1. Aligning Kinases Applying MSA Analysis to the CDK family

2. Building A Multiple Sequence Alignment

3. chite ---ADKPKRPLSAYMLWLNSARESIKRENPDFK-VTEVAKKGGELWRGLKD wheat --DPNKPKRAPSAFFVFMGEFREEFKQKNPKNKSVAAVGKAAGERWKSLSE trybr KKDSNAPKRAMTSFMFFSSDFRS----KHSDLS-IVEMSKAAGAAWKELGP mouse -----KPKRPRSAYNIYVSESFQ----EAKDDS-AQGKLKLVNEAWKNLSP ***. :::.:... :.. *. *: * chite AATAKQNYIRALQEYERNGG- wheat ANKLKGEYNKAIAAYNKGESA trybr AEKDKERYKREM--------- mouse AKDDRIRYDNEMKSWEEQMAE * :.*. : Extrapolation Motifs/Patterns Phylogeny Profiles Struc. Prediction Multiple Alignments Are CENTRAL to MOST Bioinformatics Techniques. Potential Uses of A Multiple Sequence Alignment?

4. 1 Organizing a Family Gathering The CDK example

5. Choosing the Right Sequences SwisProt Litterature Other Databases

6. Organizing the Data SRS Public Data IGS Data Aventis CDK Genecard Manual Automatic

7. Accessing the Data: The Fischer Server Fischer will Contain – A collection of Flat files – A secure SRS server – File Formats The server is a Technology Pipeline – Can be adapted in real time – Can be Transfered

8. Our CDK Data CDKs and CDK-like – Protein Information Functional Features Structural Information – Genomic Information Genes Variant SNPs

9. Our MSA dataset 29 amino acid sequences (CDKS and Aurora families, stemming from primary transcripts) – 2 isoforms of a cdk member 4 PDB structures : – 1MUO (AUR A) – 1BLX (CDK 6 ) – 1b38 (CDK 2) – 1H4L (CDK 5) Use of T-coffee release 1.78 with integration of the structure informations contained in pdb files

10. 2 Aligning The Sequences

11. Building A Multiple Sequence Alignment ClustalW T-Coffee Muscle Hand Editing Combination Comparison

12. Using Structural Information 3D-Coffee Struct Vs Struct Seq Vs Struct Thread Superpose Seq Vs Seq Local Global

13. Method

14. Accessing the Methods: Fischer Public 3D-Coffee server – igs-server.cnrs-mrs.fr/TCoffee/ Fischer – Latest version of T-Coffee – Customised parameters – Coktails of MSA methods

15. 3 Dressing Up a Multiple Sequence Alignment

16. Feature Dressing -25 Binding site -20 Phospho -40 nsSNP -50 Splice Site … Escript

17. Feature Dressing

18. 4 How Good Is The Alignment ????

19. T-Coffee CORE Evaluation

20. CORE index Specificity (  ) and Sensitivity (  )

21. Feature Based Evaluation

22. Features mapping on multiple alignment T-coffee ATP binding site Glycine loop ATP binding site Glycine loop Non-synonymous SNP ClustalW

23. Structure Based Evaluation APDB

25. Include Sequences with Known Structures – Do Not use Structural Information Score 1 – Use Structural Information:Score 2 If Score1 ~ Score 2 – Structural Information does not help much – The alignment is of reasonnable quality

26. Evaluating a Multiple Sequence Alignment T-Coffee CORE index Feature Based Library APDB

27. Maninupulating and Comparing Alignments Reformating/Processing – seq_reformat – extract_from_pdb Coloring – seq_reformat – ESCript Comparing – aln_compare

28. 5 Thinking Large ????

29. T-Coffee_dpa T-Coffee is limited to a small number of sequences T-coffee_dpa: Double Progressive Algo – Able to handle large datasets – 1000 sequences and more – Able to use structural information

30. Using A Multiple Sequence Alignment

31. 1 Exploring The Alignment

32. Cdk's signature Cdk's T-loop (orange) and aurora's Activating loop Substrat recognition motif

33. 2 Using The Alignment Does my Sequence Make Sense

34. Identifying Abnormalities within an MSA Insertion within the Nuc Binding Site…

35. Identifying Abnormalities within an MSA

37. Activation loop (orange)

38. Identifying Abnormalities within an MSA Retinoblastoma

39. 2 Using The Alignment Analysing the Structure with The Alignment

40. The Evoltionnary Trace

41. 3 Using The Alignment Spotting differences

42. What makes a CDK not and AurorA

43. 4 Clustering and Correlating

44. Function Trees Vs Lead Trees 1-Select Functionnaly Important Positions 2-Make a tree based on these positions 3-Compare the tree with the lead tree PROBLEMS: – Choose on the right positions – Describe the Leads with the right determinants