Structural biology should be computable!

1. Structural biology should be computable! • Protein structures determined by amino acid sequences • Protein structures and complexes correspond to global free energy minima • Fundamental test of understanding and huge practical relevance

2. Model of energetics of inter and intramolecular interactions Design (Given Structure, Optimize Sequence) Prediction (Given Sequence, Optimize Structure) ROSETTA Ab initio structure Protein Structure Protein design prediction Protein-protein docking Protein-protein Interface design interactions

3. Model of macromolecular interactions • Removal of single methyl groups can destabilize proteins --> jigsaw puzzle-like packing crucial • Buried polar atoms almost always hydrogen bonded --> treat hydrogen bonding as accurately as possible • Exposed charge substitutions generally have little effect --> damp long range elctrostatics • Focus on short range interactions!

4. Conformational sampling Random Start Low-Resolution Monte Carlo Search (integrate out sidechain degrees of freedom) High-Resolution Refinement with full atomic detail Select lowest energy models 105 Predictions Jeff Gray (Hopkins), Ora Furman (Hebrew University), Chu Wang

5. Docking Low-Resolution Search • Monte Carlo Search • Rigid body translations and rotations • Residue-scale interaction potentials Protein representation: backbone atoms + average centroids

7. Docking Protocol (Target 12: cohesin-dockerin; unbound-bound) • Initial Search • Refinement Energy RMSD to arbitrary starting structure (Å) RMSD to starting structure of refinement

8. Target 12Cohesin-Dockerin Side Chain Flexibility dockerin • 0.46Å interface rmsd • 87% native contacts • 6% wrong contacts cohesin Ora Furman, Chu Wang red,orange– xray blue – model; green– unbound

9. Details of T12 Interface dockerin R53 S45 D39 L22 N37 Y74 L83 E86 cohesin red,orange– xray blue - model

10. Target 15immunity protein D-colicin D tRNase Accurate Side Chain Modeling colicin • 0.23Å interface rmsd Science 310, 638-642 immunity protein red,orange– xray blue - model

11. Details of T15 Interface colicin H611 K607 K610 K608 E56 E68 D61 E59 red,orange– xray blue - model immunity protein

12. Modeling Backbone Movement Target 20HemK-RF1 • 2.34Å interface rmsd • 36% native contacts RF1 HemK Loop with methylated Gln red,orange– xray blue – model; green– unbound Chu Wang

14. Phil Bradley CASP6 T0198: PhoU domain repeat Model 2: 4A over 210 rsds (Model 1: 3.94 over 198)

15. CASP6 T0212 Model 2: 3.97 over 109 rsds (Model 1: 4.0 over 104)

18. Phil Bradley T0281 ab initio prediction (1.59Å)

20. 1r69

22. Science 309, 1868-1871 1ubq

24. 2REB

25. Boinc.bakerlab.org/rosettaDavid Kim High resolution ab initio structure prediction from single sequences by enhanced diversity “barcode” directed sampling Outreach!

30. High Resolution Refinement of CASP target 199 - remote homology model Bin Qian Calculations performed on SDSC teragrid clusters

31. High Resolution NMR Model Refinement Vatson Raman Disulfide Bond Formation Protein Blue - X-ray structure Green - NMR models Red - Rosetta models

32. Computing Structural Biology • Free energy function reasonable => Computing simple protein structures and interactions now appears to be within reach • Implications for structural genomics? • More cpu power => more accurate predictions for larger proteins • For larger complexes, experimental data essential (low resolution electron density!). • Symmetry helps! Modeling accuracy also illustrated by structures of designed proteins

33. Top7 X-ray structure has correct topology. Backbone RMSD to design only 1.2Å!! C-a Backbone Overlay Red : X-ray structure Blue : Design model Brian Kuhlman, Gautam Dantas; Science 302 1364-8

34. Q51 Y35 Q51 Y35 Y35 Q169 Q169 Q180 G177 G177 G177 Q180 interface Design of novel H bond network Design X-ray Lukasz Joachimiak

35. Design of new protein functions • Design of new protein-protein interactions • Design of enzymes catalyzing novel chemical reactions • Design of new transcription factor and endonuclease specificities • Design of HIV vaccine

36. HIV vaccine design • Present HIV coat protein epitopes locked into conformation observed in complexes with neutralizing antibodies using designed scaffolds • Preliminary results: designed proteins fold and bind neutralizing antibodies (5nM affinity). One design confirmed crystallographically. Bill Schief in collaboration with Peter Kwong

37. Computational design of non-HIV immunogens to elicit broadly-neutralizing antibodies Bill Schief Crystal structure of Mab 2F5 in complex with its HIV epitope Model of non-HIV scaffold-epitope (red)

38. Design-WT WT-WT Redesign of DNA cleavage specificity of MsoI homing endonuclease using ROSETTA Justin Ashworth, Jim Havranek Nature in press Design-Design WT-Design

39. ½ ¼ - 1 1/2n 1/29 wild-type design Cleavage targets wild-type I-Mso 5uM nuclease - wild-type design Design Specific DNA cleavage by designed nuclease

40. Design Brian Kuhlman (UNC) Gautam Dantas Justin Ashworth Jim Havranek Robetta.bakerlab.org prediction and design server: David Kim (domain parsing, boinc) and Dylan Chivian Rosetta software freely available for academic use Boinc.bakerlab.org/rosetta Protein structure prediction Phil Bradley (MIT) Rhiju Das Lars Marlstrom Bin Qian Vatson Raman Protein-protein docking Ora Furman (Hebrew University) Chu Wang Jeff Gray (Johns Hopkins) Acknowledgements