Sticky Proteins Rogues in the Network

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Sticky ProteinsRogues in the Network

• Ramazan Saeed
• 25th August 2004

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Outline

• Protein interaction networks
• Methods for measuring interactions
• Y2H
• TAP Tagging
• False positives
• Evolutionary rate and interactions
• Work in progress

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Protein Interaction Networks

• Interactions reveal the inner working of the cell
• Help in determining biological pathways
• Select protein targets for therapeutic
  intervention
• Predict side effects
• Model the cell

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Interactomes

• Low throughput and High throughput methods
• Current Interactomes
• D. melanogaster, S. cerevisiae, C. elegans, H.
  pylori, H. sapiens, E. coli, M. musculus
• Several Databases exist
• DIP, BIND, IntAct, MINT
• Interactions can be measure and predicted

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Yeast two-Hybrid

• Uses Gal4, a transcription factor
• It has two domains(AD and BD)
• Protein of interest fused with BD
• Potential interacting partner fuses with AD
• Transcription of reporter gene is measured

http//www.bioteach.ubc.ca/MolecularBiology/AYeast
TwoHybridAssay/
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Tandem-affinity purification

• Bait protein is tagged
• Put through a column
• First tag is removed
• Put through another column
• Products are separated
• Identified by mass spectrometry

http//www.nature.com/nrm/journal/v4/n1/slideshow/
nrm1007_F1.html
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False Positives

• Time and Space
• Sticky Proteins
• Bridging Proteins (TAP)
• Auto-activation (Y2H)
• Ambiguity of MS

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Evolutionary Distance
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Structural Folds
D
A
B
C
G
E
F
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Work in progress (20/8)

• Complete Structural Analysis
• Functional Categories
• Overall Bias
• Study potentially sticky proteins