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2017-07-04
1
Neutron Scattering for Soft Matter- Lipid Membranes Interacting with Proteins
- Binary Superlattices of 1D Colloids
Sung-Min Choi
Neutron Scattering and Nanoscale Materials Lab.Dept. Nuclear and Quantum Engineering
KAIST([email protected])
• Basic Concepts of Neutron Scattering• Lipid Membranes Interacting with Proteins• Binary 1D Colloidal Superlattices
Neutrons for Soft Matter
Spherical Micelle
Rod-likemicelle
LamellarVesicles
2-4 nmin Water
2017-07-04
2
Infrared UVCold Neutron
soft x-ray x-rayNeutron
Microwave
Electron Microscopy (destructive)STM (surface)
In-situ analysis : Neutron & X-rayWe need
Measurement Techniques
1 nanometer1cm 1mm 1m
10-2m 10-3m 10-4m 10-5m 10-6m 10-7m 10-8m 10-9m 10-10m
Natural
Protein
Manmade
Nanotube electronics
What Neutrons See
C. G. Shull B. N. Brockhouse
2017-07-04
3
Length scale (Å)
Tim
e sc
ale
(sec
)
Dynamics : femto sec- 100 nano sec
Neutrons probe Structures & Dynamics
DynamicsDynamics
SPIN ECHOSPIN ECHO
Back scatteringBack scattering
TOF/TASTOF/TAS
FANSFANS
SurfaceSurface TEM/SEMOptical
Internal StructureInternal Structure
Neutron DIFF
SANS/REF
Structure : 0.1 nm – 100’s nm
Neutron Sources
Use Thermal Energy
~1000’s MWth
Nuclear Power Plant
Neutrons
Research Reactor
Use
10’s MWth
Nuclear Fission
2017-07-04
4
Proton accelerator based neutron source
Spallation Neutron Source
Particle and Wave Properties of Neutron
y) x velocitmass(neutron
constant) s(Planck'
vm
h
Fast Neutron
V ~ 20,000,000 m/sec~ 0.00002 nm
tThermal Neu ron
V ~ 2,000 m/sec
~ 0.2 nmModeratorWater~35°C
Cold Neutron
V ~ 200 m/sec
~ 2 nmModeratorLiquid H2
~25 K
2017-07-04
5
Wavelength ~ Å, nm(thermal & cold neutron)
Energy ~ meV
Spin =1/2
Interacts with nuclei
No charge
Atomic & Nano length scale
Same magnitude as basic excitations in matter
Magnetic structure &dynamics
Why Neutrons ?
Contrast variation
Deep penetration
NIST, US
ORNL, US
SNS, US
ANSTO, Australia HANARO, Korea J-PARC, Japan
JRR-3, Japan
ILL, France
ISIS, UK
FRM-II, Germany
ESS, Sweden
Major Neutron Facilities in the World
2017-07-04
6
Four Circle Diffractometer
High Resolution Powder Diff.
NeutronRadiography
High IntensityPowder Diff.
Cold Neutron Guide
Bio-Diffractometer
Triple Axis Spectrometer
PGAA
Test Station & Residual Stress
Instrument
HANARO Thermal Neutron Facility
• 1995: 1st critical• Power: 30MW• Flux: ~2x1014 n/cm2sec
HANARO Cold Neutron Facility
40m SANS
18m SANS
Vertical-REF
DC-TOF
Horizontal-REF
Cold TAS
USANS
Thermal Neutron Instruments Cold Neutron Facility
2017-07-04
7
Basic Concepts of
Neutron Scattering
2017-07-04
8
Young’s Double Slit Experiment
Interference PatternIncoming plane wave Scattered wave
δ
4. Size of the slits, δ
1. Wavelength of the incident wave, λ2. Distance between the slits, d3. Distance from the slits to the detector, L
λ
d
L
InterferencePattern
Young’s Double Slit Exp. with Different Distance between Slits
• The distance between slits determines the interference pattern (periodicity).
Therefore, • If we measure the interference pattern,
we can determine the distance between slits.
d
'd’
2017-07-04
9
Young’s Double Slit Exp. with Different Slit Size
if slits are VERY small
• The envelope of interference pattern is determined by the size of slits.
