|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 4.7x 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 (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
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
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.
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
reference set with 50
reference sets are accessed using IBasis value (1 to 10) in the
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?
Q: What are RMSD and NRMSD?
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.
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.