Team:TU Eindhoven/Modeling/Script

Modeling Script

Model input

A short description, the input parameters and output data are explained for the four submodels below:

Linker Simulation (Distance)
This model calculates the effect of the linker on the distance between the intracellular domains. It's output file consists of a list of numbers. Each number represents the distance between the intracellular domains that was calculated in one iteration.

Linker Simulation (BRET/FRET)
This model calculates the effect of the linker on the distance between the intracellular domains. Then it uses these calculated data to calculate the mean BRET or FRET efficiency, while maintaining a constant distance between the membrane proteins. It's output file consists of the distance between the membrane proteins that was modeled and the corresponding simulated mean BRET or FRET, separated by a comma.

Total System Simulation
This model simulates the total system. It places an equimolar amount of the membrane proteins with the 2 different intracellular domains on a 2D membrane. Then it calculates the mean BRET or FRET efficiency between these domains. It's output file consists of the amount of membrane proteins modeled and the simulated mean BRET or FRET.

Aptamer Binding Simulation
This model predicts the amount of aptamers bound to their ligand, using the software package Smoldyn. The data in it's output file can be configured in the configuration file of Smoldyn itself. It's output file consists of the amount of particles of each type per 10 time steps, by default.

Custom Model

  • Box Size (nm): the width and height of the simulated 2d membrane, so the total area will be the square. We filled in 188 nm, to obtain an area of 35,344 nm2, which is about one hundredth of the area of an E. coli bacterium.
  • Amount of Particles: the amount of membrane proteins that will be placed on the 2d area. The amount we used was dependent on the situation
  • Radius of Membrane Protein (nm): the radius of the simulated membrane protein. The used membrane protein ompX, has a radius of 2 nm. This was measured by means of its PDB file.
  • Length of Linker (nm): the contour length of the modeled linker in nanometers. The value we used for our simulation was 28.4 nm; the linker consists of 71 amino acids that each have a contour length of 0.4 nm [1]
  • Radii of the tethered particles (nm): the radii of the two intracellular domains that are tethered to their membrane protein. We filled in 3 nm for both our first domain (Nanoluc) and our second (mNeongreen). These values were measured using PDB files. For Nanoluc a PDB was used that represents a model of the real structure. For mNeongreen, a single beta barrel in the PDB file 4HVF was used. This PDB file describes the x-ray diffraction structure of lanGFP, a relative of lanYFP, the protein which was mutated to mNeongreen. [2]
  • Persistence Length of Linker (nm): the persistence length that is characteristic for the used material. The value we used to simulate our peptide linker was 0.45 nm [3]
  • Förster Distance (nm): the förster distance of the used BRET or FRET pair. Because this distance was not known for the used pairs, 5 nm was used for this parameter, which is a typical distance [4]
  • Max Förster Distance (nm): the maximum distance the membrane proteins may be apart from each other to simulate the BRET or FRET between their intracellular domains. This parameter was set to 12 nm. Figure ? shows that intracellular domains of membrane proteins that are further away than 12 nm of each other, hardly contribute to the total BRET or FRET signal.

  • sysradius: Defines the length of the edge of the system cube. The total volume is the cube of the value spacified here. In our model we used this value to define the concentration of thrombin, while keeping the total molecules of thrombin constant
  • memradius: Defines the radius of the sphere that represents the E. coli. The membrane area is simulated with a scale of 1:100. It was set to 0.053 to give the simulated E. coli's membrane area on hundredth of a real E. coli [5]
  • Diffusion constants: The diffusion constants of thrombin and the membrane proteins were configured in the model. The diffusion constant of thrombin was set to 110 µm2/s [6], and the diffusion constant for the membrane proteins was set to 4.5µm2/s, which is an approximation for membrane proteins with an radius of 2 nm in general, not specific for ompX [7].
  • Reaction settings: The association and disassociation constants used for our model were 77450 and 0.00338 [8]. Furthermore the maximum probability of a geminate reaction was set to 99%, because the aptamers are attached to a 2d surface, the membrane [9]
  • Amount of particles: Here the amount of simulated proteins can be set. In our simulations the amount was dependent on the situation. The amount of thrombin was always set to 1000, in order to create an excess

Download model
Click here to download the model. The ".bat" files will work for windows. For Mac or Linux use a custom script to startup java in a command-line.

[1] S. R. K. Ainavarapu, J. Brujic, H. H. Huang, A. P. Wiita, H. Lu, L. Li, K. a Walther, M. Carrion-Vazquez, H. Li, and J. M. Fernandez, “Contour length and refolding rate of a small protein controlled by engineered disulfide bonds.,” Biophys. J., vol. 92, no. 1, pp. 225–233, 2007.
[2] N. C. Shaner, G. G. Lambert, A. Chammas, Y. Ni, P. J. Cranfill, M. a Baird, B. R. Sell, J. R. Allen, R. N. Day, M. Israelsson, M. W. Davidson, and J. Wang, “A bright monomeric green fluorescent protein derived from Branchiostoma lanceolatum.,” Nat. Methods, vol. 10, no. 5, pp. 407–9, 2013.
[3] T. H. Evers, E. M. W. M. Van Dongen, A. C. Faesen, E. W. Meijer, and M. Merkx, “Quantitative understanding of the energy transfer between fluorescent proteins connected via flexible peptide linkers,” Biochemistry, vol. 45, no. 44, pp. 13183–13192, 2006.
[4] A. Manuscript, “Förster distances for FRET between mCherry and other Visible Fluorescent Proteins” Changes, vol. 29, no. 6, pp. 997–1003, 2012.
[6] O. V. Kim, Z. Xu, E. D. Rosen, and M. S. Alber, “Fibrin Networks Regulate Protein Transport during Thrombus Development,” PLoS Comput. Biol., vol. 9, no. 6, 2013.
[7] S. Ramadurai, a Holt, and V. V Krasnikov, “Lateral diffusion of membrane proteins,” J. Am. Chem. Soc., no. 16, pp. 12650–12656, 2009.
[8] S. Goji and J. Matsui, “Direct detection of thrombin binding to 8-bromodeoxyguanosine-modified aptamer: effects of modification on affinity and kinetics.,” J. Nucleic Acids, vol. 2011, p. 316079, 2011.
[9] M. M. Palm, M. N. Steijaert, and P. a J. ten Eikelder, Huub M M Hilbers, “Modeling molecule exchange at membranes,” Proc. Third Int. Conf. Found. Syst. Biol. Eng. Denver, Color., pp. 40–43, 2009.