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digest
Function
Protein proteolytic enzyme or reagent cleavage digest
Description
digest finds the positions where a specified proteolytic enzyme or
reagent might cut a peptide sequence.
This programs allows you to input a protein sequence and to specify
one proteolytic agent from a list. It will then output a file
containing the positions where the agent cuts, together with the
peptides produced.
Trypsin will not normally cut after a K if it is followed by (e.g.)
another K or a P. Specifying the qualifier -unfavoured shows those
cuts. as well as the favoured ones.
If you wish to emulate a partial digestion, then using the -overlap
qualifier will display the results from a digest in which all cut
sites are used and in which one site at a time is not cut. Thus the
resulting peptide fragments from the cut sites numbered 1, 2, 3, 4
etc. are shown, plus the fragments produced by cutting at the sites
1..3, 2..4, etc.
i.e. fragments containing one potential cut site are also shown.
If you wish to emulate a very partial digestion (!) then using the
-allpartials qualifier will do what -overlap did, but also show all
possible fragments from using all possible combinations of cut sites.
For example, fragments from the cut sites numbered 1, 2, 3, 4, 5 and
from the fragments produced by cutting between sites 1..3, 1..4, 1..5,
2..4, 2..5, 3..5, etc.
i.e. fragments containing one or more potential cut site are also
shown.
If the boolean -ragging is specified then terminal break-up fragments
are also shown; this can be useful for mass spectrometry. Ragging is
further controlled by the -termini option which allows you to select
removal of residues from the N and/or C termini.
Usage
Here is a sample session with digest
% digest
Protein proteolytic enzyme or reagent cleavage digest
Input protein sequence(s): tsw:opsd_human
Enzymes and Reagents
1 : Trypsin
2 : Lys-C
3 : Arg-C
4 : Asp-N
5 : V8-bicarb
6 : V8-phosph
7 : Chymotrypsin
8 : CNBr
Select number [1]:
Output report [opsd_human.digest]:
Go to the input files for this example
Go to the output files for this example
Command line arguments
Standard (Mandatory) qualifiers:
[-seqall] seqall Protein sequence(s) filename and optional
format, or reference (input USA)
-menu menu [1] Select number (Values: 1 (Trypsin); 2
(Lys-C); 3 (Arg-C); 4 (Asp-N); 5
(V8-bicarb); 6 (V8-phosph); 7
(Chymotrypsin); 8 (CNBr))
[-outfile] report [*.digest] Output report file name
Additional (Optional) qualifiers: (none)
Advanced (Unprompted) qualifiers:
-aadata datafile [Eamino.dat] Molecular weight data for amino
acids
-unfavoured boolean Trypsin will not normally cut after a K if
it is followed by (e.g.) another K or a P.
Specifying this shows those cuts. as well as
the favoured ones.
-ragging boolean Allows semi-specific and non-specific
digestion. This option is particularly
useful for generating lists of peptide
sequences for protein identification using
mass-spectrometry.
-termini menu [1] Select number (Values: 1 (none); 2
(nterm); 3 (cterm); 4 (nterm OR cterm))
-overlap boolean Used for partial digestion. Shows all cuts
from favoured cut sites plus 1..3, 2..4,
3..5 etc but not (e.g.) 2..5. Overlaps are
therefore fragments with exactly one
potential cut site within it.
-allpartials boolean As for overlap but fragments containing more
than one potential cut site are included.
Associated qualifiers:
"-seqall" associated qualifiers
-sbegin1 integer Start of each sequence to be used
-send1 integer End of each sequence to be used
-sreverse1 boolean Reverse (if DNA)
-sask1 boolean Ask for begin/end/reverse
-snucleotide1 boolean Sequence is nucleotide
-sprotein1 boolean Sequence is protein
-slower1 boolean Make lower case
-supper1 boolean Make upper case
-sformat1 string Input sequence format
-sdbname1 string Database name
-sid1 string Entryname
-ufo1 string UFO features
-fformat1 string Features format
-fopenfile1 string Features file name
"-outfile" associated qualifiers
-rformat2 string Report format
-rname2 string Base file name
-rextension2 string File name extension
-rdirectory2 string Output directory
-raccshow2 boolean Show accession number in the report
-rdesshow2 boolean Show description in the report
-rscoreshow2 boolean Show the score in the report
-rusashow2 boolean Show the full USA in the report
-rmaxall2 integer Maximum total hits to report
-rmaxseq2 integer Maximum hits to report for one sequence
General qualifiers:
-auto boolean Turn off prompts
-stdout boolean Write standard output
-filter boolean Read standard input, write standard output
-options boolean Prompt for standard and additional values
-debug boolean Write debug output to program.dbg
-verbose boolean Report some/full command line options
-help boolean Report command line options. More
information on associated and general
qualifiers can be found with -help -verbose
-warning boolean Report warnings
-error boolean Report errors
-fatal boolean Report fatal errors
-die boolean Report dying program messages
Input file format
Any protein sequence.
