1. Proteins to Proteomes The InterPro Database

2. Origins of InterPro raw data UniProt Swiss-Prot TrEMBL 5M ??? InterPro
290K annotated
5M ???
automated annotation
InterPro

3. uncharacterised sequence feed back common annotation
Curated Annotation in InterPro
TrEMBL
uncharacterised sequence
TrEMBL
feed back common annotation
multiple signatures
InterPro
groups of related proteins
(same family or share domains)
annotated sequence
Swiss-Prot

4. Finding Conserved Signatures
Pattern
Simplest (limited)
Fingerprint
Sequence clustering
HMM
More information

5. Patterns Pattern/motif in sequence  regular expression
Can define important sites
Enzyme catalytic site Prosthetic group attachment Metal ion binding site Cysteines for disulphide bonds Protein or molecule binding
B chain xxxxxxCxxxxxxxxxxxxCxxxxxxxxx
A chain xxxxxCCxxxCxxxxxxxxCx
| |
EXAMPLE: Insulin

6. Patterns Pattern/motif in sequence  regular expression
Can define important sites
EXAMPLE: PS00262 Insulin family signature
B chain xxxxxxCxxxxxxxxxxxxCxxxxxxxxx
A chain xxxxxCCxxxCxxxxxxxxCx
| |
MALWMRLLPLLALLALWGPDPAAAFVNQHLCGSHLVEALYLVCGERGFFYTPKTRREAEDLQVGQVELGGGPGAGSLQPLALEGSLQKRGIVEQCCTSICSLYQLENYCN

7. Patterns Pattern/motif in sequence  regular expression
Can define important sites
EXAMPLE: PS00262 Insulin family signature
B chain xxxxxxCxxxxxxxxxxxxCxxxxxxxxx
A chain xxxxxCCxxxCxxxxxxxxCx
| |
MALWMRLLPLLALLALWGPDPAAAFVNQHLCGSHLVEALYLVCGERGFFYTPKTRREAEDLQVGQVELGGGPGAGSLQPLALEGSLQKRGIVEQ CCTSICSLYQLENYC N

8. Patterns C-C-{P}-x(2)-C-[STDNEKPI]-x(3)-[LIVMFS]-x(3)-C
Pattern/motif in sequence  regular expression
Can define important sites
EXAMPLE: PS00262 Insulin family signature
B chain xxxxxxCxxxxxxxxxxxxCxxxxxxxxx
A chain xxxxxCCxxxCxxxxxxxxCx
| |
MALWMRLLPLLALLALWGPDPAAAFVNQHLCGSHLVEALYLVCGERGFFYTPKTRREAEDLQVGQVELGGGPGAGSLQPLALEGSLQKRGIVEQ CCTSICSLYQLENYC N
Regular expression
C-C-{P}-x(2)-C-[STDNEKPI]-x(3)-[LIVMFS]-x(3)-C

9. Patterns PS00000 C-C-{P}-x(2)-C-[STDNEKPI]-x(3)-[LIVMFS]-x(3)-C
Sequence alignment
Insulin family motif
Define pattern
Extract pattern sequences
xxxxxx
C-C-{P}-x(2)-C-[STDNEKPI]-x(3)-[LIVMFS]-x(3)-C
Build regular expression
Pattern signature
PS00000

10. Fingerprints Several motifs  characterise family
Identify small conserved regions in divergent proteins
Different combinations of motifs describe subfamilies
EXAMPLE: PR00107 Phosphocarrier HPr signature
PTHP_ENTFA:
MEKKEFHIVAETGIHARPATLLVQTASKFNSDINLEYKGKSVNLKSIMGVMSLGVGQGSDVTITVDGADEAEGMAAIVETLQKEGLAE

11. Fingerprints Several motifs  characterise family
Identify small conserved regions in divergent proteins
Different combinations of motifs describe subfamilies
EXAMPLE: PR00107 Phosphocarrier HPr signature
PTHP_ENTFA:
MEKKEFHIVAET GIHARPATLLVQTASKF NSDINLEYKGKSVNLK SIMGVMSLGVGQGSDVTITVDGADEAEGMAAIVETLQKEGLAE
His phosphorylation site

