==== FunDi ====

=== Installation ===

Download it from https://github.com/dgaston/FunDi
=> ZIP button

Install it with these commands:
<code>
unzip dgaston-FunDi-229c4d6.zip
cd dgaston-FunDi-229c4d6/
unzip qmmraxml.zip
export PATH=$PATH:qmmraxml  # Or mv qmmraxml/qmmraxmlHPC to a PATH folder
</code>

if not working, you may need to recompile qmmraxmlHPC

<code>
unzip dgaston-FunDi-229c4d6.zip
cd dgaston-FunDi-229c4d6/
unzip qmmraxml.zip
cd qmmraxml
make  clean   # remove all files
make          # compile qmmraxmlHPC
cd ..
export PATH=$PATH:qmmraxml  # or move  qmmraxmlHPC in a path folder.
</code>

FunDi needs PERL release 5.10 or higher (maybe with threads enabled).





==== Identication of function sites in the Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) (EC 1.2.1.12)  enzyme ====

=== Execution ===
<code>
perl FunDi.pl -a GAPDH_alignment.phy -o gapdh_subtree -m LG -s gapdh_subtree.def -P qmmraxml -r 4 -t gapdh.tre
</code>
(It can take time)

Parameters:
  * -a: multiple alignment file in PHYLIP format.
  * -o: basename for output.
  * -m: Substitution matrix (qmmraxml => LG).
  * -s: definition of groups, one line = one subgroup.
  * -P: phylogenetic tool to estimate parameters.
  * -r: number of categories for the gamma distribution.
  * -t: phylogenetic tree in NEWiCK format.

More details with the help:
<code>
perl FunDi.pl -h
</code>

=== Results  ===

== Visualisation of sites ==

The most important file is "FunDi_Posterior_Scores.txt".\\
You can load it in your spreadsheet (tab separated, activate special detection of numbers).\\
The numbering of sites starts from 0.\\
Sites with a posterior probality P(FD) with 0.50 or more could be interesting.\\
In particular:\\
  * site  38 (= 39 in Jalview) (= 34 in the structure 2PKQ)
  * site 193 (=194 in Jalview) (=188 in the structure 2PKQ)
  * site 195 (=196 in Jalview) (=191 in the structure 2PKQ)

Open Jalview, load the file GAPDH_alignment.phy, remove the first line and visualisation the sites.


== 3D visualisation ==

Open PyMol and load protein 2PKQ
<code>
Plugins-> PDB Loader Service 
</code>

Change the visualisation mode
<code>
all->S->Show->as->Cartoon
all->C->Color by->chain
</code>

In the PyMol command line, select the sites of interest...
<code>
select ImportantSites, resi 34+188+191 
</code>
... and change the visualisation mode:
<code>
(ImportantSites)->S->Show->Spheres
(ImportantSites)->C->Color by->chain
</code>