Structure based drug design

Question 1

X-Ray Crystallography

Answer 1

• most commonly used technique in obtaining structural information
• requires large amount of pure protein which must be crystallised (difficult process)
• typically carried out by a vapor diffusion method where the solution is equilibrated with reagents that promote crystallisation (i.e. salts, PEG) under various conditions in a 96 well plate
• Crystallised product undergoes NMR analysis, producing a diffraction pattern which can be mathematically to give an electron density map which provides a structure
• 1-2 A resolution, cant see H atoms

Question 2

XRC; Synchrotron radiation

Answer 2

• accelerated electrons from the synchotron mean X-rays are very much brighter than those generated from those in labratories, meaning faster and higher resolution in results.
• electrons travel in a vacuum ring where the path of the electrons is directed into a circle by bending magnets
• as directions of the electrons change it emits high energy photos (x rays)

Question 3

Cyro-electron microscopy:

Answer 3

• A beam of electron is fired at a frozen sample of protein solution. Emerging scattered electrons pass through a lens to create a magnified image of the detector, from which their structure can be worked out
• Advantageous in that the protein does not need to be crystallised, however, images are of lower resolution.

Question 4

Docking

Answer 4

• In silico procedure used to determine how a molecule will fit to a protein via the use of molecular mechanics
• Each bound conformation is known as a pose
• Score is an estimate of binding affinity
• works well when the binding site is well defined and relatively small
• does not treat a protein as being flexible
• used to select compounds from a database for screening