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:

  • You can modify the default Sequest parameters for the pipeline, which will set the defaults for every search protocol defined for data files in the pipeline (see Set the LabKey Pipeline Root).
  • You can override the default Sequest parameters for an individual search protocol (see Search and Process MS/MS Data).

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 nameGROUPNAMEDefaultNotes
first_database_namepipelinedatabasen.a.Entered through the search form.
peptide_mass_tolerancespectrum

parent monoisotopic mass error plus

parent monoisotopic mass error minus

2.0fThey must be set to the same value
use an indexed ("pre-digested") fasta filepipeline

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 filepipeline

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_unitsspectrumparent monoisotopic mass error unitsDaltonsThe value for this parameter may be 'Daltons' or 'ppm': all other values are ignored
ion_seriesscoring

a ions
b ions
c ions
x ions
y ions
z ions

no
yes
no
no
yes
no

On is 1 and off is 0. No fractional values.

sequest

d ions
v ions
w ions
a neutral loss
b neutral loss
y neutral loss

no
no
no
no
yes
yes

fragment_ion_tolerancespectrumfragment mass error1.0 
num_output_linessequestnum_output_lines10 
num_resultssequestnum_results500 
num_description_linessequestnum_description_lines5 
show_fragment_ionssequestshow_fragment_ions0 
print_duplicate_referencessequestprint_duplicate_references40 
enzyme_infoproteincleavage site

[RK]|{P}

 

max_num_differential_AA_per_mod

sequestmax_num_differential_AA_per_mod3 
max_num_differential_per_peptidesequestmax_num_differential_per_peptide3 
diff_search_optionsresiduepotential modification massnone 
term_diff_search_optionsrefine

potential N-terminus modifications

potential C-terminus modifications

none 
nucleotide_reading_framen.an.a0Not settable.
mass_type_parentsequestmass_type_parent00=average masses 1=monoisotopic masses
mass_type_fragmentspectrumfragment mass type10=average masses
1=monoisotopic masses
normalize_xcorrsequestnormalize_xcorr0 

remove_precursor_peak

sequestremove_precursor_peak00=no
1=yes
ion_cutoff_percentagesequestion_cutoff_percentage0 
max_num_internal_cleavage_sitesscoringmaximum missed cleavage sites2 
protein_mass_filtern.a.n.a.0 0Not settable.
match_peak_countsequestmatch_peak_count0 
match_peak_allowed_errorsequestmatch_peak_allowed_error1 
match_peak_tolerancesequestmatch_peak_tolerance1 
create_output_filesn.a.n.a.1

Not settable.

partial_sequencen.a.n.a.noneNot settable.
sequence_header_filtern.a.n.a.noneNot settable.
add_Cterm_peptideproteincleavage C-terminal mass change0 
add_Cterm_proteinproteinC-terminal residue modification mass0 
add_Nterm_peptideproteincleavage N-terminal mass change0 
add_Nterm_proteinproteinprotein, N-terminal residue modification mass0 

add_G_Glycine
add_A_Alanine
add_S_Serine
add_P_Proline
add_V_Valine
add_T_Threonine
add_C_Cysteine
add_L_Leucine
add_I_Isoleucine
add_X_LorI
add_N_Asparagine
add_O_Ornithine
add_B_avg_NandD
add_D_Aspartic_Acid
add_Q_Glutamine
add_K_Lysine
add_Z_avg_QandE
add_E_Glutamic_Acid
add_M_Methionine
add_H_Histidine
add_F_Phenylalanine
add_R_Arginine
add_Y_Tyrosine
add_W_Tryptophan

residuemodification mass0 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

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