Q: What is CDPro?
A: CDPro is a suite of programs for
protein CD analysis. It contains three popular CD analysis programs [CONTIN,
CDSSTR, and SELCON3], a program to generate INPUT file that
is used by all three programs, and a program [CLUSTER] for
determining the tertiary structure class from CD spectrum. The CD
analysis programs use multiple sets of reference proteins [CD and
secondary structure data] and these are also provided in this site. Download
the CDPro.zip file and unzip it from your C: drive. CDPro directory
will be created with all necessary files. Open the RunCDPro.html file from a internet browser.
(Netscape 4.7 or IE6.0).
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.
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 et al. Anal. Biochem. (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 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 (Basis_1 = 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. 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.
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 Basis_1 (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.
Q: What are RMSD and
NRMSD?
A: These are the RMS and normalized RMS
differences between the calculated and experimental CD spectra. They
can be different from different reference sets and different programs.
For a given INPUT file (given CD data and a chosen reference set) both
RMSD and NRMSD decrease in the order: SELCON3 > CDSSTR > CONTINLL.
This difference has to do with the algorithm and these values should not be used
to judge the quality of the analysis. |