Sequest Parameters |
2024-05-04 |
Modifying Sequest Settings in LabKey Server
Sequest settings are based on the sequest.params file (See your Sequest documentation). For many applications, the Sequest default settings used by LabKey Server are likely to be adequate, so you may not need to change them. If you do wish to override some of the default settings, you can do so in one of two ways:
Sequest takes parameters specified in XML format. In LabKey Server, the default parameters are defined in a file named sequest_default.input.xml, at the pipeline root. When you create a new search protocol for a given data file or set of files, you can override the default parameters. Each search protocol has a corresponding Sequest Sequest analysis definition file, and any parameters that you override are stored in the file, named sequest.xml by default.
Note: If you are modifying a sequest.xml file by hand, you don't need to copy parameter values from the sequest_default_input.xml file. The parameters definitions in these files are merged by LabKey Server at runtime.
Using X!Tandem Syntax for Sequest Parameters
You don't have to be knowledgeable about XML to modify Sequest parameters in LabKey Server. You only need to find the parameter that you need to change, determine the value want to set it to, and paste the correct line into the Sequest XML section when you create your MS2 search protocol.
When possible the Sequest parameters will use the same tags already defined for X!Tandem. Most X!Tandem tags are defined in the X!Tandem documentation, available here:
http://www.thegpm.org/TANDEM/api/index.html
As you'll see in the X!Tandem documentation, the general format for a parameter is as follows:
<note type="input" label="GROUP, NAME">VALUE</note>
For example, in the following entry, the parameter group is residue, and the parameter name is modification mass. The value given for the modification mass is 227.2 daltons at cysteine residues.
<note type="residue, modification mass">227.2@C</note>
LabKey Server provides additional parameters for Sequest where X!Tandem does not have an equivlent parameter, for working with the data pipeline and for performing quantitation, described in the following sections.
The Sequest parameters the you see in a standard sequest.params file are defined here:
sequest.params name | GROUP | NAME | Default | Notes |
first_database_name | pipeline | database | n.a. | Entered through the search form. |
peptide_mass_tolerance | spectrum |
parent monoisotopic mass error plus parent monoisotopic mass error minus |
2.0f | They must be set to the same value |
use an indexed ("pre-digested") fasta file | pipeline |
use_index |
0 (no) | If set, the SEQUEST pipeline will use a fasta file index to perform the search. If the index does not already exist, the pipeline will invoke makedb.exe to create the index. 1 means yes |
name of indexed fasta file | pipeline |
index_name |
empty | (Optional). Specifies the name of the index to generate and use. If no name is specified, the SEQUEST pipeline will create a name based on the values of the search parameters, in particular the enzyme_info. |
peptide_mass_units | spectrum | parent monoisotopic mass error units | Daltons | The value for this parameter may be 'Daltons' or 'ppm': all other values are ignored |
ion_series | scoring |
a ions |
no yes no no yes no |
On is 1 and off is 0. No fractional values. |
sequest |
d ions |
no |
||
fragment_ion_tolerance | spectrum | fragment mass error | 1.0 | |
num_output_lines | sequest | num_output_lines | 10 | |
num_results | sequest | num_results | 500 | |
num_description_lines | sequest | num_description_lines | 5 | |
show_fragment_ions | sequest | show_fragment_ions | 0 | |
print_duplicate_references | sequest | print_duplicate_references | 40 | |
enzyme_info | protein | cleavage site |
[RK]|{P} |
|
max_num_differential_AA_per_mod |
sequest | max_num_differential_AA_per_mod | 3 | |
max_num_differential_per_peptide | sequest | max_num_differential_per_peptide | 3 | |
diff_search_options | residue | potential modification mass | none | |
term_diff_search_options | refine |
potential N-terminus modifications potential C-terminus modifications |
none | |
nucleotide_reading_frame | n.a | n.a | 0 | Not settable. |
mass_type_parent | sequest | mass_type_parent | 0 | 0=average masses 1=monoisotopic masses |
mass_type_fragment | spectrum | fragment mass type | 1 | 0=average masses 1=monoisotopic masses |
normalize_xcorr | sequest | normalize_xcorr | 0 | |
remove_precursor_peak |
sequest | remove_precursor_peak | 0 | 0=no 1=yes |
ion_cutoff_percentage | sequest | ion_cutoff_percentage | 0 | |
max_num_internal_cleavage_sites | scoring | maximum missed cleavage sites | 2 | |
protein_mass_filter | n.a. | n.a. | 0 0 | Not settable. |
match_peak_count | sequest | match_peak_count | 0 | |
match_peak_allowed_error | sequest | match_peak_allowed_error | 1 | |
match_peak_tolerance | sequest | match_peak_tolerance | 1 | |
create_output_files | n.a. | n.a. | 1 |
Not settable. |
partial_sequence | n.a. | n.a. | none | Not settable. |
sequence_header_filter | n.a. | n.a. | none | Not settable. |
add_Cterm_peptide | protein | cleavage C-terminal mass change | 0 | |
add_Cterm_protein | protein | C-terminal residue modification mass | 0 | |
add_Nterm_peptide | protein | cleavage N-terminal mass change | 0 | |
add_Nterm_protein | protein | protein, N-terminal residue modification mass | 0 | |
add_G_Glycine |
residue | modification mass | 0 |