Biophysics and Biomolecular Interaction Centre

Determining protein dimension and oligomeric state by SEC-MALS

Determining protein activity, stability and biophysical properties as well as the interactions between proteins, nucleic acids and small molecules is at the heart of all the biochemical processes that occur in life. The Biochemistry Department houses state of the art instrumentation to analyze all these facets.

We can use the automated Multi-angle Light Scattering (MALS) in line with Size-Exclusion Chromatography (SEC) or Analytical Ultracentrifugation (AUC) to determine the molecular mass and size of macromolecular complexes. Mass spectrometry (MALDI-ToF) is also available to determine molecular weights. Instruments employing optical measurements including absorbance, fluorescence and circular dichroism (CD) can provide important insights into protein structure, stability and activity. Differential scanning calorimetry (DSC) provides detailed quantitative measure of protein stability.

Standard tools such as Isothermal Titration Calorimetry (ITC) and the newest technology of Microscale Thermophoresis (MST) gather equilibrium binding constants between molecules. In the case of ITC differences in heat signatures that are measured when two molecules interact within a confined environment. On the other hand microscale thermophoresis measures the change in diffusion rate of a molecule when it interacts with another molecule. Furthermore, the on rate and off rate of two molecules that interact can be measured by Surface Plasmon Resonance (SPR) and Biolayer Interferometry (BLI).