                                     iep



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Function

   Calculate the isoelectric point of proteins

Description

   iep calculates the isoelectric point of a protein from its amino acid
   composition assuming that no electrostatic interactions change the
   propensity for ionization. Optionally, iep will plot the ionization
   curve with respect to pH and write an output file of the data, where
   for each pH point the number of bound electrons and charge is given.

   For peptide fragments it is possible to exclude the charges of the
   amino and carboxy terminal residues.

Usage

   Here is a sample session with iep


% iep tsw:laci_ecoli
Calculate the isoelectric point of proteins
Output file [laci_ecoli.iep]:


   Go to the input files for this example
   Go to the output files for this example

   Example 2


% iep tsw:ifna2_human -disulphide 2 -lysinemodified 2
Calculate the isoelectric point of proteins
Output file [ifna2_human.iep]:


   Go to the input files for this example
   Go to the output files for this example

Command line arguments

Calculate the isoelectric point of proteins
Version: EMBOSS:6.6.0.0

   Standard (Mandatory) qualifiers (* if not always prompted):
  [-sequence]          seqall     Protein sequence(s) filename and optional
                                  format, or reference (input USA)
*  -graph              xygraph    [$EMBOSS_GRAPHICS value, or x11] Graph type
                                  (ps, hpgl, hp7470, hp7580, meta, cps, x11,
                                  tek, tekt, none, data, xterm, png, gif, pdf,
                                  svg)
*  -outfile            outfile    [*.iep] Output file name

   Additional (Optional) qualifiers:
   -amino              integer    [1] Number of N-termini (Integer 0 or more)
   -carboxyl           integer    [1] Number of C-termini (Integer 0 or more)
   -[no]termini        boolean    [Y] Include charge at N and C terminus
   -lysinemodified     integer    [0] Number of modified lysines (Integer 0 or
                                  more)
   -disulphides        integer    [0] Number of disulphide bridges (Integer 0
                                  or more)

   Advanced (Unprompted) qualifiers:
   -pkdata             datafile   [Epk.dat] Values of pKa for amino acids
   -step               float      [.5] Step value for pH (Number from 0.010 to
                                  1.000)
   -plot               toggle     [N] Plot charge vs pH
   -[no]report         toggle     [Y] Write results to a file

   Associated qualifiers:

   "-sequence" 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
   -scircular1         boolean    Sequence is circular
   -squick1            boolean    Read id and sequence only
   -sformat1           string     Input sequence format
   -iquery1            string     Input query fields or ID list
   -ioffset1           integer    Input start position offset
   -sdbname1           string     Database name
   -sid1               string     Entryname
   -ufo1               string     UFO features
   -fformat1           string     Features format
   -fopenfile1         string     Features file name

   "-graph" associated qualifiers
   -gprompt            boolean    Graph prompting
   -gdesc              string     Graph description
   -gtitle             string     Graph title
   -gsubtitle          string     Graph subtitle
   -gxtitle            string     Graph x axis title
   -gytitle            string     Graph y axis title
   -goutfile           string     Output file for non interactive displays
   -gdirectory         string     Output directory

   "-outfile" associated qualifiers
   -odirectory         string     Output directory

   General qualifiers:
   -auto               boolean    Turn off prompts
   -stdout             boolean    Write first file to standard output
   -filter             boolean    Read first file from standard input, write
                                  first file to 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 and exit. 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
   -version            boolean    Report version number and exit


Input file format

   iep reads one or more protein sequences.

   The input is a standard EMBOSS sequence query (also known as a 'USA').

   Major sequence database sources defined as standard in EMBOSS
   installations include srs:embl, srs:uniprot and ensembl

   Data can also be read from sequence output in any supported format
   written by an EMBOSS or third-party application.

   The input format can be specified by using the command-line qualifier
   -sformat xxx, where 'xxx' is replaced by the name of the required
   format. The available format names are: gff (gff3), gff2, embl (em),
   genbank (gb, refseq), ddbj, refseqp, pir (nbrf), swissprot (swiss, sw),
   dasgff and debug.

   See: http://emboss.sf.net/docs/themes/SequenceFormats.html for further
   information on sequence formats.

