Using the interface of webESPript

The Main frame
The Buttons frame

The Main frame

Fill up the www form by, at least, uploading a multiple alignment file in the section 'Aligned Sequences'. Strings of characters in blue are hyperlinks to the manual.
PDB files and DSSP files can be uploaded in the section 'Secondary Structure'. They can also be retrieved from a SRS or 3DBBROWSE server. If you do not see the buttons labelled 'Get pdb file from server' or 'Get dssp file from server', please ask your administrator.

The Buttons frame

• When the main form is filled, click on the SUBMIT button to start the program.
• The RESULT window must automaticaly appear on your screen once the program has been executed. If not, click on the RES button.
• Once the PostScript generated and if you are curious, click on the IN button to have a look to the ESPript command file. Copy and paste it if you want to use standalone ESPript.
• To read the ESPript logfile, click on the OUT button (always check warnings).
• Click on ADV (Advanced) or EXP (Expert) to customize your output. In ADV mode, you can introduce another secondary structure file, change labels of secondary elements, fiddle with special characters and calculate similarity scores between groups of sequences. In EXP mode, you can also define your own colours, shift sequence numbering... You can navigate between BEG, ADV and EXP modes without losing information in your query.
• If you are in ADVanced or EXPert mode, you can use the + buttons (+1, +3, -1, -3) to build a non-standard command file. If you click on '+1', the form for a new data set is created. The parameters specified for the data set 0 (i.e. the first one) are copied to 1. The data set 0 is refered by a [01] displayed at each header of the main form, and the 1 by a [01]. You can switch between the different data sets by using the scroll bar or by clicking on the hyperlink 1 of [01] (or 0 of [01]). Check the last example in this form to learn more on this option.
• Use the LOAD button to load a previously saved session. Uploaded files (file.aln, file.dssp ...) are saved with the session. Use the button 'do not load data files' if these files have been modified and reenter them. Alternatively, use buttons 'new' in main frame. Save your new session when completed.
• Use the SAVE button to save your session locally, so that you can use later the same parameters and files.
• Pay attention to the TIME LEFT bar. You must at least execute one command on the server each 45 minutes. Otherwise you are disconnected.
• Before leaving, click on the EXIT button: files are removed from server. This is important for security.

The Results window

Programs SPDB and DSSP

Examples

Session vp7_beg.sav for beginners

Sequences homologous to vp7_btv10, a viral protein whose 3D structure has been solved, have been aligned with CLUSTALW. The form vp7_beg.sav allows the user to create a figure in PostScript from the alignment and to display secondary structure elements of vp7_btv10. It has been created as follows:

-> Start webESPript
-> Save the alignment file vp7.aln on your disk, and upload it in the first section 'Aligned Sequences' of the form.
-> RUN webESPript, open the resulting image. A column is framed in blue, if more than 70% of its residues are similar according to physico-chemical properties (threshold is set to 0.7 in the section 'Similarity Calculations' of the form and the button '%Equivalent' is activated). Note that vp7_btv10 is the second sequence in the alignment.
-> Save the PDB file vp7_btv10.pdb on your disk and upload it in the second section 'Secondary Structure' of the form
-> RUN, open image, observe that the secondary structure elements displayed above sequence blocks are named vp7_btv1s. This is normal, by default they are aligned with the first sequence, here vp7_btv1s, displayed by ESPript. However this is wrong, your secondary structure elements refer to vp7_btv10.
-> Browse the form to the section 'Defining Groups'. Change the string all to 2 all
-> RUN, open image, vp7_btv10 is now the first displayed sequence and secondary structure elements are correctly aligned with it.
-> Check the log file by clicking on OUT. At its end, you will find the following sentence :
   Total number of columns 360 suggestion columns per line: 60
By default, residues are written on 70 columns. Change this default value to 60 in section 'Output Layout', in order to obtain a justified figure.
-> RUN ESPript and SAVE session

Remark: Warnings are written in the Results frame (as well as in the log file available by clicking on OUT in the buttons frame) if ESPript notices mismatches between residue names of sequence and structure. The program stops if the number of mismatches is higher than 50%.

Session vp7_adv.sav for advanced users (resulting PostScript, Gif)

This session uses the same CLUSTALW alignment as in mode BEGinner. You can start with the session vp7_beg.sav decribed above.

-> Start webESPript
-> LOAD previous session by using the buttons frame
-> RUN ESPript to check if everything is ok
-> Click on ADVanced to access new options

-> Click on the button 'relative accessibility' in section 'Secondary Structure' of the main frame, to show accessibility per residues with a blue bar below sequence blocks on the image.
-> Click on 'hide names' in the same section, to remove the string vp7_btv10 above sequence blocks, at the same line as secondary structure elements
-> RUN to check
-> Type the command U R 127 250 in section 'Special Character', to add two red triangles below residues 127 and 250 of the first displayed sequence, vp7_btv10. These triangles show the separation between an alpha domain (1-127, 250-349) and a beta domain (128-249)in the 3D structure
-> RUN to check
In the same section, type S B 168-170 178-180 to mark an RGD tripeptide observed at different positions in sequences. Then type:
X B 1-126 254-349 to colour secondary structure elements of the alpha domain in blue
X G 127-253 to colour secondary structure elements of the beta domain in green
T R 1 2 to colour in red the names of the first and second diplayed sequences.
Note, you must enter a carriage return between special commands.
-> RUN to check
-> Type in subsection 'Comment' of 'Output Layout' the sentence: Alignment for protein vp7
-> RUN to check

