2. Using Structure to Elucidate Function/Mechanism

3. An experimental strategy for – determining the arrangement of atoms/molecules in space to obtain structural information

4. –understand a mechanism in detail –structure-based (rational) drug design –elucidate a novel structure –understand a conformational switch (e.g. prions) Some situations in which this strategy might be applied

5. X ray bombardment of crystals circular dichroism (CD) nuclear magnetic resonance (NMR)- size < 25 kD sequence comparison/analysis (e.g. COILS or threading)- surface residues are most conserved electron microscopy (EM) atomic force microscopy photo/chemical cross-linking protease digestion calorimetry sedimentation Some methods that provide structural information:

6. The differential absorption of radiation polarized in two directions as a function of frequency is called dichroism. Circular dichroism (CD) spectroscopy is a form of light absorption spectroscopy that measures the difference in absorbance of right- and left-circularly polarized light (rather than the commonly used absorbance of isotropic light) by a substance. Circular Dichroism Kurt D. Berndt http://broccoli.mfn.ki.se/pps_course_96/ss_960723_21.html

7. Circular Dichroism Kurt D. Berndt http://broccoli.mfn.ki.se/pps_course_96/ss_960723_21.html Circular dichroism is very sensitive to the secondary structure of polypeptides and proteins.

9. –material for study should be of homogeneous composition. How? –often useful to reduce the complexity of the problem by focusing on a subregion of a larger molecule. How? –minimize flexibility of the molecule in solution so that only a small number of conformations are possible. How? What are the experimental prerequisites for structural studies?

10. –biologically irrelevant structure? Less common than you might think. –faulty interpretation. Interfaces that form during packing of molecules into a crystal may not be biologically relevant. Solution? What are possible pitfalls of crystallographic studies?