Bullseye

Documentation

Google Code page with Wiki and Source for Bullseye:

https://code.google.com/p/bullseye-ms/

File formats:

Bullseye requires two input files. The first input file is the Hardklör results file which contains the peptide isotope distribution information for your MS full scan data. Hardklör can be found here.

The second input file is the file containing your MS/MS data. Acceptable file formats are .ms2, .bms2 and .cms2. Thermo .RAW files can also be used in the Windows compiled version of Bullseye.

Bullseye ouputs data in an ms2 style format or in the .mgf file format. A ms2 to dta file converter (source and linux build) is available here:
ms22dta

Bullseye usage:

Bullseye, v1.24, Nov 5, 2009
Usage: bullseye [flags] <HK file> <Data file> <Pos file> <Neg file>

HK files are Hardklör generated results files.
http://proteome.gs.washington.edu/software/hardklor

Data files contain the MS/MS data to be used.
Acceptable formats:
Thermo RAW (Windows only)
.ms2, .bms2, .cms2 (Extracted with MakeMS2)
http://proteome.gs.washington.edu/software/makems2/MakeMS2.zip

Pos and Neg files are output files for matches and non-matches, respectively.
Acceptable formats (specify with file extension):
.ms2, .bms2, .cms2, .mgf

Example: bullseye -p 5 Peptides.hk RawData.RAW Matches.ms2 NoMatch.ms2

Flags:
-c <num> Ignore peptides that persist for this length (in minutes). These peptides are considered contaminants.
Default value: 2

-e Use exact match to precursor ion. Rather than use wide precursor boundaries, this flag forces Bullseye to match precursors to the base isotope peak identified in Hardklor. The tolerance is set with the -p flag.

-m <num> Only consider peptides below this maximum mass in daltons.
Default value: 8000

-n <num> Only consider peptides above this minimum mass in daltons.
Default value: 600

-p <num> Sets the tolerance (+/- ppm) for exact match searches.
Default value: 10

-r <num> Sets the tolerance (+/- ppm) for finding persistent peptides.
Default value: 5

-t <num> Sets the tolerance (+/- minutes) around the retention time over which a peptide can be matched to the MS/MS spectrum.
Default value: 0.5

Comments

Bullseye attempts to match monoisotopic mass measurements, from identified persistent peptide isotope distributions (PPIDs), to MS/MS spectra. In complex samples, it is not unsual for multiple PPIDs to be found near an MS/MS spectrum. In those cases Bullseye will assign both mass measurements to the spectrum. In an .ms2 file format, multiple Z line entries will be made for the scan number.

It may be possible to reduce the number of scans with multiple PPIDs by adjusting Bullseye's parameters. For example, reducing the retention time tolerance ("-t") or reducing the tolerance for persistent peptides ("-r") will reduce the chances of multiple PPIDs being assigned.

Contact Ed Hsieh at edhsieh <at> u.washington.edu for any inquiries.