BIOPHYSICS CENTRE FOR STRUCTURE AND INTERACTIONS

             The centre comprises a wide range of analytical methods such as absorbance spectroscopy, spectrofluorimetry, circular dichroism spectrometry, diferential scanning fluorimetry, microcalorimetry and surface plasmon resonance. Proteins, as well as a wide variety of other biomolecules (such as nucleic acids and small molecules), can be studied in terms of their structure, stability, function and molecular interactions.

Structural and functional analysis of biomolecules and their stability;

• Structure
  • Tertiary structure by circular dichroism - CD
  • Secondary structure by CD
  • CD in aquose (soluble) and lipid (membrane) phases
  • Conformational changes monitored by CD
  • Folding state (e.g. of lack of function mutants)
  • Proteins, Nucleic acids, complexes, etc
  • X-ray crystallography (in house)
  • Size and shape:
    • SEC plus Ultracentrifugation a la Siegel and Monte
• Stability
  • Diferencial Scanning Fluorimetry (DSF, “Thermal shift”)
  • Circular dichroism - CD
  • Diferential Scanning Calorimetry – DSC (@ FCUP)
• Function
  • UV-Vis spectrophotometry based activity assays
  • Fluorimetry based activity assays
  • Interaction analysis (see below)

Characterization of molecular interactions

• Technologies:
  • Label free:
    • Surface Plasmon Ressonance (SPR) (aka Biacore)
    • Isothermal titration calorimetry (ITC)
    • Circular dichroism (CD)
  • With probes:
    • Fluorescence Anisotropy Measurements
    • Differential Scanning Fluorimetry (DSF)
  • Kinetics and Thermodinamic characterization of binding events
    • How fast, how slow is binding and unbinding
    • Energetics of binding
    • How specific (drug targetable) are the interactions
    • stoichiometry

• Some Applications:
  • characterization of molecular interactions of proteins, antibodies, nucleic acids, lipids, cells and other biomolecules;
  • analysis of interactions between drug compounds and target molecules,
  • active concentration - concentration based on specific binding
  • analysis of interactions of biomolecules in membranes (liposomes, reconstituted bilayers and monolayres)
  • assessment of the effect of structural changes on binding mechanisms

Please check the available resources here.