Sequence-based predictions (PKU)
Contents
Short Task Description
Go read..
and while you're at it, read some more
Our task is to use the primary sequence of our protein (and some other example proteins) to predict comparatively simple features like secondary structure, signal peptides and transmembrane regions and more advanced like GO terms and similar functional annotations with different tools. The used commands and programms are listed at the appropriate places (if short and interesting enough) or linked at their own site.
Secondary Structure Prediction
Test some SS-predictors ans compare the results to the 'gold standard' of dssp
ReProfSeq
reprof -i <uniprotID>.fasta
egrep -v "^#|^No" <uniprotID>.reprof |awk '{print $3}'|tr -d '\n' > <uniprotID>_reprof.secstruc
PsiPred
Webserver here
grep Pred: <uniprotID>.psipred_out |cut -d " " -f2|tr -d '\n' > <uniprotID>_psipred.secstruc
DSSP
Download dssp here and get PDB files matching the Uniprot entries
dssp -i <PDBID>.pdb > <PDBID>.dssp tail -n+29 <PDBID>.dssp |cut -c17|tr ' ' '-'|tr -d '\n' > <PDBID>_dssp.secstruc
PDB-Files contains only part of the structure or more than 1 chain! e.g. 117aa-424aa for PAH.
Aligned structure to sequence manually..
Script to calculate Q3 and SOV: ss_score.py
results
| Protein | ReprofSeq | |||||||
|---|---|---|---|---|---|---|---|---|
| UniprotID | Q_E | Q_H | Q_L | Q_3 | SOV_E | SOV_H | SOV_L | SOV |
| P00439 | 45.5 | 75.7 | 74.8 | 71.2 | 47.5 | 86.7 | 71.8 | 76.4 |
| P10775 | 23.1 | 71.9 | 62.0 | 61.8 | 21.9 | 87.2 | 66.9 | 70.6 |
| Q9X0E6 | 26.8 | 91.9 | 39.1 | 53.5 | 36.1 | 91.1 | 41.0 | 57.6 |
| Q08209 | 69.1 | 34.0 | 73.1 | 57.9 | 62.4 | 45.6 | 76.7 | 63.0 |
| Protein | PsiPred | |||||||
|---|---|---|---|---|---|---|---|---|
| UniprotID | Q_E | Q_H | Q_L | Q_3 | SOV_E | SOV_H | SOV_L | SOV |
| P00439 | 57.6 | 85.1 | 89.0 | 83.8 | 57.6 | 88.5 | 54.4 | 71.1 |
| P10775 | 92.3 | 90.3 | 94.7 | 92.5 | 92.7 | 95.5 | 95.7 | 95.2 |
| Q9X0E6 | 75.6 | 86.5 | 78.3 | 80.2 | 93.44 | 100.0 | 94.5 | 96.1 |
| Q08209 | 58.2 | 77.3 | 90.7 | 81.0 | 64.1 | 85.8 | 64.8 | 72.5 |
Disorderd Regions
Transmembreane Helices
Signalpeptides
P47863 from rat, rest human => type eukaryotes; command:
signalp -trunc 70 -t euk <ID>.fasta > <ID>.sigP_out
| SignalP v3 | SignalP v4 | Signal Peptide Database | |||
|---|---|---|---|---|---|
| UniprotID | SignalP-HMM | SignalP_NN | summary prediction | summary prediction | experimental |
| P00439 | 0% signal peptide or anchor | 5 times No | no signal peptide or anchor | no signal peptide or anchor | no evidence for peptide or anchor |
| P02768 | 100% signal peptide, 0% signal anchor | 5 times Yes | signal peptide | signal peptide | confirmed signal peptide |
| P11279 | 100% signal peptide, 0% signal anchor | 5 times Yes | signal peptide | signal peptide | confirmed signal peptide |
| P47863 | 52.6% signal peptide, 45.7 signal anchor | 4 No, 1 Yes | signal peptide | no signal peptide or anchor | no evidence for peptide or anchor |
P47863 reaches a high maximal signal peptide score (S-score) in signalP-NN and probabilities above the cut-off in signalP-HMM in version 3. In version 4 (webserver) neither version predicts a signal peptide for this protein and the probabilities/scores are well below the cut-off values. P47863 is a multipass membrane protein and signalPv3 most likely predicted a possible cleavage site for the N-terminal membrane region, classifying it wrongly as signal sequence.
GO Terms
GOPet
| GOPet predicted GO terms | ||||
|---|---|---|---|---|
| GO ID | Aspect | Confidence | GO Term | True/False |
| GO:0003824 | F | 94% | catalytic activity | true |
| GO:0016491 | F | 88% | oxidoreductase activity | true |
| GO:0004497 | F | 87% | monooxygenase activity | true |
| GO:0004505 | F | 84% | phenylalanine 4-monooxygenase activity | true |
| GO:0004510 | F | 80% | tryptophan 5-monooxygenase activity | false |
| GO:0004511 | F | 79% | tyrosine 3-monooxygenase activity | false |
| GO:0046872 | F | 78% | metal ion binding | true |
| GO:0005506 | F | 78% | iron ion binding | true |
| GO:0008199 | F | 72% | ferric iron binding | false |
| GO:0008198 | F | 72% | ferrous iron binding | false |
| GO:0016597 | F | 71% | amino acid binding | true |
ProtFun
# Functional category Prob Odds Amino_acid_biosynthesis => 0.210 9.530 Biosynthesis_of_cofactors 0.229 3.180 Cell_envelope 0.034 0.563 Cellular_processes 0.063 0.867 Central_intermediary_metabolism 0.061 0.970 Energy_metabolism 0.343 3.815 Fatty_acid_metabolism 0.025 1.889 Purines_and_pyrimidines 0.392 1.615 Regulatory_functions 0.020 0.125 Replication_and_transcription 0.118 0.438 Translation 0.204 4.630 Transport_and_binding 0.024 0.060 # Enzyme/nonenzyme Prob Odds Enzyme => 0.724 2.527 Nonenzyme 0.276 0.387 # Enzyme class Prob Odds Oxidoreductase (EC 1.-.-.-) 0.154 0.738 Transferase (EC 2.-.-.-) 0.271 0.785 Hydrolase (EC 3.-.-.-) 0.083 0.261 Lyase (EC 4.-.-.-) 0.047 1.002 Isomerase (EC 5.-.-.-) => 0.100 3.138 Ligase (EC 6.-.-.-) 0.019 0.370 # Gene Ontology category Prob Odds Signal_transducer 0.075 0.350 Receptor 0.003 0.016 Hormone 0.001 0.206 Structural_protein 0.005 0.166 Transporter 0.025 0.229 Ion_channel 0.010 0.168 Voltage-gated_ion_channel 0.005 0.232 Cation_channel 0.010 0.215 Transcription 0.043 0.334 Transcription_regulation 0.032 0.255 Stress_response 0.010 0.118 Immune_response 0.012 0.140 Growth_factor 0.006 0.407 Metal_ion_transport 0.009 0.020
Additional Predictions:
Feature Output summary SignalP 3.0 No signal peptide cleavage site predicted ProP 1.0 1 propeptide cleavage site predicted at position: 74 TargetP 1.1 No high confidence targeting predition NetPhos 2.0 22 putative phosphorylation sites at positions 16 23 40 70 110 196 250 303 339 391 411 22 105 189 278 418 24 77 198 268 277 317 NetOGlyc 3.1 No O-glycosylated sites predicted NetNGlyc 1.0 2 putative N-glycosylated sites at positions 61 376 TMHMM 2.0 No TM helices predicted