Therefore, • If you measure the envelope the interference pattern, we can determine the size of slits.
Fourier Transform
Fourier Transform in Nature : Rainbow
Prism
Fourier Transform is decomposition or separation of a function into a sum of sinusoidsof different frequencies.
t
h(t)
T
Frequency :T
f1
Period
2017-07-04
10
Fourier Transform in 1D
time
T
x
Time domain Real space
: wavelength (cm)T : period (sec)
Tf
1 : time frequency (sec-1)
Tf
22 : angular frequency
(radians sec-1)
2
Q : spatial frequency (radians cm-1)
: wave vector
dtethH ti )()(
dxexhQH iQx)()(
)(xh)(th
dQeQHxh iQx)()(
deHth ti)()(
Fourier Transform : Examples
xQAxh ocos)(
x
2
oQ
Q
H(Q)
real-Qo +Qo
F.T.
dxexQAQH iQxo )cos()(
)()(222
)()(oo
xQQixQQiiQxxiQxiQ
QQQQA
dxedxeA
dxeee
A oo
oo
2017-07-04
11
Fourier Transform : Examples
Q
h(x)
-xo +xo
F.T.
H(Q)
x
Fourier Transform: Examples
x Q4
a
4
a
a
4a
8a
4
a
8
h(x) H(Q)
F.T.Ao
x Q2
a
2
a
a
2a
4
a
6a
2
a
4
a
6
h(x) H(Q)
F.T.
Top Hat function Sinc function (sinx/x)
Ao
2/
)2/sin()(
Qa
QaaAQH o
2017-07-04
12
Fourier Transform: Convolution Theorem
Convolution
Multiplication
F.T.
H(Q) = G(Q) × F(Q)
dxfgxh )()()(
)()( xfxg
)()()( xfxgxh
Fourier Transform: Convolution Theorem
=
g(x) f(x)h(x)
x
F.T.
F(Q)
Q
F.T.
=
H(Q)
Q
F.T.
×
G(Q)
Q
2017-07-04
13
Re-Visit Young’s Double Slit Exp.
if slits are VERY small
Q
h(x)
-xo +xo
F.T.
H(Q)
x
The interference pattern is|Fourier Transform of two slits|2
Re-Visit Young’s Double Slit Exp.
h(x)
-xo +xo
F.T.
x
The interference pattern is|Fourier Transform of two slits|2
Q
2017-07-04
14
The interference pattern ofYoung’s double slit experiment
is|Fourier Transform of two slits|2
Young’s Double Slit Experiment
Interference PatternIncoming plane wave Scattered wave
δ
λ
d
L
1
2
21 2
21
2 I
2017-07-04
15
Young’s Experiments with Neutron Wave and Atoms
Neutron Scattering
AtomsIncident Neutron WaveDetector
(counts neutrons)
2
j
RQ-ij
jebdΩ
dσ
Add up phase factors of the scattered waves from all the scattering centers in the sample.
Differential Scattering Cross Section
Φd
d
d
d into secondper scattered photons of No.
ddr
drdA
sin 2
2
inin Ek ,
outout Ek ,
x
y
z
incident photons
scatteredphotons
transmittedphotons
Sample
Detector
secondper per neutronsincident of No. 2cmΦ
ddr
dAd sin
2detector
detector
2017-07-04
16
jout Rn
jR
0
ink
outk
ie
iRkki ee jinout
)(
Scattering Vector Q and Scattering Cross Section
2
j
RQ-ij
jebdΩ
dσ
Add up phase factors of the scattered waves from all the scattering centersin the sample.
jb = scattering length of the j-th nucleus
jin Rn
ininin kkn
/
outoutout kkn
/jin Rk
jout Rk
inout kkQ
LengthQ
1 ofUnit
jRQie
inout kkQ
2
koutin kk
ink
outk
Q
For elastic scattering
2sin2
kQ
2sin
4
Q
Scattering vector Q
Note: The dimension of Q = 1/Length
dQ
2
Qd
2or
2017-07-04
17
What Contrast do we use in Neutron Scattering ?