Input files for usage example
'tsw:opsd_human' is a sequence entry in the example protein database
'tsw'
Database entry: tsw:opsd_human
ID OPSD_HUMAN Reviewed; 348 AA.
AC P08100; Q16414; Q2M249;
DT 01-AUG-1988, integrated into UniProtKB/Swiss-Prot.
DT 01-AUG-1988, sequence version 1.
DT 20-MAR-2007, entry version 91.
DE Rhodopsin (Opsin-2).
GN Name=RHO; Synonyms=OPN2;
OS Homo sapiens (Human).
OC Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi;
OC Mammalia; Eutheria; Euarchontoglires; Primates; Haplorrhini;
OC Catarrhini; Hominidae; Homo.
OX NCBI_TaxID=9606;
RN [1]
RP NUCLEOTIDE SEQUENCE [GENOMIC DNA].
RX MEDLINE=84272729; PubMed=6589631;
RA Nathans J., Hogness D.S.;
RT "Isolation and nucleotide sequence of the gene encoding human
RT rhodopsin.";
RL Proc. Natl. Acad. Sci. U.S.A. 81:4851-4855(1984).
RN [2]
RP NUCLEOTIDE SEQUENCE [GENOMIC DNA].
RA Suwa M., Sato T., Okouchi I., Arita M., Futami K., Matsumoto S.,
RA Tsutsumi S., Aburatani H., Asai K., Akiyama Y.;
RT "Genome-wide discovery and analysis of human seven transmembrane helix
RT receptor genes.";
RL Submitted (JUL-2001) to the EMBL/GenBank/DDBJ databases.
RN [3]
RP NUCLEOTIDE SEQUENCE [LARGE SCALE MRNA].
RC TISSUE=Retina;
RG The German cDNA consortium;
RL Submitted (JUN-2003) to the EMBL/GenBank/DDBJ databases.
RN [4]
RP NUCLEOTIDE SEQUENCE [LARGE SCALE MRNA].
RX PubMed=15489334; DOI=10.1101/gr.2596504;
RG The MGC Project Team;
RT "The status, quality, and expansion of the NIH full-length cDNA
RT project: the Mammalian Gene Collection (MGC).";
RL Genome Res. 14:2121-2127(2004).
RN [5]
RP NUCLEOTIDE SEQUENCE [GENOMIC DNA] OF 1-120.
RX PubMed=8566799; DOI=10.1016/0378-1119(95)00688-5;
RA Bennett J., Beller B., Sun D., Kariko K.;
RT "Sequence analysis of the 5.34-kb 5' flanking region of the human
RT rhodopsin-encoding gene.";
RL Gene 167:317-320(1995).
RN [6]
RP REVIEW ON RP4 VARIANTS.
RX MEDLINE=94004905; PubMed=8401533;
RA Al-Maghtheh M., Gregory C., Inglehearn C., Hardcastle A.,
RA Bhattacharya S.;
[Part of this file has been deleted for brevity]
FT /FTId=VAR_004816.
FT VARIANT 209 209 V -> M (effect not known).
FT /FTId=VAR_004817.
FT VARIANT 211 211 H -> P (in RP4).
FT /FTId=VAR_004818.
FT VARIANT 211 211 H -> R (in RP4).
FT /FTId=VAR_004819.
FT VARIANT 216 216 M -> K (in RP4).
FT /FTId=VAR_004820.
FT VARIANT 220 220 F -> C (in RP4).
FT /FTId=VAR_004821.
FT VARIANT 222 222 C -> R (in RP4).
FT /FTId=VAR_004822.
FT VARIANT 255 255 Missing (in RP4).
FT /FTId=VAR_004823.
FT VARIANT 264 264 Missing (in RP4).
FT /FTId=VAR_004824.
FT VARIANT 267 267 P -> L (in RP4).
FT /FTId=VAR_004825.
FT VARIANT 267 267 P -> R (in RP4).
FT /FTId=VAR_004826.
FT VARIANT 292 292 A -> E (in CSNBAD1).
FT /FTId=VAR_004827.
FT VARIANT 296 296 K -> E (in RP4).
FT /FTId=VAR_004828.
FT VARIANT 297 297 S -> R (in RP4).
FT /FTId=VAR_004829.
FT VARIANT 342 342 T -> M (in RP4).
FT /FTId=VAR_004830.
FT VARIANT 345 345 V -> L (in RP4).
FT /FTId=VAR_004831.
FT VARIANT 345 345 V -> M (in RP4).
FT /FTId=VAR_004832.
FT VARIANT 347 347 P -> A (in RP4).
FT /FTId=VAR_004833.
FT VARIANT 347 347 P -> L (in RP4; common variant).
FT /FTId=VAR_004834.
FT VARIANT 347 347 P -> Q (in RP4).
FT /FTId=VAR_004835.
FT VARIANT 347 347 P -> R (in RP4).
FT /FTId=VAR_004836.
