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CIRCULAR DICHROISM SPECTROSCOPY structural analysis of nmol samples of macromolecules Prof. Eric Wickstrom.

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Presentation on theme: "CIRCULAR DICHROISM SPECTROSCOPY structural analysis of nmol samples of macromolecules Prof. Eric Wickstrom."— Presentation transcript:

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 orthogonal electronic (E) and magnetic (B) components of linearly polarized light EE BB

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,  (degree·cm 2 /dmol), and the difference in molar extinction coefficients called the molar circular dichroism, ε L -ε R =Δε (liter/mol·cm). The molar ellipticity [  ] is related to the difference in extinction coefficients by [  ] = 3298 Δε.

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)

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