-> Type 2 1 3-6, carriage return, 7-8, carriage return, 9 in section 'Defining Groups', in order to define three groups of sequences according to phylogeny (the first group is made of btv sequences only) and to calculate in-group as well as cross-group similarities. This necessits the use of a scoring matrix, and the option 'Risler' is selected instead of '%Equivalent' in section 'Similarity Calculations'.
-> RUN

You can now observe
(i) at column 1 that all residues are identical and are boxed in red
(ii)at column 10 that residues of the first group are identical and are in red (all threonines) and that those of the second are similar (threonine and serine) and are in red ; the third group is made of a single sequence and the residue is in black. The global similarity score calculated from all groups is > 0.7 (which is the threshold entered in the section 'Similarity Calculation') and the column is framed in blue.
(iii) at column 19 that residues are not framed, the global similarity score being < 0.7. Moreover, residues of the second group are in black, because they are not similar (lysine and threonine) according to a in-groups score < 0.7 (the same threshold is used by the program for global scores and in-groups scores).
(iv) at column 79 that the column has a yellow background. Residues are similar in each group (in-groups scores > 0.7) but significantly different from the first group (all prolines) to the second (all histidines). Thus, the cross-groups score is > 0.5, which is the current threshold according to the section 'Similarity Calculation'.

Remark: Try this if you want a more colourful PostScript:
-> Select option 'Thermal' in section 'Layout' to obtain bold characters.
-> Type the command M Y all in section 'Special Characters' to add a yellow background on similar residues
-> RUN to check

Again, same CLUSTALW alignment, but information on the 3D structure of vp7_btv1s is now entered. The button +1 of the button frames is used for this purpose. The following form can be generated in mode ADVanced or EXPert. We will start from scratch.

-> Open webESPript
-> Upload vp7.aln in the section 'Aligned Files'
-> Upload vp7_btv10.pdb in the section 'Secondary Structures'
-> Type X B 1 in the section 'Special Commands and Characters', to colour in blue secondary structure elements and align them with the first displayed sequence.
-> Type h in subsection 'Replace labels' of 'Special Commands and Characters', in order to remove the label, h1, from the 310 helix
-> Type 2 all in the section 'Defining Groups', to display vp7_btv10 as first sequence in the alignment.
-> Type 60 in subsection 'Columns' of 'Output Layout' to justify sequences
-> RUN ESPript to check the result. It should be fine.

-> Click on 'hide sequence' in the first section and RUN again. Only secondary structures elements are now displayed.
-> Click on +1 in the buttons frame to duplicate the form. The main frame is now divided in two independant parts named [01] and [01]. Check that the alignment file vp7.aln has been copied to [01].

All following commands must be entered in the second part [01].

-> Disable option 'hide sequence', then enter a 'Vertical Shift' of -1 in section 'Output Layout'
-> RUN ESPript. You now have a gap between secondary structure elements of vp7_btv10 and sequences.

-> Enter vp7_btv1.dssp in the section 'Secondary Structures', which corresponds to vp7_btv1s
-> Type X R 2 in the section 'Special Commands and Characters', to colour in red secondary structure elements and align them with the second displayed sequence.
-> Type 2 all in the section 'Defining Groups'
-> RUN ESPript to check the result

Information on secondary structure elements are well aligned with sequences but labels of vp7_btv1s need to be removed

-> Click on 'hide labels' in section 'Secondary Structure'
-> RUN ESPript to check the result.

All information is now well displayed. We will add extra boxing on residues.

->Type Q P 1-3 169-171 in section 'Special Command and Characters'. This means that columns 169-171 of sequences 1-3 are boxed in pink. Note that the program uses columns numbering instead of residues numbering for the special commands Q, V and W. You can visualize columns numbers by clicking on the button 'Ruler' in the same section.

-> Type Q P 5-7 169-171, carriage return, Q P 8 180-182 in the same section to box all RGD tripeptides.
-> Select the mode 'Flashy' in section 'Output Layout' to add a yellow background on similar residues.
-> RUN ESPript

We will now colour sequence names of vp7_btv10 and vp7_btv1s.
Type in the same section 'Special Command and Characters'
-> T B 1 to colour the name of the first displayed sequence in blue
-> T R 2 to colour the name of the second displayed sequence in red

Finally, we will number all sequences.
-> Click on 'number sequences' in section 'Aligned sequences'
-> RUN. You should observe that secondary structure elements in blue are not aligned with sequences any more. Indeed, information entered in [01] and in [01] are treated separately by the program, and extra columns have been added in [01] so as to number all sequences. Thus, you must:
-> Browse to the first part [01] of the form and click on 'number sequences' too.
-> RUN
Remark: You may have the same problem with justification, if you use the option 'Replace sequences names' in mode EXPert. Sequences names must be changed in [01] and in [01].

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