Presentation homolgy modeling
- 2. Homology Modeling Process Template recognition Alignment Determining structurally conserved regions Backbone generation Building loops or variable regions Conformational search for side chains Refinement of structure Validating structures
- 3. Template Recognition First we search the related proteins sequence(templates) to the target sequence in any structural database of proteins The accuracy of model depends on the selection of proper template FASTA and BLAST from EMBL-EBI and NCBI can be used This gives a probable set of templates but the final one is not yet decided After intial aligments and finding structurally conserved regions among templates, we choose the final template
- 4. Alignment in Homology Modeling Sequence alignment is central technique in homology modeling Used in determining which areas of the reference proteins are conserved in sequence Hence suggesting where the reference proteins may also be structurally conserved After SCRs are found,it is used to establish one to one correspondence between the amino acids of reference proteins and the target in SCRs Thus providing basis of the transforming of coordinates from the reference to the model
- 5. Comparison Matrix Between the Two Sequences
- 6. Final Maximum Pathway and Corresponding Sequence Alignment
- 7. Main Heuristic Approaches Progressive multiple Alignment – example - ClustalW and ClustalX Iterative Multiple Alignment – example SAGA
- 8. Alignment of Target Protein with SCR After doing alignments and finding SCRs We align the unknown sequence with the aligned reference proteins with the knowledge of SCRs Since SCR cant contain insertions and deletions The enhancement of standard alignmnet algorithm is used After finding the suitable template and aligning the unknown sequence with the template Assignment of coordinates within conserved regions is done
- 9. Alignment of Model sequence with Reference sequences having SCRs
- 10. Mapping the Pathway Through the Matrix
- 11. Assignment of coordinates within conserved region Once the correspondence between amino acids in the reference and model sequences has been made, the coordinates for an SCR can be assigned The reference proteins' coordinates are used as a basis for this assignment Where the side chains of the reference and model proteins are the same at corresponding locations along the sequence, all the coordinates for the amino acid are transferred Where they differ, the backbone coordinates are transferred , but the side chain atoms are automatically replaced to preserve the model protein's residue types
- 12. Assignment of coordinates in loop or variable region Two main methods Finding similar peptide segments in other proteins Generating a segment de-novo Finding similar peptide segment in other proteins Advantage: all loops found are guaranteed to have reasonable internal geometries and conformations Disadvantage: may not fit properly into the given model protein’s framework In this case, de-novo method is advisable
- 13. Selection Of Loops Check the loops on the basis of steric overlaps A specified degree of overlap can be tolerated Check the atoms within the loop agains each other Then check loop atoms against rest of the protein’s atoms
- 14. Side Chain Conformation Search With bond lengths, bond angles and two rotable backbone bonds per residue φ and ψ, its very difficult to find the best conformation of a side chain In addition, side chains of many residues have one or more degree of freedom. Hence Side chain conformational search in loop regions is must Side chain residues replaced during coordinate transformations should also be checked
- 15. Selection Of Good Rotamer Fortunately, statstical studies show side chain adopt only a small number of many possible conformations The correct rotamer of a particular residue is mainly determined by local environment Side chain generally adopt conformations where they are closely packed
- 16. Optimisation Approaches Energy Minimisation is used to produce a chemically and conformationally reasonable model protein structure Two mainly used optimisation algorithms are Steepest Descent Conjugate Gradients Molecular Dynamics is used to explore the conformational space a molecule could visit
- 17. Model Validation Every homology model contains errors.Two main reasons % sequence identity between reference and model The number of errors in templates Hence it is essential to check the correctness of overall fold/ structure, errors of localized regions and stereochemical parameters: bond lengths, angles, geometries