1. Proteins Structure Predictions Structural Bioinformatics

2. Reminder 2 3.1 Final date to chose a project 10.1 Submission project overview (one page) -Title -Main question -Major Tools you are planning to use to answer the questions 11.1 /18.1– meetings on projects 9.3 Poster submission 16.3 Poster presentation

3. 3 In 22.12.2015 there were 114,402 protein structures in the protein structure database. Was solved in 1958 by Max Perutz John Kendrew of Cambridge University. (Won the 1962 and Nobel Prize in Chemistry ) The first high resolution structure of a protein-myoglobin

4. The 3D structure of a protein is stored in a coordinate file Each atom is represented by a coordinate in 3D (X, Y, Z)

5. The coordinate file can be viewed graphically RBP

6. 6 Predicting the three dimensional structure from sequence of a protein is very hard (some times impossible) However we can predict with relative high precision the secondary structure MERFGYTRAANCEAP…. What can we do to bridge the gap?? >10,000,000>100,000

7. What do we mean by Secondary Structure ? Secondary structure are the building blocks of the protein structure: =

8. 8 What do we mean by Secondary Structure ? Secondary structure is usually divided into three categories: Alpha helix Beta strand (sheet) Anything else – turn/loop

9. 9 The different secondary structures are combined together to form the Tertiary Structure of the Proteins

10. 10 RBP Globin Tertiary Secondary ? ? ?

11. Secondary Structure Prediction Given a primary sequence ADSGHYRFASGFTYKKMNCTEAA what secondary structure will it adopt (alpha helix, beta strand or random coil) ? 11

12. 12 Secondary Structure Prediction Methods Statistical methods –Based on amino acid frequencies –HMM (Hidden Markov Model) Machine learning methods –SVM, Neural networks

13. 13 Chou and Fasman (1974) Name P(a) P(b) P(turn) Alanine 142 83 66 Arginine 98 93 95 Aspartic Acid 101 54 146 Asparagine 67 89 156 Cysteine 70 119 119 Glutamic Acid 151 037 74 Glutamine 111 110 98 Glycine 57 75 156 Histidine 100 87 95 Isoleucine 108 160 47 Leucine 121 130 59 Lysine 114 74 101 Methionine 145 105 60 Phenylalanine 113 138 60 Proline 57 55 152 Serine 77 75 143 Threonine 83 119 96 Tryptophan 108 137 96 Tyrosine 69 147 114 Valine 106 170 50 The propensity of an amino acid to be part of a certain secondary structure (e.g. – Proline has a low propensity of being in an alpha helix or beta sheet  breaker) Not very useful for predictions Statistical Methods for SS prediction

14. What is missing? 14

15. 15 HMM enables us to calculate the probability of assigning a sequence to a specific secondary structure TGTAGPOLKCHIQWML HHHHHHHLLLLBBBBB p = ? HMM (Hidden Markov Model) An approach for predicting Secondary Structure considering dependency between the position

16. 16 The probability of observing a residue which belongs to an α-helix followed by a residue belonging to a turn = 0.15 The probability of observing Alanine as part of a β- sheet Table built according to large database of known secondary structures α-helix followed by α-helix Beginning with an α- helix

17. Example What is the probability that the sequence TGQ will be in a helical structure?? TGQ HHH p = 0.45 x 0.041 x 0.8 x 0.028 x 0.8x 0.0635 = 0.0020995 What can we learn from secondary structure predictions??

18. csc Mad Cow Disease PrP c to PrP sc PRP c PRP sc

19. Predicting 3D Structure based on homology Comparative Modeling/homology modeling Similar sequences suggests similar structure

20. Sequence and Structure alignments of two Retinol Binding Protein

21. How do we evaluate structure similarity?? Structure Alignment

22. Structure Alignments The outputs of a structural alignment are a superposition of the atomic coordinates and a minimal Root Mean Square Distance (RMSD) between the structures. There are many different algorithms for structural Alignment.

23. Atoms in Protein V Atoms in Protein W Atom N (x, y, z) The RMSD of two aligned structures indicates their divergence from one another. Low values of RMSD mean similar structures

24. 24 Different sequences can result in similar structures 1ecd2hhd RMSD<1

25. 25 We can learn about the important features which determine structure and function by comparing the sequences and structures ?

26. 26 The Globin Family

27. 27 Why is Proline 36 conserved in all the globin family ?

28. 28 Where are the gaps?? The gaps in the pairwise alignment are mapped to the loop regions

29. 29 How are remote homologs related in terms of their structure? b-lactoglobulin RBD

30. 30 PSI-BLAST alignment of RBP and  -lactoglobulin: iteration 3 Score = 159 bits (404), Expect = 1e-38 Identities = 41/170 (24%), Positives = 69/170 (40%), Gaps = 19/170 (11%) Query: 3 WVWALLLLAAWAAAERD--------CRVSSFRVKENFDKARFSGTWYAMAKKDPEGLFLQ 54 V L+ LA A + S V+ENFD ++ G WY + K Sbjct: 1 MVTMLMFLATLAGLFTTAKGQNFHLGKCPSPPVQENFDVKKYLGRWYEIEKIPASFE-KG 59 Query: 55 DNIVAEFSVDETGQMSATAKGRVRLLNNWDVCADMVGTFTDTEDPAKFKMKYWGVASFLQ 114 + I A +S+ E G + K V + ++ +PAK +++++ + Sbjct: 60 NCIQANYSLMENGNIEVLNKELSPDGTMNQVKGE--AKQSNVSEPAKLEVQFFPL----- 112 Query: 115 KGNDDHWIVDTDYDTYAVQYSCRLLNLDGTCADSYSFVFSRDPNGLPPEA 164 +WI+ TDY+ YA+ YSC + ++ R+P LPPE Sbjct: 113 MPPAPYWILATDYENYALVYSCTTFFWL--FHVDFFWILGRNPY-LPPET 159

31. 31 The Retinol Binding Proteinb-lactoglobulin

32. 32 MERFGYTRAANCEAP…. Taken together FUNCTION

33. Comparative Modeling Builds a protein structure model based on its alignment (sequence) to one or more related protein structures in the database Similar sequence suggests similar structure

34. Comparative Modeling General algorithm Modeling of a sequence based on known structures Consist of four major steps : 1.Finding a known structure(s) related to the sequence to be modeled (template), using sequence comparison methods such as PSI-BLAST 2. Aligning sequence with the templates 3. Building a model 4. Assessing the model

35. Comparative Modeling Accuracy of the comparative model is usually related to the sequence identity on which it is based >50% sequence identity = high accuracy 30%-50% sequence identity= 90% can be modeled <30% sequence identity =low accuracy (many errors) However other parameters (such as identify length) can influence the results

36. What is a good model? ModBase- for homology modelling https://modbase.compbio.ucsf.edu/

37. What is a good model?

39. Extra Slides (for your interest) 39

40. 40 3.6 residues 5.6 Å Alpha Helix : Pauling (1951) A consecutive stretch of 5-40 amino acids (average 10). A right-handed spiral conformation. 3.6 amino acids per turn. Stabilized by Hydrogen bonds

41. 41 Beta Strand : Pauling and Corey (1951) > An extended polypeptide chains is called β –strand (consists of 5-10 amino acids > The chains are connected together by Hydrogen bonds to form b-sheet β -strand β -sheet

42. 42 Loops Connect the secondary structure elements (alpha helix and beta strands). Have various length and shapes.