Modeling DNA unzipping in the presence of bound proteins

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Information about Modeling DNA unzipping in the presence of bound proteins

Published on March 18, 2008

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Modeling DNA unzipping in the presence of DNA binding proteins Farhat Habib, Dr. Ralf Bundschuh Department of Physics, The Ohio State University

Overview Importance of understanding DNA-protein interactions Single molecule experimental techniques The problem statement Theory and description of the applied model Results and conclusions Future directions

Importance of understanding DNA-protein interactions

Single molecule experimental techniques

The problem statement

Theory and description of the applied model

Results and conclusions

Future directions

DNA Carries genetic information Double stranded polymer consisting of monomer units called nucleotides 4 nucleotides labeled A, G, C, and T Basepairing A ≡T, G≡C Each strand carries complete information of the other

Carries genetic information

Double stranded polymer consisting of monomer units called nucleotides

4 nucleotides labeled A, G, C, and T

Basepairing A ≡T, G≡C

Each strand carries complete information of the other

Proteins Most diverse of macromolecules Intermediaries in most biological reactions Composed of smaller units called amino acids Proteins play a vital role in DNA replication, transcription, recombination, repair, and in activating/inhibiting gene expression

Most diverse of macromolecules

Intermediaries in most biological reactions

Composed of smaller units called amino acids

Unzipping force analysis of protein association (UFAPA) Single molecule experiment Right sensitivity for probing DNA-protein interactions F ~ 10 – 20 pN Distances ~ nm

Single molecule experiment

Right sensitivity for probing DNA-protein interactions

F ~ 10 – 20 pN

Distances ~ nm

Goals To investigate the limitations of UFAPA The minimum binding energy for which the protein can be detected Minimum distance between two proteins for which they can be resolved

To investigate the limitations of UFAPA

The minimum binding energy for which the protein can be detected

Minimum distance between two proteins for which they can be resolved

Theory and methods We describe the protein-DNA system’s thermodynamic behavior or properties in terms of the partition function of the system Model Break the DNA-protein system into two parts The double stranded (ds)DNA with (or without) proteins The single stranded (ss)DNA on which force is being applied

We describe the protein-DNA system’s thermodynamic behavior or properties in terms of the partition function of the system

Model

Break the DNA-protein system into two parts

The double stranded (ds)DNA with (or without) proteins

The single stranded (ss)DNA on which force is being applied

Model Partition function for dsDNA The ssDNA In the highly stretched regime we will operate the Extended Freely Jointed Chain (EFJC) model is the most accurate one Where E(i) is the stacking energy of the i th basepair

Partition function for dsDNA

The ssDNA

In the highly stretched regime we will operate the Extended Freely Jointed Chain (EFJC) model is the most accurate one

Model (cont.) The protein Include protein-DNA interaction by adding the extra free energy due to the presence of the protein at the binding site Partition function for the entire system where m 0 is the protein binding site To obtain force at a given extension once we have the partition function, we use

The protein

Include protein-DNA interaction by adding the extra free energy due to the presence of the protein at the binding site

Partition function for the entire system

where m 0 is the protein binding site

To obtain force at a given extension once we have the partition function, we use

Minimum protein strength Top plot shows the force-extension curves from a protein of progressively lower binding energy at same position Average force = 15.3 pN; Std deviation = 0.7 pN At less than 10 kJ/mol the peak from the protein is within one standard deviation of the mean

Top plot shows the force-extension curves from a protein of progressively lower binding energy at same position

Average force = 15.3 pN; Std deviation = 0.7 pN

At less than 10 kJ/mol the peak from the protein is within one standard deviation of the mean

Minimum resolvable distance between two proteins 1400 1500 1700 1800 50 40 20 10 12 14 16 18 20 22 30 1600 R(nm) Protein separation (bp) Force (pN)

Averaged minimum resolvable distance Minimum resolvable distance for pairs of proteins with 3 different relative binding energies

Minimum resolvable distance for pairs of proteins with 3 different relative binding energies

Conclusions and Future Directions We investigate the limits of the UFAPA technique by considering the protein-DNA system thermodynamically Average force for bare DNA was found to be 15.3 pN with a standard deviation of 0.7 pN Minimum binding energy for a protein to be detected using UFAPA in the absence of FEC of bare DNA is around 10 kJ/mol Minimum resolvable distance between two proteins can be up to 50 basepairs depending on relative protein strengths and the underlying DNA sequence Future: Consider the kinetics of the process

We investigate the limits of the UFAPA technique by considering the protein-DNA system thermodynamically

Average force for bare DNA was found to be 15.3 pN with a standard deviation of 0.7 pN

Minimum binding energy for a protein to be detected using UFAPA in the absence of FEC of bare DNA is around 10 kJ/mol

Minimum resolvable distance between two proteins can be up to 50 basepairs depending on relative protein strengths and the underlying DNA sequence

Future:

Consider the kinetics of the process

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