)()( rBμr MV
)(2
)(2
rr N
nN b
mV
Neutron Interaction Potentials
: Neutron-Nucleus
: Neutron-Magnetic Field
Scattering Length
b = bN ± bM
Nuclear Magnetic
V
bn
jj
Scattering length density,
= bound coherent scattering length of atom jjb
V = volume containing the n atoms
Neutron Scattering Length Density
2017-07-04
18
SL
D (
1010
cm-2
)
0
H2O-0.56
D2O6.65
SLD bi
i
n
V
NA
mass
Mw
i
bi molecule
N A Avogadro' s number = 6 1023
Mw molecular weight
bi bound coherent scattering length of atom i
H2O + D2O(1:1 volume)
3.05
• H2O + D2O mixture (1:1 volume)SLD ,Mixture xH2O SLD ,H2O (1 xH2O ) SLD ,D2O
0.5 (0.56 1010 cm 2 ) 0.5 (6.65 1010 cm 2 )
3.05 1010 cm 2
- bound coherent scattering length (10-13 cm) bH = -3.74 bD = 6.67 bO = 5.80
• H2O
• D2O
SLD ,H2O (6 10 23 / mol )1.0g / cm 3
18g / mol
2 bH bO
0.56 10 10 cm 2
SLD , D2O (6 1023 / mol )1.1g / cm 3
20 g / mol
2 bD bO
6.65 1010 cm 2
Calculation of Neutron Scattering Length Density
Neutron Contrast Variation
Neutron interacts with Nuclei.
HD
CO
Si
Isotope substitution (H/D)
Neutron cross section.
• VERY sensitive to Hydrogen• H and D are VERY different
2017-07-04
19
H2O/D2O mixtures can contrast match most materials
SL
D (
x10-1
0cm
-2)
Mole fraction D2O
H2O
D2O
2
)( j
RQij
jebQd
d
Neutron Scattering : Fourier Transform
Differential scattering cross-section
2
)()(
rderQd
d rQisld
)()( jj
Rrrn
: Atomic number density
j
jj Rrbr )()( sld
: Scattering length density
1)r( rd
)R()R-r()(
frdrf
Dirac delta function
jRQi
jj
rQij
jj
rQi ebrdeRrbrderr )()()(F.T. sldsld
2017-07-04
20
Young’s Experiments with Neutron Wave and Atoms
Neutron Scattering
AtomsIncident Neutron WaveDetector
(counts neutrons)
Fourier Transformof atomic positions
2
)()(
rderQd
d rQisld
Small-Angle Neutron Scattering
2017-07-04
21
range 0.001 Å-1 to 0.8 Å-1
Small-Angle Neutron Scattering
ink
outkQ
inout kkQ
2sin
4
2sin2
kQ
sampleincident neutrons
Q
Length scale, 1 nm to 500 nm 5 to3.0
d
Qd 2
Small scattering angle,
We measure the interference pattern from structures.
)(Q
d
d= differential scattering cross-section
• SANS measures the bulk nanostructures of 1 nm – 100’s nmin solids, liquids, gel or mixtures.
• Practically, anything that has proper1) length scale, 2) neutron contrast and 3) sample volume
0.1 ~ 10 mm
1.0 ~ 2.5 cm (beam diameter)
Typical sample volumeNeutron scattering length density
Systems that SANS Can Measure
2017-07-04
22
HANARO Cold Neutron Research Facility
40m SANS
18m SANS
Bio-REF
USANS
Cold TAS
Vertical-REF
DC-TOF
SANS Measures the Inhomogeneity of the Scattering Length Density in Sample
2)( of TransfomFourier )( rQ
d
d
2
)(1
)(1
)(
rrQQ rQ deVd
d
Vd
d i
2017-07-04
23
What Information from SANS ? Particulate Systems
2( ) Fourier Transfom of ( )
d
d
Q r
)( )(
1)( )(2
Q RRQ
SPn
eN
FV
N
p
N
i
N
j
i
P
pp p
ji
Structure factorInter-Particle interference
Interactionbetween particles
Form factorIntra-Particle interference
Shape and dimensionsof particles
Remember the convolution theorem
Positions of particlesShape and Size
of particles
Convolution
pN
jjRrrn )()( )( rf
)(r
Particles dispersed in solvent
FT FT
p
j
N
j
RQ-ie
rderfQF rQi
)()(×
2017-07-04
24
2
3
3(sin( ) sin( ))( )
( )
QR QR QRP Q
QR
221
0
2 ( sin ) sin(( cos ) / 2)( ) sin
sin cos ) / 2
J QR QLP Q d
QR QL
2
1 13 ( ) ( ) 3 ( ) ( )( ) c c s c s s solv s
c s
V J QR V J QRP Q
QR QR
Sphere
Cylinder
Core-Shell
Particle Form Factors
Thiolated PEG
Attaching Molecules on the Surface of Gold Nanoparticles
What is the shell thickness of the functionalized gold nanoparticles in water ?
