KcsA Stability Analysis

Author: Julietta Sophia Mendivelso

This project presents a simple stability analysis of the KcsA membrane protein when is embedded in a POPC membrane.

Motivation

The KcsA (K channel of streptomyces A) membrane protein, an archetypal potassium channel with very interesting properties, was the first ion channel to be characterized using X-ray crystallography. The channel itself is a tetramer composed of four identical, single-domain subunits.

KcsA belongs to a family of channels found in almost all organisms and it is one of the most studied ion channels to this day. Research on KcsA has provided important insight on the molecular basis for K+ ion; as a result, KcsA is a template for research on K+ channel function and its elucidated structure underlies computational modeling of channel dynamics for both prokaryotic and eukaryotic species.

Studying KcsA when interacting with a lipid membrane results relevant to the understanding of important structural and mechanistic properties of the $K^+$ channel function. Here, KcsA was embedded in atomistic lipid bilayer of POPC (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) and atomistic molecular dynamics (MD) simulations were conducted to determine the structural integrity of the structure over simulations. The output of the simulations is the data analysed in this project. RMSD and RMSF analyses were conducted, as well as a moving average analysis of the pressure trace along the simulation.

Methods

The tertiary structure of KcsA was obtained using X-ray crystallography. The data was deposited as a .pdb file in the Protein Data Bank and its freely accessible on the Internet.

The .pdb file contains a monomer of the KcsA protein and everything else solved from the crystal structure. The tetramer was built and embedded in the POPC membrane following the steps on the Membrane Protein Tutorial, using the VMD software.

The simulations for minimization, equilibration and production runs of the membrane-protein system were performed using NAMD, a parallel molecular dynamics code designed for high-performance simulation of large biomolecular systems.

The file with the pressure values during the simulation was obtained using the NAMD plot plugin for VMD.

Data

• The Protein Database file for KcsA is available here
• The output files of the simulation are public and available in the Required tutorial files for the Membrane Proteins Tutorial (Advanced).

Requirements

Libraries

The following packages are necessary to run the scripts:

• reshape2
• tidyverse
• zoo
• ggplot2
• bio3d

Files

It is necessary to download the tutorial files to obtain the trajectory (.dcd) file if you do not have the necessary software to run build the tetramer and run the simulations. 510MB of storage are needed.

In the Tutorials website, search the Membrane Protein Tutorial (Advanced) entry and click on Required tutorial files: full version, [.tar.gz, 508M]. Extract the 03-MINEQ/example-output/kcsa_popcwieq-04.dcd file and save it in the data/raw directory in this repository.

Project structure

Project/
*    ├── data/
     │  ├── raw
        │   ├── pressure.dat
        │   ├── kcsa_popcwi.pdb
     |  └── processed
            │   ├── pressure_trace.cvs
*    ├── docs/
        ├── Final_Project_Report.pdf
        ├── Final_project.tex
        ├── references.bib
*    ├── output/
        ├── figs/
*    ├── R/
     │   ├── 01_package_installation.R
     │   ├── 02_data_manipulation.R
     │   ├── 03_rmsd.R
     │   ├── 04_rmsf.R
     │   ├── 05_pressure_trace.R
*    └── README.md

• In the data/raw directory you will find the .pdb file for the KcsA and the pressure values measured during the simulation.

• In the R directory you will find all the necessary scripts to run the project:

  • 01_package_installation.R install the necessary packages.
  • 02_data_manipulation.R clean the data for the pressure stability analysis.
  • 03_rmsd.R calculate and plot the Root Mean Square Deviations during the simulation.
  • 04_rmsf.R calculate and plot the Root Mean Square Fluctuations during the simulation.
  • 05_pressure_trace.R Calculate and plot the pressure and its moving average during the simulation.

• In the data/processed you will find the pressure.dat file processed in the script 02_data_manipulation.R

• In the docs directory you will find a pdf version of report of the project, and a .tex version with its respective bibliography (.bib) file and the images loaded in the .tex file.

• In the output/figs directory you will find the output plots from each script.