12. Fingerprints Several motifs  characterise family
Identify small conserved regions in divergent proteins
Different combinations of motifs describe subfamilies
EXAMPLE: PR00107 Phosphocarrier HPr signature
PTHP_ENTFA:
MEKKEFHIVAET GIHARPATLLVQTASKF NSDINLEY KGKSVNLK SIMGVMSL GVGQGSDVTITVDGADEAEGMAAIVETLQKEGLAE
His phosphorylation site
Ser phosphorylation site

13. Fingerprints Several motifs  characterise family
Identify small conserved regions in divergent proteins
Different combinations of motifs describe subfamilies
EXAMPLE: PR00107 Phosphocarrier HPr signature
PTHP_ENTFA:
MEKKEFHIVAET GIHARPATLLVQTASK FNSDINLEY KGKSVNLK SIMGVMSL GVGQGSDVTITVDGADE AEGMAAIVETLQKEGLAE
His phosphorylation site
Conserved site
Ser phosphorylation site

14. Fingerprints Several motifs  characterise family
Identify small conserved regions in divergent proteins
Different combinations of motifs describe subfamilies
EXAMPLE: PR00107 Phosphocarrier HPr signature
PTHP_ENTFA:
MEKKEFHIVAET GIHARPATLLVQTASK FNSDINLEY KGKSVNLK SIMGVMSL GVGQGSDVTITVDGADE AEGMAAIVETLQKEGLAE
1) GIHARPATLLVQTASKF
2) KGKSVNLKSIMGVMSL
3) LGVGQGSDVTITVDGADE
3-motif fingerprint

15. Fingerprints 1 2 3 PR00000 Correct order Correct spacing
Ser phosphorylation site
Conserved site
His phosphorylation site
Define motifs
Sequence alignment
Extract motif sequences
xxxxxx
Fingerprint signature 1 2 3
Correct order
Correct spacing
PR00000

16. Recruit homologous domains
Sequence clustering
Automatic clustering of homologous domains
**Rarely covers entire domain (conserved core)
**Signature size can change with release
Known domain families
Recruit homologous domains
PSI-BLAST
MKDOM2
Automatic clustering
ProDomAlign
Align domain families

17. Hidden Markov Models (HMM)
Can characterise protein over entire length
Models conserved and divergent regions (position-specific scoring)
Models insertions and deletions
Outperform in sensitivity and specificity
More flexible (can use partial alignments)

18. (residue frequency at each position in alignment)
Sequence alignment
Sequence 1:
Sequence 2:
Sequence 3:
Sequence 4:
Sequence 5:
Sequence 6:
Sequence 7:
Scoring matrix
(residue frequency at each position in alignment)
Profile

19. Phe, Tyr and Leu found at position 1 of alignment
Sequence 1:
Sequence 2:
Sequence 3:
Sequence 4:
Sequence 5:
Sequence 6:
Sequence 7:
Phe, Tyr and Leu found at position 1 of alignment
Phe most conserved
highest match value

20. Probability method gauges scoring parameters
Sequence 1:
Sequence 2:
Sequence 3:
Sequence 4:
Sequence 5:
Sequence 6:
Sequence 7:
Tyr and Leu found at equal frequency at position 1
Tyr closer to Phe than Leu
Scores: F > Y > L
Probability method gauges scoring parameters

21. Hidden Markov Models (HMM)
Sequence alignment M1 M2 M3 M4
Begin
End
M = match state

22. Hidden Markov Models (HMM)
I = insert state, I2 I3 M1 M2 M3 M4
Begin
End D1 D4 D2
D = delete state D3
M = match state,

23. SAM Profile HMMs Homologous structural superfamilies
Start with single seed sequence
Create 1 model for every protein in superfamily  combine results
Few proteins in family have PDB structures
Proteins in superfamily may have low sequence identity

24. Specialisation of Databases
PRINTS
Describe sibling families
PROSITE
Identify binding and active sites
PRODOM
Describe conserved core of domains
PFAM
Wide coverage of domains & families
SMART
Signalling, extracellular & nuclear domains
TIGRFAM
Functional classification of families
PIRSF
Families conserved in domain composition
PANTHER
Functional classification of families
GENE3D
Structural-based domain classification
Superfam
Structural-based domain classification

25. Foundations of InterPro
Integration of signatures
InterPro
Manual curation

26. InterPro Entry Groups similar signature together
Links related signatures
Adds extensive annotation
Linked to other databases
Structural information and viewers

