Module 2 : Spectroscopic Methods

Lecture 10 : Circular dichroism of Proteins – II

  1. Question/Instruction 8: Hit ENTER to continue
      Ctrl-C to ABORT

a. If all the details you have entered are correct, press ENTER. The basis sets available for analysis are displayed.

  1. Question/Instruction 9: Select a IBasis value
    1. Select the reference set with maximum number of proteins but encompassing the smallest wavelength the CD data is recorded at.

    2. The software suggests an option which basis set should be used. It is recommended to use the suggested option.

    3. Type the suggested IBasis number and press ENTER.
  2. The above steps generate a file, INPUT.SMP; this file is used as the input file for CD spectrum deconvolution. Important : If there is an INPUT.SMP file already present in the CDPro directory, it gets overwritten when steps 12 – 19 are followed.

  3. Execute the CDSSTR.exe application by double clicking it. The application calculates the fractions of the secondary structure components using the INPUT file.

  4. Press ENTER.

  5. Execute the CONTINLL.exe application by double clicking it. The application calculates the fractions of the secondary structure components using the INPUT file.

  6. Press ENTER.

  7. Execute the SELCON3.exe application by double clicking it. The application calculates the fractions of the secondary structure components using the INPUT file.

  8. Press ENTER.

  9. Open the CDSSTR.out, CONTIN.out, and SELCON3.out to see the output of CDSSTR, CONTINLL, and SELCON3 algorithms, respectively.

  10. Open ProtSS.out file using Notepad. The file displays the CD deconvolution results obtained from all the three algorithms.

  11. Choose the appropriate results based on the RMSD (root mean squared deviation) and NRMSD (normalized root mean square deviation).

Notes:

  1. It is a common mistake not to include ‘.txt' after the file name. Make sure the file extension is included.

References:

  1. Sreerema, N. and Woody, R.W. (1993) A self-consistent method for the analysis of protein secondary structure from circular dichroism. Analytical Biochemistry, 209, 32-44.

  2. Sreerama, N. and Woody, R. W. (2000) Estimation of Protein Secondary Structure from Circular Dichroism Spectra: Comparison of CONTIN, SELCON, and CDSSTR Methods with an Expanded Reference Set . Analytical Biochemistry, 287, 252-260.

  3. Whitmore, L. and Wallace, B. A. (2004) DICHROWEB: an online server for protein secondary structure analyses from circular dichroism spectroscopic data. Nucleic Acids Research, 32, W668-W673.


    Greenfield, N. J. (2006) Using circular dichroism spectra to estimate protein secondary structure. Nature Protocols, 1, 2876-2890.