Biomolecular Interactions

The binding of proteins into reversible complexes underlies much of biology as we know it, whether for purposes of signaling, formation of cellular structures, cellular division, immune response or other processes crucial to maintaining a healthy, functioning organism. Many of these protein-protein interactions are difficult to analyze by traditional means such as cellular assays, gel-shift assays, or even biophysical techniques such as surface plasmon resonance or isothermal titration calorimetry, as a result of complex phenomena such as multi-valence, co-operativity, multi-protein assembly, or combined self- and hetero-association.

One of the most versatile techniques for characterizing protein and other biomolecular interactions in solution, without labeling or immobilization, is composition-gradient multi-angle light scattering (CG-MALS). In this technique, the Calypso II automatically prepares a series of compositions or concentrations and delivers them as stop-flow injections to a DAWN or miniDAWN MALS detector. The CALYPSO software uses these MALS measurements in order to determine the weight-averaged molar mass of the solution as a function of composition, then assess the fit of the data to an association scheme in order to decide if the data support the specific model and quantify the binding constants (Kd). The software provides an essentially infinite variety of association models consisting of one or two interacting species, permitting the analysis of self-association, hetero-association, combined self- and hetero-association, multi-valent interactions and co-operative binding.

The real power of CG-MALS derives from its direct assessment of molar mass, which removes all ambiguity from the determination of oligomeric state and stoichiometry of multi-protein complexes. Method development is simple and straightforward, assisted by the simulation capabilities of the CALYPSO software.