Help

Input

File name and File type

OMSSA can take one of the following input formats:

• a single dta file
• a set of dta files merged into a single file, separated by blank lines.
• a set of dta files merged into a single file, separated by xml-like tags that allow tracking of spectra by file name and by number.  A Perl script to do the merge is included in the distribution.  It is called dta_merge_OMSSA.pl and is described in the README file.
• an mgf file
• a pkl file

Sequence Library

Sequence library to search.  Refseq is comprehensive, consistent, and stable library of sequences meant as a reference standard. Nr is a non-redundant set of most protein sequences in NCBI databases.
 

Enzyme and Maximum missed cleavages

The enzyme used to theoretically cleave the sequence library. If you cannot find the enzyme you want to use in the list, please contact Lewis Geer.

Species to search

Limits your search to a particular organism or organisms. Multiple organisms can be selected by holding down the "Ctrl" key while clicking on the organism names.  Public search service users must select the organisms to search. If you want to search all organisms at once, you must download OMSSA and install it on your own computer.

If the organism you wish to search is not listed, please contact Lewis Geer.

Hitlist max length and E-value cutoff

Both options affect the number of hits kept in memory during the search and returned to the user. The hitlist max length is the maximum number of hits retained per spectrum per precursor charge. The E-value cutoff is the maximum E-value allowed in the hit list.

Fixed and Variable modifications

Enter post translational modifications to be searched. Fixed modifications are modifications that always occur. Variable modifications are modifications that may occur -- OMSSA will search both the modified and unmodified versions of peptides. Multiple modifications can be selected by holding down the "Ctrl" key and clicking.

If a modification you wish to search is not listed, please contact Lewis Geer.

Max mod combinations per peptide

To reduce the combinatorial expansion that results when specifying multiple variable modifications, it is possible to put an upper bound on the number of mass ladders generated per peptide using this option.  The ladders are generated in the order of the least number of modifications to the most number of modifications so that modifications are applied sparingly at first. If you set this number too low, you will miss highly modified peptides. If you set it too high, it will make the e-values less significant.

To give an example what this means, assume that the hard limit is 11 and that the theoretical peptide is STYY and you've selected phosphorylation of S, T, and Y as variable mods. The combinations that OMSSA will test are:

STYY
----
0000
1000
0100
0010
0001
1100
1010
1001
0110
0101
0011

where 0 represents no modification and 1 represents a modification at the site indicated by the column the digit is in. OMSSA tries the combinations with the least number of variable modifications and then adds modifications until the upper bound is reached.

Precursor and Product Mass Tolerance

The precursor ion is the ion before fragmentation and the product ions are the ions generated after fragmentation. The mass tolerance is the instrumental error in determining the mass of these ions. These values are specified in Daltons +/- the measured value, e.g. a value of 2.0 means +/- 2.0 Daltons of the measured value.

Scaling of precursor mass tolerance

Allows you to specify how the mass tolerance scales with the charge of the precursor. For example, you may search a precursor assuming that it has a charge state of 2+ and 3+. If you set the parameter to 1, then the mass tolerance for the 2+ charge state will be 2 times the precursor mass tolerance, and for the 3+ charge state it will be 3 times the precursor mass tolerance. If you set the parameter to 0, the mass tolerance will always be equal to the precursor mass tolerance, irrespective of charge state.

Precursor and Product Search type

These settings specify the use of average or monoisotopic amino acid masses when calculating theoretical precursor and product ion masses, respectively.

Charge Handling

Determination of precursor charge and product ion charges.  Presently, OMSSA estimates which precursors are 1+ using the fraction of peaks below the precursor as an indicator. If the number of peaks is below the specified fraction, then the spectrum is searched as 1+.   All other spectra are searched with precursor charge ranging from the minimum to maximum specified.

OMSSA ignores charges specified in the input file, except when using it to calculate the precursor m/z. For example, if you are searching using a dta file, OMSSA will search over the minimum and maximum precursor charge you specified, not the charge specified in the dta file. In the case of dta files, this means that you do not have to search multiple versions of the same file, where each is calculated using a different precursor charge.

The "Charge at which to start considering multiply charged products" setting is the lowest precursor charge at which OMSSA should start considering multiply charge product ions.

Peak intensity cutoff and Number of top intensity peaks

Preprocessing is the process of eliminating noise from a spectrum. Normally, you do not need to adjust options associated with preprocessing as OMSSA automatically adjusts its preprocessing for best results.

The "Peak intensity cutoff" eliminates all peaks whose intensity is less than a fraction of the most intense peak.

The "Number of top intensity peaks in first pass" is the number of top intensity peaks from which OMSSA must find at least one match between the a peak m/z value and the m/z values calculated from the protein sequence library peptide.

Ions to search

OMSSA searches two ions series, both of which can be specified.  Normally one of the specified ion series is a forward ion series and the other is a reverse ion series.

Output

OMSSA can create output in CSV (excel), XML and ASN.1 using the form on the results page. The CSV format is a convenient summary of the detailed results in the XML and ASN.1 format files. The XML and ASN.1 format files are logically equivalent. 

A sample Perl parser for the xml output is included (see the README file).  To use the sample parser, type "perl readOMSSA.pl test.xml" where test.xml is a file containing xml output from OMSSA.