CD  (Tutorials:   Szilagyi  /  Rupp)  

Circular Dichroism is a useful tool for monitoring protein and nucleic acid structures and tracking conformational changes.  This information can be used to obtain information about secondary structure, folding and other conformational changes.  CD sprectrum of representative protein structures.  

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Review:  Properties of polarized light

Linearly Polarized Light – direction of electric vector is constant but magnitude

          is modulated.      Andras Szilagyi demos: linear  combo

Circularly Polarized Light – magnitude is constant, but direction is modulated.

          Circularly polarized light may be either “right-handed” or “left-handed.”  

                        Andras Szilagyi demos: +90combo   +90/-90both  R/Lcombo

Birefringence - "birefringence" means double refraction - it refers to those materials 

         whose refractive index depends on the direction of the light.    

Dichroism – phenomenon in which light absorption differs for different directions of polarization.

                      Andras Szilagyi demos:    Lin_refraction     Cir_refraction

     .

Linear and Circular Dichroism are special kinds of absorption spectroscopy and

          occur at energies where absorption takes place.

                      Andras Szilagyi demos:   CD   

 LD (Linear Dichroism) -       LD (l) = A_J (l)  - A_^ (l) 

           Linearly polarized light oriented along the transition dipole will be absorbed

           strongly, light oriented perpendicular to dipole will not be absorbed.

 Compare absorption parallel vs. perpendicular to helix axis.

Circular Birefringence - occurs when the refractive index is different for left and right circularly

        polarized light.  Circular birefringence rotates the plane of polarization of plane-polarized

        light (Optical Rotation).   

Andras Szilagyi demos:  CD_birefring1   CD_birefrin2

ORD (Optical Rotary Dispersion) – optical activity as seen in rotation of linearly

          polarized light due to difference in refractive index of two types of

circularly polarized light.  (n_L ¹ n_R) ORD vs. l

 CD (Circular Dichroism) – difference in absorption or left and right circularly

          polarized light.   (e_L  ¹  e_R)  Very small effects, far UV (170 – 300 nm)   Tutorial

      D (l) = A_L(l)  - A_R(l)  =  [e_L(l)  -  e_R(l)] lc  =  De lc  

    Historically – CD results reported as “ellipticity”   q =   DA (32.98)

          or  molar ellipticity  [q]  =  3298 De   (deg · dl/mol · dm).

For proteins, CD_{207 nm} ~ % helix  = q /-57,000.

 Note:  CD and ORD can be interconverted using the Kronig-Kramer relationships.  Most experiments today are reported as CD results.

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   Figures / Illustrations of Data:

   1. A) CD and ORD poly-L-Lys in a-helical, b-sheet, and random coil.

        B) Experimentally observed CD for helical protein myoglobin.

   2. Expanded wavelength range for CD of “pure” secondary structures.

   3. CD sprectrum of representative protein structures.

   4. Use of CD spectra to estimate secondary structure of proteins.

                   Fraction helix content  =   (theta)q₂₀₇  / -57000 

                   Better way: Singular Value Decomposition Analysis

                   (Find the best matrix that can be used to generate the best fit for

                                 converting CD sprectra into secondary structure.)

    5. Use of CD to monitor folding or denaturation of proteins

    6. CD sprecta of DNA to monitor folding or denaturation of DNA

       CD spectra of DNA to estimate secondary structure of nucleic acids