2017-07-04
25
SANS Measurements and Form Factor Analysis
Spherical Core-Shell Form Factor
0.5 vol % Au-PEG-1 and -2 in D2O Measured using the 40 m SANS at
HANARO
Parameters Au-PEG-1 Au-PEG-2
core radius (Rcore , nm) 6.5 6.5
thickness of shell 1 (Tshell 1 , nm) 2.4 1.6
thickness of shell 2 (Tshell 2 , nm) 1.2 2.0
thickness of shell 3 (Tshell 3 , nm) 0.9 3.4
volume fraction of PEG in shell 1 0.98 0.94
volume fraction of PEG in shell 2 0.51 0.75
volume fraction of PEG in shell 3 0.40 0.37
total shell thickness (nm) 4.5 7.0
Attractive square well
Hard sphere
Coulomb repulsion
Structure Factors
S(Q
)
Q × diameter
S(Q) ≈ 1 when sample is very dilute.
2017-07-04
26
SANS Intensity
• S(Q) ≈ 1 when sample is very dilute. )()( QPnQI p
)()()( QSQPnQI p
Q
P(Q), scaled
S(Q)
I(Q)
drrQr
QrrQ
d
d
V
22 )sin(
)(4)(
Contrast Correlation function
Orientation average
')'()'(
')'()'()(
rrr
rrrr
d
dr
What Information from SANS ? Non-Particulate Systems
2017-07-04
27
2
j
RQ-ij
jebdΩ
dσ
2
)()(
rderQd
d rQisld
)()()( QSQPnQd
dp
Summary of Small Angle Neutron Scattering
SANS and SAXS Studies ofBinary Superlattices of 1D Colloids
Hierarchically Self-Assembled Hexagonal, Honeycomb, and Kagome Superlattices
of Binary 1D Colloids
Sung-Hwan Lim, Taehoon Lee, Younghoon Oh, Theyencheri Narayanan, Bong June Sung, and Sung-Min Choi
Highly Ordered and Highly Aligned 2D BinarySuperlattice of a SWNT/Cylindrical-Micellar System
Sung-Hwan Lim, Hyung-Sik Jang, Jae-Min Ha, Tae-Hwan Kim, Pawel Kwasniewski,Theyencheri Narayanan, Kyeong Sik Jin, and Sung-Min Choi
2017-07-04
28
Salt
Semiconductor
Diamond
SuperconductorLi-ion battery
Magnets
Atomic Crystals
Nanoparticles as Building Blocks
2017-07-04
29
2017-07-04
30
Optical
Electrical
Magnetic
Chemical
Emergent Properties
Nanoparticle Supercrystals
Nanoparticle Supercrystals
Closely packed silica nanoparticles (150-300 nm) Diffraction of Light
OPAL
2017-07-04
31
Binary Superlattice of Spherical Nanoparticles
Kalsin et al. Science 312, 420 (2006) C.B. Murray et al Nature (2009)
C.B. Murray et al Nature (2006)
Slow Solvent Evaporation Induced
Electrostatic Interaction
DNA-Programmed Nanoparticle Superlattice
S-A10-AAGACGAATATTTAACAATTCTGCTTATAAATTGTT-A-GCGC
S-A10- AAGACGAATATTTAACAATTCTGCTTATAAATTGTT-A-ATGC
X
TACG - A -TTGTTAAATATTCGTCTTAACAATTTATAAGCAGAA-A10-S
Y
G C
T A Z
CGCG - A - TTGTTAAATATTCGTCTTAACAATTTATAAGCAGAA-A10-S
Z
FCC
BCC
C.A. Mirkin et al, Nature (2008)
Linker Z Linker Z
Linker X Linker Y
C.A. Mirkin et al, Science (2013)
2017-07-04
32
Binary Superlattices of 1D Nanoparticles
Spherical Nanoparticles
1DNanoparticles
?
