CIRCULAR DICHROISM SPECTROSCOPY

1
CIRCULAR DICHROISM SPECTROSCOPY

• structural analysis of nmol samples of
  macromolecules
• Prof. Eric Wickstrom

2
Circular Dichroism is the difference in
absorption between left and right hand circularly
polarised light in chiral molecules. A chiral
molecule is one with a low degree of symmetry
which can exist in two mirror image isomers.
Illustrated above is an example of circular
dichroism in glucose, a simple sugar.
3
? E
? B
orthogonal electronic (E) and magnetic (B)
components of linearly polarized light
4
electronic field of linearly polarized light
(left) right-handed circularly polarized light
(right)
5
Ellipticity, ?
6
? ? Prism Polarizer
7
The most commonly used units are mean residue
ellipticity, ? (degreecm2/dmol), and the
difference in molar extinction coefficients
called the molar circular dichroism, eL-eR ?e
(liter/molcm). The molar ellipticity ? is
related to the difference in extinction
coefficients by ? 3298 ?e.
8
Typical Initial Concentrations

• Protein Concentration 0.5 mg/ml
• Cell Path Length 0.5 mm
• Stabilizers (Metal ions, etc.) minimum
• Buffer Concentration 5 mM or as low as possible
  while maintaining protein stability
• Contaminants Unfolded protein, peptides,
  particulate matter (scattering particles)

9
(No Transcript)