Data

1. Protein-protein interaction data
   A comprehensive list of interactions for the protein family under study should be compiled. As interaction data are typically generated only for proteins whose sequences have been deposited in databases, a recently published comprehensive phylogeny of the protein family under study may yield an upper estimate of the number and phylogenetic breadth of interaction data to be expected. In many cases recently published phylogenetic relationships need to be extracted from the publications itself, however a growing number of phylogenies are being uploaded in online databases such as TreeBASE (http://treebase.org/treebase-web/home.html) or Dryad (http://datadryad.org/).
   
   1. For obtaining data on protein-protein interactions, databases might be used. Prominent examples of such databases include BioGRID (http://thebiogrid.org/) (Chatr-Aryamontri et al., 2013), the Database of Interacting Proteins (http://dip.doe-mbi.ucla.edu/) (Salwinski et al., 2004), IntAct (http://www.ebi.ac.uk/intact/) (Orchard et al., 2014) and String (http://string-db.org/) (Franceschini et al., 2013), to mention but a few. Above all, UniProt (http://www.uniprot.org/) (UniProt 2015) provides cross-references to a number of these database, thus facilitating searches for potential interaction partners.
      
   2. Whereas database searches provide a good starting point, they very often do not capture all of the information available. It is therefore advisable to undertake a literature search. Special emphasize should be put on obtaining information from phylogenetically informative proteins, i.e. from proteins that occupy a position in the phylogeny that is critical for resolving the state of a particular trait (i.e. the character-state). Very often these are the early-diverging lineages, as their inclusion (together with more derived taxa) ensures that the whole phylogenetic breadth of a taxonomic group is captured. It might prove useful to obtain new experimental data for proteins that are phylogenetically especially informative. Indeed, generation of new protein-protein interaction data is often combined with character-state reconstruction to better understand the evolution of protein-protein interactions (Liu et al., 2010; Melzer et al., 2014; Li et al., 2015).
   
   
2. Sequence retrieval
   Protein or nucleotide sequences for phylogenetic reconstructions can be retrieved from the NCBI nucleotide collection (http://www.ncbi.nlm.nih.gov/nuccore) or the NCBI protein collection (http://www.ncbi.nlm.nih.gov/protein).

Software

1. For sequence alignment and subsequent phylogenetic reconstructions one or several of the following programs may be used:
   
   Table 1. Programs for sequence alignments and phylogenetic reconstructions
   

   Program	Purpose	Reference
   ExPASy translate	Translation of nucleotide sequences into amino acid sequences.	(Artimo et al., 2012) http://web.expasy.org/translate/
   Clustal 2	Sequence alignment. Suited especially for closely related sequences.	(Larkin et al., 2007) http://www.clustal.org/clustal2/
   MAFFT7	Sequence alignment. Suited for closely as well as more distantly related sequences.	(Katoh and Standley, 2013) http://mafft.cbrc.jp/alignment/software/
   RevTrans 1.4	Converting amino acid alignment into codon alignment.	(Wernersson and Pedersen, 2003) http://www.cbs.dtu.dk/services/RevTrans/
   MEGA 6	Sequence alignment and phylogenetic reconstruction.	(Tamura et al., 2013) http://www.megasoftware.net/
   MrBayes 3	Phylogenetic reconstruction.	(Ronquist and Huelsenbeck, 2003) http://mrbayes.sourceforge.net/index.php

   
   
2. To collate the character matrix
   Microsoft Excel or a similar spreadsheet application
   
3. For character-state reconstruction:
   Mesquite 3.02 (Maddison and Maddison, 2015) (http://mesquiteproject.org/)
   Mesquite also provides extensive documentation: (http://mesquiteproject.wikispaces.com/)
   

Procedure