Biomodeling: DNA-binding proteins (PCNA)
Intro
Download link: https://www.dropbox.com/scl/fo/793kcxhjhj7q7ye5sh51y/h?rlkey=1rrx9y0h1iuzc7suh2q15sfbq&dl=0
In PCNA folder, you can find the structure and XTC file from my simulation.
Intro
There are several well known DNA-binding motif proteins.
In those DNA-binding proteins, basic residues bind to
Backbone phosphates: non-specific (e.g., RNA polymerase).
Major grooves: motif-level, specific and relatively weaker (e.g., zinc finger, helix-turn-helix etc.).
Minor grooves: single residue-level, specific and relatively stronger (e.g., histone, exonuclease etc.).
DNA-binding proteins diffuse along DNA to find the target sites!!!
References
You, S., Lee, H. G., Kim, K. & Yoo, J. Improved Parameterization of Protein-DNA Interactions for Molecular Dynamics Simulations of PCNA Diffusion on DNA. J Chem Theory Comput 16, 4006-4013 (2020). https://doi.org/10.1021/acs.jctc.0c00241
Blainey, P. C. et al. Nonspecifically bound proteins spin while diffusing along DNA. Nat Struct Mol Biol 16, 1224-1229 (2009). https://doi.org/10.1038/nsmb.1716
Kochaniak, A. B. et al. Proliferating cell nuclear antigen uses two distinct modes to move along DNA. J Biol Chem 284, 17700-17710 (2009). https://doi.org/10.1074/jbc.M109.008706
Diffusion of PCNA in MD simulations
In Kochaniak et al., you can find the single-molecule measurements of a PCNA complex as a function of time, y(t).
Using y(t), we can calculate the mean-squared-displacement (MSD) curve.
Questions
Questions
Diffusion coefficient of the protein complex?
Translational vs. rotational?
What is the physical nature of the protein-DNA contacts?
Visual inspection
Radial distribution functions?
What else?
PCNA in MD simulations
Prepare a 20-bp or 21-bp dsDNA under PBC using our web server.
dna.pdb contains the coordinates of DNA.
PCNA is a homotrimeric ring-shaped protein complex.
Download PDB: 5L7C or 6GIS.
Orientation of DNA in 6GIS and our DNA from the web server are different. We want to align DNA of 6GIS parallel to the z-axis.
# Align the principal axis of DNA to the x-axis.
gmx editconf -f 6GIS.pdb -o 6GIS_orient.pdb -princ
# Rotate by 90 degree with respect to the y-axis.
gmx editconf -f 6GIS_orient.pdb -o 6GIS_orient.pdb -rotate 0 90 0
Now, DNA molecules in dna.pdb 6GIS_orient.pdb are parallel to the z-axis.
Update the box information in dna.pdb and 6GIS_orient.pdb using text editor.
CRYST1 115.000 115.000 68.000 90.00 90.00 60.00 P 1 1
Put dna.pdb and 6GIS_orient.pdb to the center of the box we defined.
gmx editconf -f dna.pdb -o dna_c.pdb -c
gmx editconf -f 6GIS_orient.pdb -o 6GIS_c.pdb -c
Confirm if dna_c.pdb and 6GIS_c.pdb are aligned well using VMD.
We need to convert 6GIS_c.pdb for Gromacs using gmx pdb2gmx.
gmx pdb2gmx -f 6GIS_c.pdb -ignh
But, pdb2gmx returns an error saying that some atoms are missing in 6GIS_c.pdb.
This problem often occurs because crystal structure don't have all atomic coordinates in many cases.
There are multiple ways to fill the missing atoms: AF server (https://alphafoldserver.com), CHARMM, modeller, ..., charmm-gui.org
We will use alphafoldserver.com to get a whole complex structure without missing atoms or residues.
Put protein sequence downloaded from RCSB sites in FASTA format.
Generate a trimer structure using AF server.
Load model 0 from AF to VMD.
Make 6GIS_c.pdb as a top molecule in VMD.
VMD->Extensions->Analysis->RMSD caculator.
Align the AF model to 6GIS_c.pdb using the selection "chain A B C and name CA and resid 1 to 100"
Save the aligned AF model structure to AF.pdb and copy AF.pdb to the server.
Redo gmx pdb2gmx using AF.pdb as an input.
gmx pdb2gmx -f AF.pdb -o conf.pdb -ignh
# outputs
# topol.top
# conf.pdb
Put DNA information to topol.top and conf.pdb
In topol.top, put the following lines for DNA.
; Include DNA topologies
#include "dna.itp"
#include "dna.pbc.itp"
#include "dna.hbonds.itp"
At the end of conf.pdb, append dna.pdb.
Modify CRYST1 line in conf.pdb to
CRYST1 115.000 115.000 68.000 90.00 90.00 60.00 P 1 1
Put the PCNA structure to an infinitely long DNA from the web server.
Then, follow a typical MD simulation protocol.
Perform the simulation for a few microseconds.
In the Dropbox folder (PCNA), you can find a trajectory file.
Mean-Squared Displacement (MSD) and Diffusion Coefficient
For a 1D diffusion case, the diffusion coefficient is defined as MSD(∆ t)= 2D∆.t
A Python example.
https://stackoverflow.com/questions/31264591/mean-square-displacement-python
import numpy as np
# calculate COM of PCNA and put the z coordinates to zdata array.
r = np.sqrt(zdata**2)
diff = np.diff(r) #this calculates r(t + dt) - r(t)
diff_sq = diff**2
MSD = np.mean(diff_sq)
Rotation and Tilt angles of PCNA
