DNA/Protein%20structure-function%20analysis%20and%20prediction

1
DNA/Protein structure-function analysis and
prediction

• Protein-protein Interaction (PPI)
• Protein-protein Interaction
• Interfaces
• Solvation
• Energetics
• Conformational change
• Allostery

• Prediction
• Gene Cluster
• Phylogenetic Profile
• Rosetta Stone
• Sequence co-evolution
• Random Decision forest
• Docking
• Examples

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PPI Characteristics

• Universal
• Cell functionality based on protein-protein
  interactions
• Cyto-skeleton
• Ribosome
• RNA polymerase
• Numerous
• Yeast
• 6.000 proteins
• at least 3 interactions each
• 18.000 interactions
• Human
• estimated 100.000 interactions
• Network
• simplest homodimer (two)?
• common hetero-oligomer (more)?
• holistic protein network (all)?

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Interface Area

• Contact area
• usually gt1100 Å2
• each partner gt550 Å2
• each partner loses 800 Å2 of solvent accessible
  surface area
• 20 amino acids lose 40 Å2
• 100-200 J per Å2
• Average buried accessible surface area
• 12 for dimers
• 17 for trimers
• 21 for tetramers
• 83-84 of all interfaces are flat
• Secondary structure
• 50 ?-helix
• 20 ?-sheet
• 20 coil
• 10 mixed
• Less hydrophobic than core, more hydrophobic than
  exterior

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Complexation Reaction

• A B ? AB
• Ka AB/AB ? association
• Kd AB/AB ? dissociation
• Free energy
• DGd -RTln Kd
• Kd exp(-DGd / RT)?
• (R 8.3144 J mol-1 K-1 )?

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Experimental Methods

• 2D (poly-acrylamide) gel electrophoresis ? mass
  spectrometry
• Liquid chromatography
• e.g. gel permeation chromatography
• Binding study with one immobilized partner
• e.g. surface plasmon resonance
• In vivo by two-hybrid systems or FRET
• Binding constants by ultra-centrifugation,
  micro-calorimetry or competition
• experiments with labelled ligand
• e.g. fluorescence, radioactivity
• Role of individual amino acids by site directed
  mutagenesis
• Structural studies
• e.g. NMR or X-ray

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PPI Network
http//www.phy.auckland.ac.nz/staff/prw/biocomplex
ity/protein_network.htm
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Protein-protein interactions

• Complexity
• Multibody interaction
• Diversity
• Various interaction types
• Specificity
• Complementarity in shape and binding properties

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Binding vs. Localization
strong
Non-obligatetriggered transient e.g. GTPPO4-
Non-obligatepermanente.g. antibody-antigen
Obligateoligomers
Non-obligateco-localised e.g. in membrane
Non-obligateweak transient
weak
co-expressed
different places
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Some terminology

• Transient interactions
• Associate and dissociate in vivo
• Weak transient
• dynamic oligomeric equilibrium
• Strong transient
• require a molecular trigger to shift the
  equilibrium
• Obligate PPI
• protomers not stable structures on their own
• (functionally obligate)?

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Strong medium weak

• Nanomolar to sub-nanomolar
• Kd lt 10-9
• Micromolar to nanomolar
• 10-6 gt Kd gt 10-9
• Micromolar
• 10-3 gt Kd gt 10-6
• A B ? AB
• Kd AB/AB ? dissociation

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Analysis of 122 Homodimers

• 70 interfaces single patched
• 35 have two patches
• 17 have three or more

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Patches

• Cluster in different domains
• (structurally defined units often with specific
  function)?

two domains
anticodon-binding
catalytic
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Interfaces

• 30 polar
• 70 non-polar

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Interface

• Rim is water accessible

rim
core
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Interface composition

• Composition of interface essentially the same as
  core
• But surface area can be quite different!

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Propensities

• Interface vs. surface propensities
• as ln(fint/fsurf)?

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Conformational Change

• Chaperones
• extreme conformational changes upon complexation
• ligand unfolds within the chaperone GroEL/GroES
• Allosteric proteins
• conformational change at 'active' site
• ligand binds to 'regulating' site
• Peptides
• often adopt 'bound' conformation
• different from the 'free' conformation

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Allostery 1

• Regulation by 'remote' modulation of binding
  affinity (complex strength)?

www.blc.arizona.edu/courses/181gh/rick/energy/allo
stery.html
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Allostery 2

• Substrate binding is cooperative
• Binding of first substrate at first active site
• stimulates active shape
• promotes binding of second substrate

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Allostery 3

• Committed step of metabolic pathway
• regulated by an allosteric enzyme
• Pathway end product
• can regulate the allosteric enzyme for the first
  committed step
• Inhibitor binding favors inactive form

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DNA/Protein structure-function analysis and
prediction

• Protein-protein Interaction (PPI)
• Protein-protein Interaction
• Interfaces
• Solvation
• Energetics
• Conformational change
• Allostery