  Input files for usage example

   'tsw:laci_ecoli' is a sequence entry in the example protein database
   'tsw'

  Database entry: tsw:laci_ecoli

ID   LACI_ECOLI              Reviewed;         360 AA.
AC   P03023; O09196; P71309; Q2MC79; Q47338;
DT   21-JUL-1986, integrated into UniProtKB/Swiss-Prot.
DT   19-JUL-2003, sequence version 3.
DT   13-JUN-2012, entry version 136.
DE   RecName: Full=Lactose operon repressor;
GN   Name=lacI; OrderedLocusNames=b0345, JW0336;
OS   Escherichia coli (strain K12).
OC   Bacteria; Proteobacteria; Gammaproteobacteria; Enterobacteriales;
OC   Enterobacteriaceae; Escherichia.
OX   NCBI_TaxID=83333;
RN   [1]
RP   NUCLEOTIDE SEQUENCE [GENOMIC DNA].
RX   MEDLINE=78246991; PubMed=355891; DOI=10.1038/274765a0;
RA   Farabaugh P.J.;
RT   "Sequence of the lacI gene.";
RL   Nature 274:765-769(1978).
RN   [2]
RP   NUCLEOTIDE SEQUENCE [GENOMIC DNA].
RA   Chen J., Matthews K.K.S.M.;
RL   Submitted (MAY-1991) to the EMBL/GenBank/DDBJ databases.
RN   [3]
RP   NUCLEOTIDE SEQUENCE [GENOMIC DNA].
RA   Marsh S.;
RL   Submitted (JAN-1997) to the EMBL/GenBank/DDBJ databases.
RN   [4]
RP   NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC   STRAIN=K12 / MG1655 / ATCC 47076;
RA   Chung E., Allen E., Araujo R., Aparicio A.M., Davis K., Duncan M.,
RA   Federspiel N., Hyman R., Kalman S., Komp C., Kurdi O., Lew H., Lin D.,
RA   Namath A., Oefner P., Roberts D., Schramm S., Davis R.W.;
RT   "Sequence of minutes 4-25 of Escherichia coli.";
RL   Submitted (JAN-1997) to the EMBL/GenBank/DDBJ databases.
RN   [5]
RP   NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC   STRAIN=K12 / MG1655 / ATCC 47076;
RX   MEDLINE=97426617; PubMed=9278503; DOI=10.1126/science.277.5331.1453;
RA   Blattner F.R., Plunkett G. III, Bloch C.A., Perna N.T., Burland V.,
RA   Riley M., Collado-Vides J., Glasner J.D., Rode C.K., Mayhew G.F.,
RA   Gregor J., Davis N.W., Kirkpatrick H.A., Goeden M.A., Rose D.J.,
RA   Mau B., Shao Y.;
RT   "The complete genome sequence of Escherichia coli K-12.";
RL   Science 277:1453-1474(1997).
RN   [6]
RP   NUCLEOTIDE SEQUENCE [LARGE SCALE GENOMIC DNA].
RC   STRAIN=K12 / W3110 / ATCC 27325 / DSM 5911;
RX   PubMed=16738553; DOI=10.1038/msb4100049;
RA   Hayashi K., Morooka N., Yamamoto Y., Fujita K., Isono K., Choi S.,
RA   Ohtsubo E., Baba T., Wanner B.L., Mori H., Horiuchi T.;
RT   "Highly accurate genome sequences of Escherichia coli K-12 strains


  [Part of this file has been deleted for brevity]

FT   CHAIN         1    360       Lactose operon repressor.
FT                                /FTId=PRO_0000107963.
FT   DOMAIN        1     58       HTH lacI-type.
FT   DNA_BIND      6     25       H-T-H motif.
FT   VARIANT     282    282       Y -> D (in T41 mutant).
FT   MUTAGEN      17     17       Y->H: Broadening of specificity.
FT   MUTAGEN      22     22       R->N: Recognizes an operator variant.
FT   CONFLICT    286    286       L -> S (in Ref. 1, 4 and 7).
FT   HELIX         6     11
FT   TURN         12     14
FT   HELIX        17     24
FT   HELIX        33     45
FT   HELIX        51     56
FT   STRAND       63     69
FT   HELIX        74     89
FT   STRAND       93     98
FT   STRAND      101    103
FT   HELIX       104    115
FT   TURN        116    118
FT   STRAND      122    126
FT   HELIX       130    139
FT   TURN        140    142
FT   STRAND      145    150
FT   STRAND      154    156
FT   STRAND      158    161
FT   HELIX       163    177
FT   STRAND      181    186
FT   HELIX       192    207
FT   STRAND      213    217
FT   HELIX       222    234
FT   STRAND      240    246
FT   HELIX       247    259
FT   TURN        265    267
FT   STRAND      268    271
FT   HELIX       277    281
FT   STRAND      282    284
FT   STRAND      287    290
FT   HELIX       293    308
FT   STRAND      314    319
FT   STRAND      322    324
FT   STRAND      334    338
FT   HELIX       343    353
FT   HELIX       354    356
SQ   SEQUENCE   360 AA;  38590 MW;  347A8DEE92D736CB CRC64;
     MKPVTLYDVA EYAGVSYQTV SRVVNQASHV SAKTREKVEA AMAELNYIPN RVAQQLAGKQ
     SLLIGVATSS LALHAPSQIV AAIKSRADQL GASVVVSMVE RSGVEACKAA VHNLLAQRVS
     GLIINYPLDD QDAIAVEAAC TNVPALFLDV SDQTPINSII FSHEDGTRLG VEHLVALGHQ
     QIALLAGPLS SVSARLRLAG WHKYLTRNQI QPIAEREGDW SAMSGFQQTM QMLNEGIVPT
     AMLVANDQMA LGAMRAITES GLRVGADISV VGYDDTEDSS CYIPPLTTIK QDFRLLGQTS
     VDRLLQLSQG QAVKGNQLLP VSLVKRKTTL APNTQTASPR ALADSLMQLA RQVSRLESGQ
//