27. Assigning Type Family Full-length signatures grouping related proteins
Domain
Biological units with defined boundaries
Repeat
Signature repeated as a series of short motifs
Site
Protein feature described by a Prosite pattern
Region
Any signature that doesn’t fit the above

28. Grouping Signatures Together
PFAM
PROSITE 1)
(100)
Same positions
Same protein hits
IPR000001
Same positions
Different protein hits 2)
PFAM
PROSITE
(100)
(50)
IPR000001
IPR000002
PROSITE
PFAM 3)
(100)
Different positions
Same protein hits
IPR000001
IPR000002
Different positions 4)
PFAM
PROSITE
(100)
IPR000001
IPR000002

29. Applies to domains and families
Link related signatures - relationships
1) Parent - Child
(subgroup of more closely related proteins) *
PFAM
(100)
Protein kinase
PFAM
(75)
(100)
SMART
Protein kinase
Serine kinase
PFAM
Protein kinase
SMART
PROSITE
Serine kinase
Tyrosine kinase
Parent
Children
PROSITE
(25)
Tyrosine kinase
No proteins in common
SMART
PROSITE
Applies to domains and families

30. Both families and domains can contain domains
Link related signatures - relationships
2) Contains – Found in
(Describes domain composition)
PFAM
Receptor family
PROSITE
C-terminal domain
SMART
N-terminal domain
Found in
(Pfam)
Contains
(Smart and Prosite)
PFAM
Receptor Family
SMART
PROSITE
N-terminal domain
C-terminal domain
Both families and domains can contain domains

31. Link related signatures - relationships
2) Contains – Found in
Coverage
Signature must cover the entire (>90%) sequence of contained signature
Contains
PFAM
Found in
SMART
PFAM
SMART
Contains
Found in
Overlapping

32. Criteria for Signature InterPro Relationship
Relationships – evolutionary context
Criteria for Signature
InterPro Relationship
Structural family
Grandparent
GENE3D
Parents
PFAM
Sequence families
Children
TIGRFAM
Functional families
Unique to InterPro

33. Extensive Annotation Annotation Fields in InterPro Name and short name
Entry type (family, domain, site)
Relationships (links related signatures)
GO mapping ( large scale classification)
Abstract
Taxonomy (search/download using taxonomy)
Examples
Publications

34. Select species-specific protein sets
Extensive Annotation
Annotation Fields in InterPro
Name and short name
Entry type (family, domain, site)
Relationships (links related signatures)
GO mapping ( large scale classification)
Abstract
Taxonomy (search/download using taxonomy)
Examples
Publications
Select species-specific protein sets

35. Links to Other Databases
Annotation Fields in InterPro
Blocks (family alignments)
IntEnz (enzymes)
Prosite documents
COME (bioinorganic motifs)
CAZy (carbohydrate-active enzymes)
IUPHAR (GPCR receptors)
CluS-Tr (protein clusters)
Pandit (phylogenetic trees of PFAMs)
Merops (peptidases & inhibitors)

36. Structural information
Structures
PDB
Classification
CATH
SCOP
Homology Models
Swiss-Model
ModBase

37. Sequence-Structure Display
Signatures predictive of protein annotation
Structural data for specific proteins
AstexViewer® for structure

38. Structure Viewer Manipulate structures
Navigate between structure and sequence

39. Other Features – splice variants

40. Each ‘balloon’ represents a linked InterPro domain
Other Features – domain architecture
Each ‘balloon’ represents a linked InterPro domain
Select data set of these proteins

41. Other Features – protein-protein interactions
Lists proteins in entry known to be involved in protein-protein interactions
IntAct database of interactions

42. Protein Sequence Coverage
InterPro signatures cover:
95% of UniProt/Swiss-Prot proteins
79% of UniProt/TrEMBL proteins
>4 million matches in InterPro
>50,000 signature methods
>16,000 InterPro entries

43. Searching InterPro Search tools include: Text Search
InterProScan (sequence search)

44. InterPro Text Search Text search box Search results Search using: text
protein ID
InterPro ID
GO term
Search results
Direct links to entry

45. Use ftp site to run multiple sequences simultaneously
InterProScan Search
Use ftp site to run multiple sequences simultaneously
Member database search engines
Paste in sequence (protein/nucleotide)

46. Direct links to signature databases
InterProScan Search Results
single InterPro entry
Direct links to entry
Direct links to signature databases