized university course. Both proteins are easy to obtain from natural sources and therefore commercially available. They interact with Kd ~ 1 μM, being suitable system for analysis through various techniques including isothermal titration calorimetry, microscale thermophoresis, analytical ultracentrifugation [3] or surface-based techniques. The system can be therefore used for multiple methods in parallel, making it a useful tool for their comparison, highlighting the advantages and disadvantages of each approach. It might be also used for other purposes, such as routine instrument check or cross-instrument or laboratory benchmarking.
Measurements at CF Biomolecular Interactions and Crystallography of CIISB, Instruct-CZ Centre are supported by MEYS CR (LM2023042) and European Regional Development Fund-Project „UP CIISB“ (No. CZ.02.1.01/0.0/0.0/18_046/0015974).