  Input files for usage example 2

  Database entry: tsw:ifna2_human

ID   IFNA2_HUMAN             Reviewed;         188 AA.
AC   P01563; P01564; Q14606; Q96KI6;
DT   21-JUL-1986, integrated into UniProtKB/Swiss-Prot.
DT   21-JUL-1986, sequence version 1.
DT   16-MAY-2012, entry version 128.
DE   RecName: Full=Interferon alpha-2;
DE            Short=IFN-alpha-2;
DE   AltName: Full=Interferon alpha-A;
DE            Short=LeIF A;
DE   Flags: Precursor;
GN   Name=IFNA2;
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 / MRNA].
RX   MEDLINE=81052322; PubMed=6159538; DOI=10.1038/287411a0;
RA   Goeddel D.V., Yelverton E., Ullrich A., Heyneker H.L., Miozzari G.,
RA   Holmes W., Seeburg P.H., Dull T.J., May L., Stebbing N., Crea R.,
RA   Maeda S., McCandliss R., Sloma A., Tabor J.M., Gross M.,
RA   Familletti P.C., Pestka S.;
RT   "Human leukocyte interferon produced by E. coli is biologically
RT   active.";
RL   Nature 287:411-416(1980).
RN   [2]
RP   NUCLEOTIDE SEQUENCE [GENOMIC DNA / MRNA].
RX   MEDLINE=81148795; PubMed=6163083; DOI=10.1038/290020a0;
RA   Goeddel D.V., Leung D.W., Dull T.J., Gross M., Lawn R.M.,
RA   McCandliss R., Seeburg P.H., Ullrich A., Yelverton E., Gray P.W.;
RT   "The structure of eight distinct cloned human leukocyte interferon
RT   cDNAs.";
RL   Nature 290:20-26(1981).
RN   [3]
RP   NUCLEOTIDE SEQUENCE [GENOMIC DNA / MRNA].
RX   MEDLINE=82060261; PubMed=6170983; DOI=10.1073/pnas.78.9.5435;
RA   Lawn R.M., Gross M., Houck C.M., Franke A.E., Gray P.V., Goeddel D.V.;
RT   "DNA sequence of a major human leukocyte interferon gene.";
RL   Proc. Natl. Acad. Sci. U.S.A. 78:5435-5439(1981).
RN   [4]
RP   NUCLEOTIDE SEQUENCE [MRNA].
RC   TISSUE=Bone marrow tumor;
RX   MEDLINE=86069501; PubMed=3906813;
RA   Oliver G., Balbas P., Valle F., Soberon X., Bolivar F.;
RT   "Cloning of human leukocyte interferon cDNA and a strategy for its
RT   production in E. coli.";
RL   Rev. Latinoam. Microbiol. 27:141-150(1985).
RN   [5]
RP   NUCLEOTIDE SEQUENCE [GENOMIC DNA].