FT VARIANT 347 347 P -> S (in RP4).
FT /FTId=VAR_004837.
SQ SEQUENCE 348 AA; 38893 MW; 6F4F6FCBA34265B2 CRC64;
MNGTEGPNFY VPFSNATGVV RSPFEYPQYY LAEPWQFSML AAYMFLLIVL GFPINFLTLY
VTVQHKKLRT PLNYILLNLA VADLFMVLGG FTSTLYTSLH GYFVFGPTGC NLEGFFATLG
GEIALWSLVV LAIERYVVVC KPMSNFRFGE NHAIMGVAFT WVMALACAAP PLAGWSRYIP
EGLQCSCGID YYTLKPEVNN ESFVIYMFVV HFTIPMIIIF FCYGQLVFTV KEAAAQQQES
ATTQKAEKEV TRMVIIMVIA FLICWVPYAS VAFYIFTHQG SNFGPIFMTI PAFFAKSAAI
YNPVIYIMMN KQFRNCMLTT ICCGKNPLGD DEASATVSKT ETSQVAPA
//
Output file format
The output is a standard EMBOSS report file.
The results can be output in one of several styles by using the
command-line qualifier -rformat xxx, where 'xxx' is replaced by the
name of the required format. The available format names are: embl,
genbank, gff, pir, swiss, trace, listfile, dbmotif, diffseq, excel,
feattable, motif, regions, seqtable, simple, srs, table, tagseq
See: http://emboss.sf.net/docs/themes/ReportFormats.html for further
information on report formats.
By default digest writes a 'seqtable' report file.
Output files for usage example
File: opsd_human.digest
########################################
# Program: digest
# Rundate: Sun 15 Jul 2007 12:00:00
# Commandline: digest
# -seqall tsw:opsd_human
# Report_format: seqtable
# Report_file: opsd_human.digest
########################################
#=======================================
#
# Sequence: OPSD_HUMAN from: 1 to: 348
# HitCount: 14
#
# Complete digestion with Trypsin yields 14 fragments
#
#=======================================
Start End Mol_Weight Cterm Nterm Sequence
70 135 7129.319 R Y TPLNYILLNLAVADLFMVLGGFTSTLYTSLHGYFVFGP
TGCNLEGFFATLGGEIALWSLVVLAIER
178 231 6335.495 R E YIPEGLQCSCGIDYYTLKPEVNNESFVIYMFVVHFTIP
MIIIFFCYGQLVFTVK
22 69 5788.873 R T SPFEYPQYYLAEPWQFSMLAAYMFLLIVLGFPINFLTL
YVTVQHKKLR
253 296 5004.085 R S MVIIMVIAFLICWVPYASVAFYIFTHQGSNFGPIFMTI
PAFFAK
136 177 4600.460 R Y YVVVCKPMSNFRFGENHAIMGVAFTWVMALACAAPPLA
GWSR
1 21 2257.495 . S MNGTEGPNFYVPFSNATGVVR
297 311 1728.089 K Q SAAIYNPVIYIMMNK
232 245 1490.538 K A EAAAQQQESATTQK
326 339 1403.457 K T NPLGDDEASATVSK
315 325 1186.476 R N NCMLTTICCGK
340 348 902.950 K . TETSQVAPA
249 252 503.550 K M EVTR
312 314 449.504 K N QFR
246 248 346.378 K E AEK
#---------------------------------------
#---------------------------------------
The header information contains the program name, date of run, name of
the reagent used to digest the protein and the number of fragments
reported. The header will report if complete or partial digestion was
chosen.
The rest of the file consists of columns holding the following data:
* The start position of the fragment
* The end position of the fragment
* The molecular weight of the fragment
* The residue before the cut site ('.' if start of sequence)
* The residue after the second cut site ('.' if end of sequence)
* The sequence of the fragment.
Data files
None.
Notes
None.
References
None.
Warnings
None.
Diagnostic Error Messages
None.
Exit status
It always exits with a status of 0.
Known bugs
None.
See also
Program name Description
antigenic Finds antigenic sites in proteins
epestfind Finds PEST motifs as potential proteolytic cleavage sites
fuzzpro Protein pattern search
fuzztran Protein pattern search after translation
helixturnhelix Report nucleic acid binding motifs
oddcomp Find protein sequence regions with a biased composition
patmatdb Search a protein sequence with a motif
patmatmotifs Search a PROSITE motif database with a protein sequence
pepcoil Predicts coiled coil regions
preg Regular expression search of a protein sequence
pscan Scans proteins using PRINTS
sigcleave Reports protein signal cleavage sites
Author(s)
Alan Bleasby (ajb ebi.ac.uk)
European Bioinformatics Institute, Wellcome Trust Genome Campus,
Hinxton, Cambridge CB10 1SD, UK
History
Written (1999) - Alan Bleasby
Target users
This program is intended to be used by everyone and everything, from
naive users to embedded scripts.
Comments
None
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