Q: How do I use CDPro?
A: CDPro can be used either in a PC or on any computer. The executables and the browser-based linker for analysis are provided at this site; download CDPro.zip. The programs need to be compiled for any other computer. The INPUT file need to be generated before executing the CDPro programs (Use CRDATA program and a set of wavelength-CD values). For a unix machine CDSSTR analysis program is differrent.

Q: What spectral range should be used for a good analysis?
A: Normally, one should use CD spectra at least in the range 190-240 nm. But sometimes experimental conditions limit the range to 200 nm or so, and the flexibility of the refernce sets in the more recent programs allows the use of 200-240 nm in CD analyses. The reliability, however, is slightly reduced as one reduces the wavelength range. Comparison of performances of the two spectral ranges are available in Sreerama and Woody. Anal. Biochem., 287, 252, (2000)

Q: What are units of CD data used in the analysis?
A: The CD spectra should be in per residue molar absorbance units (De M^-1 cm^-1). Most CD instruments have the software to convert raw CD data to either molar ellipticity or molar absorbance units. Be sure to divide the CD data by the number of residues in the protein. The CD data in either molar ellipticity or molar absorbance units can be used in the CRDATA program to generate the INPUT file.

Q: How do I generate the Input file?
A: The analysis programs are written is FORTRAN and they require CD data in a certain format. The file "INPUT" need to be created containing the CD data, wavelength range, etc. It can be done using a program called "CRDATA" which converts CD data from any CD instrument in a text file to the required format. You will have to refer to the manual or check the software that came with your CD instrument. The text file should contain only wavelength and CD value in each line. Check the examples provided.

Q: How do I choose a reference set?
A: The reference set with the largest number of proteins should normally be chosen. This often depends on your CD data. If your CD data is only down to 190 nm or so (say, 190-250 nm) then you can select the reference set with 43 proteins (IBasis =  4). But if you have CD data down to 180 nm (180-250 nm) then it would be wise to use the reference set with 29 proteins, which has CD data in the range 178-260 nm (one can also use 43-protein reference set and compare results with 29-protein set in this special situation). Our studies have suggested that the limitaions introduced in the analysis by the lack of short-wavelength data (180 to 190 nm) is partially overcome by the use of a larger reference set. 
       Special reference set are also available, developed for specific applications. If you are studying denaturation(e.g., thermal denaturation) then you shoud use a reference set that includes denatured proteins (SDP42 or SDP48). If you are studying membrane proteins then you should use a reference set that includes membrane proteins (SMP50 or SMP56).
       For a set of CD spectra monitoring denaturation use SDP42 or SDP48. You may get different results from reference sets of different quality (e.g, SP with SDP, SDP with SMP); Do not mix them.

Q: What is a reference set? What reference sets are available?
A: A reference set is a set of proteins whose CD spectra and Secondary Structures are used in the analysis. It is also referred to as Basis set, Database, etc. CDPro has three classes of reference sets, developed from  (1) only Soluble Proteins, (2) Soluble and Denatured Proteins, and (3)  Soluble and Membrane Proteins. They are referenced as SPxx, SDPxx, SMPxx, where xx denotes the number of proteins in the set. (e.g., SMP50 corresponds to Soluble and MembraneProtein reference set with 50 proteins). The reference sets are accessed using IBasis value (1 to 10)  in the INPUT file.

Q: Which values can I change, and How do I change them, in the INPUT file? 
A: The only values that may be changed in a properly generated INPUT file are IPRINT (from 0 to 1) and IBasis (any value between 1-10). Changing IPRINT to 1 would generate the output for every iteration and this should be done if a problem is suspected in the analysis; otherwise the standard output is sufficient to examine the results. Basis_1 value determines the reference set to be used in the analysis. Normally, the largest reference set available for the wavelength range should be used. Value of 8 selects a reference set based on the tertiary class determined by the CLUSTER program which should be run prior to its selection. Values 6 and 7 select reference sets with denatured CD spectra, and should be used for studying CD spectra under denaturing conditions. Consult the Readme and other help files provided. The changes in the INPUT file can be made using a text editor; Edit_Input button at the top of the screen can be used to open the DOS editor on your PC.

Q: How do I read the results?
A: Each program (CDSSTR, CONTINLL, and SELCON3) has a separate output (program_name.OUT) which contains results at various stages. Summary of the run (title, reference set, program and secondary structure) is added to ProtSS.OUT (execution of each of these programs write to this file). The calculated CD spectrum (corresponding to the result) is also output -- see next Q.

Q: Can I compare the Calculated CD spectra with Experiment?
A: Yes. The calculated CD spectrum (from each program) is compared with the experimental spectrum in files: CalcCD.OUT, reconCD.OUT and CONTIN.CD. They can be imported into a plotting software, such as Axum or SigmaPlot . The programs also calculate the RMSD between the two and print the result in program_name.out and ProtSS.out files.