  [Part of this file has been deleted for brevity]

DR   GO; GO:0060338; P:regulation of type I interferon-mediated signaling pathwa
y; TAS:Reactome.
DR   GO; GO:0009615; P:response to virus; IEA:UniProtKB-KW.
DR   GO; GO:0060337; P:type I interferon-mediated signaling pathway; TAS:Reactom
e.
DR   Gene3D; G3DSA:1.20.1250.10; 4_helix_cytokine_core; 1.
DR   InterPro; IPR009079; 4_helix_cytokine-like_core.
DR   InterPro; IPR012351; 4_helix_cytokine_core.
DR   InterPro; IPR000471; Interferon_alpha/beta/delta.
DR   PANTHER; PTHR11691; Interferon_abd; 1.
DR   Pfam; PF00143; Interferon; 1.
DR   PRINTS; PR00266; INTERFERONAB.
DR   SMART; SM00076; IFabd; 1.
DR   SUPFAM; SSF47266; 4_helix_cytokine; 1.
DR   PROSITE; PS00252; INTERFERON_A_B_D; 1.
PE   1: Evidence at protein level;
KW   3D-structure; Antiviral defense; Complete proteome; Cytokine;
KW   Direct protein sequencing; Disulfide bond; Glycoprotein;
KW   Pharmaceutical; Polymorphism; Reference proteome; Secreted; Signal.
FT   SIGNAL        1     23
FT   CHAIN        24    188       Interferon alpha-2.
FT                                /FTId=PRO_0000016360.
FT   CARBOHYD    129    129       O-linked (GalNAc...).
FT                                /FTId=CAR_000049.
FT   DISULFID     24    121
FT   DISULFID     52    161
FT   VARIANT       6      6       A -> D (in dbSNP:rs35971916).
FT                                /FTId=VAR_055972.
FT   VARIANT      46     46       K -> R (in alpha-2B and alpha-2C;
FT                                dbSNP:rs1061959).
FT                                /FTId=VAR_004012.
FT   VARIANT      57     57       H -> R (in alpha-2C).
FT                                /FTId=VAR_013001.
FT   VARIANT     177    177       S -> L (in a breast cancer sample;
FT                                somatic mutation).
FT                                /FTId=VAR_036329.
FT   HELIX        33     44
FT   TURN         49     54
FT   HELIX        63     66
FT   STRAND       67     69
FT   STRAND       71     75
FT   HELIX        76     91
FT   HELIX        93     98
FT   HELIX       101    123
FT   HELIX       134    155
FT   HELIX       160    178
FT   TURN        179    182
SQ   SEQUENCE   188 AA;  21550 MW;  101DD21D394CBF97 CRC64;
     MALTFALLVA LLVLSCKSSC SVGCDLPQTH SLGSRRTLML LAQMRKISLF SCLKDRHDFG
     FPQEEFGNQF QKAETIPVLH EMIQQIFNLF STKDSSAAWD ETLLDKFYTE LYQQLNDLEA
     CVIQGVGVTE TPLMKEDSIL AVRKYFQRIT LYLKEKKYSP CAWEVVRAEI MRSFSLSTNL
     QESLRSKE
//

Output file format

   With -plot specified, a graphical output is produced.

   The output is to the specified graphics device.

   The results can be output in one of several formats by using the
   command-line qualifier -graph xxx, where 'xxx' is replaced by the name
   of the required device. Support depends on the availability of
   third-party software packages.

   The device names that output to a file are: ps (postscript), cps
   (colourps), png, gif, pdf, svg, hpgl, hp7470, hp7580, das, data.

   The other available device names are: meta, x11 (xwindows), tek
   (tek4107t), tekt (tektronix), xterm, text.

   Output can be turned off by specifying none (null).

   See: http://emboss.sf.net/docs/themes/GraphicsDevices.html for further
   information on supported devices.

  Output files for usage example

  File: laci_ecoli.iep

IEP of LACI_ECOLI from 1 to 360
Isoelectric Point = 6.8385

   pH     Bound    Charge
  1.00    81.96    37.96
  1.50    81.89    37.89
  2.00    81.65    37.65
  2.50    80.91    36.91
  3.00    78.79    34.79
  3.50    73.70    29.70
  4.00    65.15    21.15
  4.50    56.73    12.73
  5.00    51.75     7.75
  5.50    49.35     5.35
  6.00    47.60     3.60
  6.50    45.48     1.48
  7.00    43.37    -0.63
  7.50    41.85    -2.15
  8.00    40.66    -3.34
  8.50    39.41    -4.59
  9.00    38.01    -5.99
  9.50    36.14    -7.86
 10.00    32.99   -11.01
 10.50    28.45   -15.55
 11.00    23.58   -20.42
 11.50    19.41   -24.59
 12.00    15.19   -28.81
 12.50     9.75   -34.25
 13.00     4.64   -39.36
 13.50     1.75   -42.25
 14.00     0.59   -43.41

