1. Protein Structure Modelling Many sequences - few structures Homology Modelling - Based on Sequence Similarity with Sequences of Known Structures.

3. COMPLEXES

4. Protein:DNA Complex

5. Initiation of Transcription.

7. Structures of Macromolecular Complexes Very few experimentally determined –e.g. antibodies:antigens ~1000 antibody sequences known ~100 antibody structures known ~10 antibody:antigen complex structures known Can we use calculation?

8. Structures of Isolated Components? - crystallography - NMR - Homology Modelling Structure of Complex? Rigid-Body Shape Complementarity (based on hydrophobic effect and van der Waals packing) Conformational Change on Complexation? Electrostatic Complementarity? Solvation Effects? Experiment?

9. Complementarity Determining Region Loops (CDRs): (i) Uniform Conformational Searching (ii) Canonical Loop Modelling (iii) Data-Base Searching of Loop Conformations (iv) Molecular Dynamics in vacuo and with solvated CDRs. > 90 models. Clustering and Screening for Consistency with Experimental Antibody Structures. 4 Dynamically Interconvertible Models. Homology Model of Framework Residues. Modelling of Isolated Antibody

10. Clustering and Screening for: (i) Buried Surface Area. (ii) Electrostatic Complementarity. (iii) Consistency with existing Ab:Ag complex structures. > 18 models. Refinement of Atomic-Detail Models with Molecular Dynamics in Explicit Solvent. 6 Models. Initial Generation Low -Resolution Shape Complementarity. > 41,585 models Modelling of Ab:Ag Complex

11. Which Experiments can help Model the Structures of Complexes ? Site-Directed Mutagenesis –Mutate then measure affinity. –Swap Mutations e.g. Glu:Lys for Lys:Glu Optical Spectroscopy. –e.g. Fluorescence energy transfer gives direct information on distances between sites.

12. Functional Binding Site on Toxin  Red: Affinity Lowered >100-fold Yellow: Affinity Lowered 10-100 fold

14. Three Models of Calculated M  23 Paratope Red: Residues contacting antigen energy core Yellow: Residues contacting functional epitope