• Prediction
• Gene Cluster
• Phylogenetic Profile
• Rosetta Stone
• Sequence co-evolution
• Random Decision forest
• Docking
• Examples

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Predicting Protein-Protein Interactions

• Gene Cluster
• Gene neighborhood
• Phylogenetic Profile
• Co-occurrence across species/genomes
• Rosetta Stone
• Occurrence of protein with domains linked
• Sequence co-evolution
• Tree correlation indicated functional relation
• Random Decision forest
• Using data on domain interactions
• Shoemaker Panchenko, PLOS-CB 2007 3 e43

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Gene Cluster / Neighborhood
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Gene Cluster / Neighborhood

• Genes with closely related functions encoding
  potentially interacting proteins
• transcribed as a single unit (operon) in bacteria
  
• co-regulated in eukaryotes
• Operons can be predicted from intergenic distance
• Neutral evolution tends to shuffle gene order
  between distantly related organisms
• but gene clusters or operons that encode
  co-regulated genes are usually conserved
• operons found by gene neighbor methods provide
  additional evidence about functional linkage

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Phylogenetic Profile
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Phylogenetic Profile

• hypothesis that functionally linked and
  potentially interacting nonhomologous proteins
  co-evolve and have orthologs in the same subset
  of organisms
• components of complexes and pathways should be
  present simultaneously in order to perform their
  functions.
• phylogenetic profile is a vector of N elements
  (number of genomes)
• presence/absence of protein in genome is 1 or
  0 at each position of a profile.
• clustered using bit-distance measure
• proteins in a cluster are considered functionally
  related.
• also for protein domains instead of entire
  proteins

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Rosetta Stone
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Rosetta Stone

• infer protein interactions from sequences in
  different genomes
• some interacting proteins/domains have homologs
  that are fused into one protein
• a so-called Rosetta Stone protein
• Apparently, gene fusion can occur to optimize
  co-expression of genes encoding for interacting
  proteins.

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Sequence co-evolution
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Sequence co-evolution

• interacting proteins often co-evolve so changes
  in one protein leading to the loss of function or
  interaction can be compensated by changes in the
  other
• orthologs of coevolving proteins also tend to
  interact
• infer unknown interactions in other genomes
• similarity between phylogenetic trees of two
  non-homologous interacting protein families
• correlation coefficient between the distance
  matrices
• requires correspondence between the matrix
  elements / tree branches (i.e. ortholog
  relations)?
• align distance matrices to minimize difference
• predicted interactions correspond to aligned
  cols
• max. 30 proteins in a family

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Classification / Random Decision Forest
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Random Forest Decision

• Decision trees based on domains of interacting
  and non-interacting proteins
• All possible combinations of interacting domains
• vector of length N (different domain types or
  features)?
• 2, 1, or 0 found in both, one, or no protein of
  pair
• experimental training set of interacting protein
  pairs
• decision tree (many trees)
• defines the best splitting feature at each node
• from a randomly selected feature subspace
• best feature is selected based on goodness of
  fit,
• can discriminate interacting and non-interacting
• stops growing the tree when all pairs at a given
  node are well-separated
• Traverse the tree to classify an unknown protein
  pair

33
DNA/Protein structure-function analysis and
prediction

• Protein-protein Interaction (PPI) and Docking
• Protein-protein Interaction
• Interfaces
• Solvation
• Energetics
• Conformational change
• Allostery

• Prediction
• Docking
• Search space
• Docking methods
• Examples

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Docking - ZDOCK

• Protein-protein docking
• 3-dimensional (3D) structure of protein complex
• starting from 3D structures of receptor and
  ligand
• Rigid-body docking algorithm (ZDOCK)
• pairwise shape complementarity function
• all possible binding modes
• using Fast Fourier Transform algorithm
• Refinement algorithm (RDOCK)?
• top 2000 predicted structures
• three-stage energy minimization
• electrostatic and desolvation energies
• molecular mechanical software (CHARMM)?
• statistical energy method (Atomic Contact
  Energy)?
• 49 non-redundant unbound test cases
• near-native structure (lt2.5Å) for 37 test cases
• for 49 within top 4

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Protein-protein docking

• Finding correct surface match
• Systematic search
• 2 times 3D space!
• Define functions
• 1 on surface
• ? or ? inside
• 0 outside

?
?
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Protein-protein docking

• Correlation function
• C????? 1/N3 ?o ?p ?q exp2?i(o? p? q?)/N
  Co,p,q

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Docking Programs

• ZDOCK, RDOCK
• AutoDock
• Bielefeld Protein Docking
• DOCK
• DOT
• FTDock, RPScore and MultiDock
• GRAMM
• Hex 3.0
• ICM Protein-Protein docking
• KORDO
• MolFit
• MPI Protein Docking
• Nussinov-Wolfson Structural Bioinformatics Group