marcin pacholczyk, silesian university of technology

Biophysics and Thermodynamics Approaches for Modeling and Testing NF-B Binding Sites

Marcin Pacholczyk, Silesian University of Technology

Physics vs Knowledge – based potentials

Physics-based Laws of Physics – electrostatics, van der

Waals, molecular flexibility, geometry of hydrogen bonds

Computationally intensive, some effects difficult to model (desolvation)

Knowledge-based Relatively simple, based on observation Training set!

Physics-Based Potential

e=80

Na+

Cl-

e=2- 4

h

e=80

e=2- 4

C VdWG E E=

Poisson-Boltzmann equation + Lenard-Jones potential

Knowledge-Based Statistical Potential

ln ln , ,P DN N

ij i ji j

G P C D P C d t t- =-

Robertson and Varani 2007

Gibbs energy probability of „correctness”


, ,, ,

, ,

ij i j

ij i j

ij i j

P d t t CP C d t t P C

P d t t=

Probability of individual atomic contact

P(C) – Bayesian prior of observing native-like protein-DNA complex – set to 1.



, ,

, , , ,, ,

ij

obs ij i j

ij i j ij i j

obs ij i jd

N d t tP d t t C f d t t

N d t t =

Probability function

Continous dij is mapped to a set of discrete distance bins b0, b1, … , bn with distance cutoffs db0, db1, … , dbn

A count is assigned to bi if dbi-1 dij < dbi

3 Å, 4 Å, 5 Å, 6 Å, 7 Å, 8 Å, 9 Å, 10 Å



, ,

, , i j

obs ij i jt t

ij i j ijC

N d t t

P d t t f dN

=

Marginal distribution

NC – total number of observed contacts between interface atoms of all types, at all distances in the training set


Training set – Nucleic Acid Database ndbserver.rutgers.edu

Computation of the PWMs

Almanova et al. 2010

Three members of the NF-B family of TF p50p50 homodimer (1NFK) p50RelB heterodimer (2V2T) p50p65 heterodimer (1VKX)

Complexes with DNA fragmentsDNA chains were mutated (MMTSB – Multiscale Modeling Tools for Structural Biology) one base pair at each step (backbone fixed)



Three members of the NF-B family of TF p50p50 homodimer (1NFK) p50RelB heterodimer (2V2T) p50p65 heterodimer (1VKX)

Complexes with DNA fragments (PDB)

NF-B family

p50p50 p50p65

p50RelB


AX b=

DNA chains were mutated (MMTSB – Multiscale Modeling Tools for Structural Biology) one base pair at each step (backbone fixed) 4N + R

All weights w(i, u) in the PWM predicted by solving the linear equation:

X is a vector of 4N dimensions of the estimated weights

A is a binary matrix of dimensions (4N, 4N + R), with all random DNA sequences whose free binding energy was computed.

The free binding energy vector b consists of 4N + R values obtained with the protein-DNA scoring procedure

p50p50

p50RelB

p50p65

TRANSFACV$NFKAPPAB_01

Comparison

Almanova DDNA2 TRANSFAC

p50p50 2.65 -

2.29p50RelB 2.84 -

p50p65 2.38 3.28

Relative entropy


, ,,

logi j i ji j

H p p=-

Comparison

69 human genes regulated by NF-B with 124 promoter sequences (TRANSPRO)

Experimentally confirmed 31 out of 124 promoters belonging to 25 genes

Matrix scan with Match on 58 confirmed binding sites

Almanova TRANSFAC

p50p50 30 (5) 25 V$P50P50_Q3

p50p65 25 (5) 26 (6) V$P50RELAP65_Q5_01

Binding site discovery



Comparison

Comparison

Almanova DDNA2 TRANSFAC

p50p50 0.825 0.833 0.855

p50p65 0.798 0.792 0.863

AUC


Implementation

http://cogangs.biobase.de/3dtf/

Conclusions

Discovery of novel NF-B binding sites Investigation of postranslational modifications like RelA Ser276 phosphorylation (Nowak et al. 2008)

It is possible to compute PWMs which perform comparably to the ones derived from experimental data (TRANSFAC)

Thermodynamic based models of transcriptional regulation including Synergistic Activation, Cooperative Binding and Short-Range Repression (He et al. 2010)

marcin pacholczyk, silesian university of technology

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