Fixes for known SEQUEST problems
Yet Another SEQUEST tip!
5/3/2001 Bill Mahn of ThermoFinnigan recently sent me a list of fixes for known SEQUEST/TurboSEQUEST problems as of April 2001. These issues are relevant for SEQUEST/TurboSEQUEST 2.0 on Xcalibur 1.2. Please contact Bill Mahn at techsupport@thermofinnigan.com if you wish to acquire the software patches.
- We detailed how to fix the SEQUEST Browser Run Summary buttons a while back. This is an easy fix that you can perform yourself. The information is available on our website here.
- There is a problem with applying static modifications to proteins when performing SEQUEST searches. This problem may prevent you from identifying peptides that have the static modifications. The fix requires a new version of the sequest.params file and an updated version of the biobrowser.dll. Contact ThermoFinnigan to obtain these software patches.
- The TIC threshold applied during DTA file creation on SEQUEST searches has been known to fail occasionally and can be fixed with a new version of the lcq_dta.exe file. Contact ThermoFinnigan to obtain this software patch.
- There was a problem with the Database Manager utility which prevented changing the drive for indexing databases without having to re-install TurboSEQUEST. This fix requires a new version of DatabaseManager.exe. Contact ThermoFinnigan to obtain this software patch.
5/17/2001 We have recently discovered a few errors in the SEQUEST BROWSER code (Perl scripts) which are detailed below. We have fixed these errors and provided the fixes to ThermoFinnigan. Unfortunately, we are not authorized to distribute these as the software is under copyright protection. Contact ThermoFinnigan if you think these may have an impact on your applications. These are relevant to Xcalibur 1.2 running Bioworks 2.0.
- SEQUEST BROWSER does not display modifications to termini (N or C) properly upon any display ions request from either the SEQUEST SUMMARY page or the SHOWOUT pages. The SEQUEST searches are performed correctly, but the ion display does not reflect the modification properly. For example, in the case of an N-terminal modification, the B ions are not displayed correctly. The runsummary.pl and showout.pl Perl scripts were modified to fix this problem.
- SEQUEST BROWSER does not handle the 3rd amino acid differential modification properly, even though one can input up to 3 differential modifications on the form on the Run SEQUEST page. Errors were found in the sequest_launcher.pl, runsummary.pl, and showout.pl Perl scripts. The error in sequest_launcher.pl prevented the 3rd AA modification from being submitted to SEQUEST for searching. Problems with runsummary.pl, and showout.pl affected the proper displaying of sequence ions with these modifications.
- The displayions.exe cannot handle more than 2 differential modifications. The displayions.exe program is used by the SEQUEST BROWSER to generate the MS/MS spectrum display with labeled sequence ions. Qual Browser uses web_display.exe for this function. We have changed the relevant Perl scripts to call web_diplay.exe instead of displayions.exe if more that 2 modifications are specified.
5/17/01 Other miscellaneous known issues that we are aware of:
- A SEQUEST search run on a peptide with a differential modification which happens to reside on the N-terminal amino acid does not return the proper result if an enzyme is defined. An example of this would be pyro-Glu (DM = -17 on Q) on the N-terminus of an Ig chain. If the modification on the N-terminal peptide is on an internal amino acid (e.g., an internal Cys), it seems to work ok. The search works properly if no enzyme is defined. This appears to be a SEQUEST bug and would be apparent if SEQUEST were run from either Qual Browser or the SEQUEST BROWSER. The only work around right now is to run the search without an enzyme defined, or use a static modification.
- SEQUEST run from Qual Browser may produce an inaccurate MS/MS ions display if the differential modification is negative (e.g., -17 as with pyro-Glu discussed above). The only work around is to manually correct the URL link to correctly display the MS/MS ions in the browser. For example, a bad link will look something like this (for a search where C had a diff mod of +57 and Q had a diff mod of -17):
http://mymachine/cgi-shl/web_display.exe?Dta=
C:\Xcalibur\results/datafile.0268.0268.2.dta&MassType=1&NumAxis=1&DMass1=
57.00&DSite=0000000000000&Pep=Q#VQLVQSGSELK
While a good link looks something like this for a Q modified by -17 on the N-terminus. The changes we've made to the link are highlighted in red text.
http://mymachine/cgi-shl/web_display.exe?Dta=
C:\Xcalibur\results/datafile.0268.0268.2.dta&MassType=1&NumAxis=1&DMass1=
57.00&DMass2=-17;DSite=2000000000000&Pep=QVQLVQSGSELK
©Copyright 2000-2008 Novatia LLC
Disclaimer and terms of usage
e-mail: info@enovatia.com
|
