Task 6 - EVfold
For the proteins used in this practical, structures have been determined. However, in real-life projects, you often do not have protein structures only sequences. However, structure often provides crucial information and furthers understanding of the proteins function. During this practical, you already predicted secondary structure elements from protein sequence and generated homology models from protein structures with sequence similarity to your protein.
correlated mutations
local method: mutual information
your protein
+ example P01112 (RASH_HUMAN)
http://pfam.sanger.ac.uk/family/Ras
calculating the evolutionary couplings
0. aligment (clustalw.... Pfam alignment good) Many sequences
1. a2m2lm.... => aligment 2. freecontact -> standard (installed on student computers)
Output -> all couplings + evolutionary coupling score (last column)
rank by score => look at distrubution, values, range
Meaning of score unclear
Take only scores for i+6, i.e. neighboring residues neglected, minimal 5 residues between coupled residues
Take ranking, check for each coupled pair the actual distance in the structure. TP: distance <= 5 AA (minimal distance of all pairs of all atoms of both residues)
EVcoupling
Check evolutionary hot spots, i.e. relevant residues, functionally important sites.
Take L couplings (L=length of protein sequence), sum scores for each residue. Analyze.
(Cell paper)
- compare to conservation, single site conservation
EVfold.org
create model
choose number of contacts: optimum ~ 60-70% of L 40% of L 100% of L
=> RMSD will be calculated by server, if you give PDB ID
PLM