Applications of Homology Modeling Hanka Venselaar

This seminar…. Homology Modeling… Why? What? When? How? And a few real world examples….

Hearing loss No structure: MGTPWRKRKGIAGPGLPDLSCALVLQPRAQVGTMSPAI ALAFLPLVVTLLVRYRHYFRLLVRTVLLRSLRDCLSGLRI EERAFSYVLTHALPGDPGHILTTLDHWSSRCEYLSHMG PVKGQILMRLVEEKAPACVLELGTYCGYSTLLIARALPP GGRLLTVERDPRTAAVAEKLIRLAGFDEHMVELIVGSSE DVIPCLRTQYQLSRADLVLLAHRPRCYLRDLQLLEAHAL LPAGATVLADHVLFPGAPRFLQYAKSCGRYRCRLHHTG LPDFPAIKDGIAQLTYAGPG DFNB 63 Sequence:

Homology modeling in short… Prediction of structure based upon a highly similar structure 2 basic assumptions: Structure defines function During evolution structures are more conserved than sequence Use one structure to predict another

Homology modeling Example: by 80 residues  30% identity sufficient # residues % identity * * Actually, modelling is possible, but we cannot get an alignment… O

Homology modeling in short… Prediction of structure based upon a highly similar structure Add sidechains, Molecular Dynamics simulation on model Unknown structure NSDSECPLSHDG || || | || NSYPGCPSSYDG Alignment of model and template sequence Known structure Back bone copied Copy backbone and conserved residues Model!

The 8 steps of Homology modeling

1: Template recognition and initial alignment

BLAST your sequence against PDB Best hit  normally template Initial alignment  NSDSECPLSHDGYCLHDGVC || || | ||||| ||| NSYPGCPSSYDGYCLNGGVC

1: Template recognition and initial alignment 2: Alignment correction

Functional residues  conserved Use multiple sequence alignments Deletions  shift gaps CPISRTGASIFRCW CPISRTA---FRCW CPISRT---AFRCW CPISRTAAS-FRCW CPISRTG-SMFRCW CPISRTA--TFRCW CPISRTAASHFRCW CPISRTGASIFRCW CPISRTA---FRCW Both are possible Multipe sequence alignment Correct alignment  Sequence with known structure  Your sequence

2: Alignment correction Core residues  conserved Use multiple sequence alignments Deletions in your sequence  shift gaps Known structure FDICRLPGSAEAV Model FNVCRMP---EAI Model FNVCR---MPEAI S G P L A E R C IV C R M P E V C R M P E  Correct alignment F-D- -A-V

1: Template recognition and initial alignment 2: Alignment correction 3: Backbone generation

Making the model…. Copy backbone of template to model Make deletions as discussed (Keep conserved residues)

1: Template recognition and initial alignment 2: Alignment correction 3: Backbone generation 4: Loop modeling

Known structure GVCMYIEA---LDKYACNC Your sequence GECFMVKDLSNPSRYLCKC Loop library, try different options

1: Template recognition and initial alignment 2: Alignment correction 3: Backbone generation 4: Loop modeling 5: Sidechain modeling

5: Side-chain modeling Several options Libraries of preferred rotamers based upon backbone conformation

1: Template recognition and initial alignment 2: Alignment correction 3: Backbone generation 4: Loop modeling 5: Sidechain modeling 6: Model optimization

Molecular dynamics simulation Remove big errors Structure moves to lowest energy conformation

1: Template recognition and initial alignment 2: Alignment correction 3: Backbone generation 4: Loop modeling 5: Sidechain modeling 6: Model optimization 7: Model validation

7: Model Validation Second opinion by PDBreport /WHATIF Errors in active site?  new alignment/ template No errors?  Model!

1: Template recognition and initial alignment 2: Alignment correction 3: Backbone generation 4: Loop modeling 5: Sidechain modeling 6: Model optimization 7: Model validation 8: Iteration

Model! 1: Template recognition and initial alignment 2: Alignment correction 3: Backbone generation 4: Loop modeling 5: Sidechain modeling 6: Model optimization 7: Model validation 8: Iteration

8 steps of homology modeling 1: Template recognition and initial alignment 2: Alignment correction 3: Backbone generation 4: Loop modeling 5: Side-chain modeling 6: Model optimization 7: Model validation 8: Iteration Alignment Modeling Correction

Hearing loss Structure! MGTPWRKRKGIAGPGLPDLSCALVLQPRAQVGTMSPAI ALAFLPLVVTLLVRYRHYFRLLVRTVLLRSLRDCLSGLRI EERAFSYVLTHALPGDPGHILTTLDHWSSRCEYLSHMG PVKGQILMRLVEEKAPACVLELGTYCGYSTLLIARALPP GGRLLTVERDPRTAAVAEKLIRLAGFDEHMVELIVGSSE DVIPCLRTQYQLSRADLVLLAHRPRCYLRDLQLLEAHAL LPAGATVLADHVLFPGAPRFLQYAKSCGRYRCRLHHTG LPDFPAIKDGIAQLTYAGPG DFNB 63 Sequence:

Saltbridge between Arginine and Glutamic acid is lost in both cases Arginine 81 -> Glutamic acid Glutamic acid 110 -> Lysine Mutations:

Mutation: Tryptophan 105 -> Arginine Hydrophobic contacts from the Tryoptohan are lost, introduction of an hydrophilic and charged residue

The three mutated residues are all important for the correct positioning of Tyrosine 111 Tyrosine 111 is important for substrate binding Accepted in Nature Genetics

Homology Modeling… What? Prediction of an unknown structure based on an homologous and known structure Why? To answer biological and medical questions when the “real” structure is unknown When? A template with enough identity must be available How? 8 Steps Real world examples: mutations in DFNB63 gene can lead to hearing disorders. To conclude….

And now…. Go to the course website: Follow the steps on the site A few hints…. Login with username: c01 – c13 (number of your pc is written on the pc itself) Click on the icon to open a terminal Run Yasara by typing “yasara” in that terminal Save your files on the Desktop