  Output files for usage example 2

  File: ifna2_human.iep

IEP of IFNA2_HUMAN from 1 to 188
Isoelectric Point = 5.7240

   pH     Bound    Charge
  1.00    52.98    22.98
  1.50    52.93    22.93
  2.00    52.77    22.77
  2.50    52.28    22.28
  3.00    50.87    20.87
  3.50    47.47    17.47
  4.00    41.62    11.62
  4.50    35.67     5.67
  5.00    32.10     2.10
  5.50    30.46     0.46
  6.00    29.48    -0.52
  6.50    28.46    -1.54
  7.00    27.44    -2.56
  7.50    26.58    -3.42
  8.00    25.80    -4.20
  8.50    24.95    -5.05
  9.00    23.99    -6.01
  9.50    22.70    -7.30
 10.00    20.46    -9.54
 10.50    17.02   -12.98
 11.00    13.16   -16.84
 11.50    10.04   -19.96
 12.00     7.49   -22.51
 12.50     4.72   -25.28
 13.00     2.23   -27.77
 13.50     0.84   -29.16
 14.00     0.28   -29.72

   For each pH point it gives the number of bound electrons and the
   charge.

Data files

   iep reads in local data file Epk.dat which contains amino acid pK
   values used to calculate the isoelectric point of the whole protein

   EMBOSS data files are distributed with the application and stored in
   the standard EMBOSS data directory, which is defined by the EMBOSS
   environment variable EMBOSS_DATA.

   To see the available EMBOSS data files, run:

% embossdata -showall

   To fetch one of the data files (for example 'Exxx.dat') into your
   current directory for you to inspect or modify, run:

% embossdata -fetch -file Exxx.dat


   Users can provide their own data files in their own directories.
   Project specific files can be put in the current directory, or for
   tidier directory listings in a subdirectory called ".embossdata". Files
   for all EMBOSS runs can be put in the user's home directory, or again
   in a subdirectory called ".embossdata".

   The directories are searched in the following order:
     * . (your current directory)
     * .embossdata (under your current directory)
     * ~/ (your home directory)
     * ~/.embossdata

   Here is the default Epk.dat file:
# pK values for amino acids
# O=Ornithine J=Hydroxyproline
#
# Amino acid    pK
Amino           8.6
Carboxyl        3.6

C               8.5
D               3.9
E               4.1
H               6.5
K               10.8
R               12.5
Y               10.1

Notes

   Adjusting the pH of an aqueous protein solution to the point where the
   numbers of positive and negative charges on the protein are equal
   brings the protein to its isoelectric point. This is often the point of
   lowest solubility, presumably because it is the point at which there
   are fewest intermolecular repulsions, so that the molecules tend to
   form aggregates.

   Modified lysines and disulphide bridges affect the contributions of
   those residues to the protein charge. A set number of each residue can
   be excluded.

References

   None.

Warnings

   None.

Diagnostic Error Messages

   None.

Exit status

   0 if successful.

Known bugs

   None.

See also

   Program name     Description
   abiview          Display the trace in an ABI sequencer file
   charge           Draw a protein charge plot
   cirdna           Draw circular map of DNA constructs
   hmoment          Calculate and plot hydrophobic moment for protein sequence(s)
   lindna           Draw linear maps of DNA constructs
   octanol          Draw a White-Wimley protein hydropathy plot
   pepinfo          Plot amino acid properties of a protein sequence in parallel
   pepnet           Draw a helical net for a protein sequence
   pepstats         Calculate statistics of protein properties
   pepwheel         Draw a helical wheel diagram for a protein sequence
   pepwindow        Draw a hydropathy plot for a protein sequence
   pepwindowall     Draw Kyte-Doolittle hydropathy plot for a protein
                    alignment
   plotorf          Plot potential open reading frames in a nucleotide sequence
   prettyplot       Draw a sequence alignment with pretty formatting
   prettyseq        Write a nucleotide sequence and its translation to file
   remap            Display restriction enzyme binding sites in a nucleotide sequence
   showfeat         Display features of a sequence in pretty format
   showpep          Display protein sequences with features in pretty format
   sixpack          Display a DNA sequence with 6-frame translation and ORFs

Author(s)

   Alan Bleasby
   European Bioinformatics Institute, Wellcome Trust Genome Campus,
   Hinxton, Cambridge CB10 1SD, UK

   Please report all bugs to the EMBOSS bug team
   (emboss-bug (c) emboss.open-bio.org) not to the original author.

History

   Completed 1st August 1999.

Target users

   This program is intended to be used by everyone and everything, from
   naive users to embedded scripts.

Comments

   None
