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A NUMERICAL MODEL FORELECTROPORATION IN BACTERIA
J.L. Moran, P.A. Garcia, N.N. Dingari, C.R. BuieDepartment of Mechanical EngineeringMassachusetts Institute of Technology
October 8, 2015
Electroporation
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J.C. Neu, W. Krassowska, Phys. Rev. E, 1999.
Hydrophilic Lipid heads
Hydrophobic pores
Hydrophobic tails
Hydrophilic pores
~5 nm
~5 nm
Schwan 1957, Tsong, Biophysical Journal, 1990.
1.5 cosmV ER θ=
1 VmV <
1 VmV ≥
Electroporation is governed by the Transmembrane Voltage (TMV)
2
Our Focus: Transformation of Bacteria
• Electroporation protocols exist for a tiny fraction of bacteria on earth
• Goal: understand the principles governing successful DNA transfer into bacteria
• Here: COMSOL model to understand effect of bacterial physiology on electroporation
Microbial Fuel Cell SchematicCredit: Logan, B. Nature Rev. Microbio. 2009
Deepwater Horizon Oil SpillCredit: Getty Images
3
Bacterial Physiology
www.daviddarling.info/encyclopedia/P/pilus.html
micro.magnet.fsu.edu/cells/procaryotes/images/procaryote.jpg
Pili on the surface of an Escherichia coli bacterium
• Outside the (inner) plasma membrane, most bacteria exhibit a “soft layer” consisting of (e.g.) fimbriae, sex pili, capsules, flagella, etc.
• The soft layer generally carries a net charge• Charge distribution in the soft layer affects polarizability (Dingari &
Buie, 2014) 4
• Electric potential inside/outside cell (σ = conductivity)
• Transmembrane voltage (TMV)
• Pore creation/destruction rate
• Pore radius evolution
• Membrane current density
( )β πγ πσ
= + − + + +
42max * 14 2 2
1 /m
effh t
V F rdr D rdt kT r r r r r
Electrical energy Steric repulsion of lipid heads Line tension
Effective membrane/liquid interfacial tension
Governing Equations
σ ε ε φ∂ ∇ ⋅ + ∇ = ∂ 0 0r t
( )φ φ≡ −m inside soft membraneV
( ) ( )α
= −
2exp / 1m ep
eq m
dN NV Vdt N V
Pore density [#/cm2]
Characteristic voltage for electroporation
Capacitive chargingProtein channels Current through electropores
( ) ( )σ πσ ∂= + +
∂
2,0 m p pm m m
m
V N t r AV VJ t Ch h t
DeBruin & Krassowska, Biophys. J. 1999 (Part I)Pucihar, Miklavcic & Kotnik, IEEE Trans. Biomed. Eng. 2009
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Numerical Implementation• COMSOL Multiphysics • 2D axisymmetric geometry• 36,353 mesh elements• Time-dependent solve (through
pulse duration)
SEM of E. coli (Wikimedia commons)
2 µm
12.5 kV/cmE =
0.5 µm
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Imposed Conductivity Profile in Soft Layer
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Induced Transmembrane VoltageNo Soft Layer 100-nm Soft Layer
All data shown at t = 1 ms (pulse truncated)Insets show TMV vs. time at positive-facing pole
E ( )φ φ≡ −m inside soft membraneV
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Pore Density vs. Position
All data shown at t = 1 ms (pulse truncated)Insets show mesh used for each case
No Soft Layer 100-nm Soft Layer
E
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Pore Radius vs. Position
All data shown at t = 1 ms (pulse truncated)Insets show mesh used for each case
No Soft Layer 100-nm Soft Layer
E
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Total Pores, Average Radius
membraneavg
membrane
NrdAr
NdA≡
∫∫
∫∫pores
membrane
N NdA= ∫∫
All data shown at t = 1 ms (pulse truncated)
Both the number and the size of pores created depend more strongly on buffer concentration when the soft layer is present
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Conclusions• The presence of a soft layer tends to amplify the
effect of varying background conditions (e.g. conductivity) on the pore size and number
• This work elucidates the effect of buffer concentration on bacterial electroporation
Ongoing Work• Enhance model of soft-layer transport to include
• Dissociation of pH-dependent ionogenic groups• Donnan potential
• Explore correlation between electroporation amenability and cell envelope properties (e.g. polarizability)
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Thank you
13
Simulation Details
• Customized Mesh
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