Purdy et al, Phys. Rev. Lett. (2005)
fd-virus (thin) + fd-PEG (thick)
1.1 < Dthick/Dthin< 3.7
Binary Mixture of 1D Nanoparticles
fd-v
irus
(mg
/ml)
fd-PEG (mg/ml)
Gold Nanorods
Slow evaporation on substrate
+
18 nm 45 nm
Aminah and Choi
2017-07-04
33
Binary Superlattices of 1D Nanoparticles
+
Medhage et al., J. Phys. Chem. 97, 7753 (1993)
Phase separation ? Random mixing ? Something lese ?
?
C12E5/Water
5 nm
4.4 nm
SAXS Measurements at Hexagonal Phase (10 oC)
p-SWNT/C12E5/D2O (15/45/55)
C12E5/D2O (45/55)
Without p-SWNT
With p-SWNT
2017-07-04
34
40m SANS at HANARO Cold Neutron Facility
40m SANS
SANS: Contrast Matching
Deuteratedp-SWNTs+
Neutron Scattering Cross Section
D2O100%
100
101
102
103
104
105
106
107
I(Q
)
2 3 4 5 6 7 8 90.1
2 3 4
Q (1/A)
100
101
102
103
104
105
106
107
I(Q
)
2 3 4 5 6 7 8 90.1
2 3 4
Q (1/A)
80%
60%
100
101
102
103
104
105
106
107
I(Q
)
2 3 4 5 6 7 8 90.1
2 3 4
Q (1/A)
40%
100
101
102
103
104
105
106
107
I(Q
)
2 3 4 5 6 7 8 90.1
2 3 4
Q (1/A)
20%
100
101
102
103
104
105
106
107
I(Q
)
2 3 4 5 6 7 8 90.1
2 3 4
Q (1/A)
15%
100
101
102
103
104
105
106
107
I(Q
)
2 3 4 5 6 7 8 90.1
2 3 4
Q (1/A)
10%
100
101
102
103
104
105
106
107
I(Q
)
2 3 4 5 6 7 8 90.1
2 3 4
Q (1/A)
100
101
102
103
104
105
106
107
I(Q
)
2 3 4 5 6 7 8 90.1
2 3 4
Q (1/A)
5%
0%10
0
101
102
103
104
105
106
107
I(Q
)
2 3 4 5 6 7 8 90.1
2 3 4
Q (1/A)
D2O100%
45% C12E5 in Water (D2O/H2O)
Contrast Matching
HydrogenatedC12E5
100% D2O
60% D2O
Surfactants C12E5 are not visible by neutron
5% D2O
2017-07-04
35
2 3 4 5 6 7 8 90.1
2 3
Q(1/A)
SAXS
SANS (without p-SWNT)C12E5 are contrast matched with D2O/H2O
New peak
p-SWNT/C12E5/Water = 15/45/55 at 10 ºC
1
7
1
3
2
SANSDeutrated p-SWNT
1
3
+
Shear-Induced Alignment of p-SWNT/C12E5/D2O
Isotropic
Hexa
Cooled Down from Isotropic Phase to Hexagonal PhaseUnder Oscillatory Shear
Shear Stress Amplitude: 500 PaFrequency: 5 Hz
ID02 BeamlineESRF, Grenoble, France
Oscillatory Shear
with T. Narayanan
2017-07-04
36
Shear-Induced Alignment of p-SWNT/C12E5/D2O
(Radial) (Tangential)
Lim and Choi et al, Angew. Chem. Int. Ed. 53, 12548 (2014)
+
Hexagonal Array in Honeycomb Lattice
AB2-Type
Lim and Choi et al, Angew. Chem. Int. Ed. 53, 12548 (2014)
5.0 nm 4.4 nm
• Maximization of Free Volume Entropy
2017-07-04
37
C12E5/water
Binary Superlattice of 1D Nanoparticleswith Varying Diameter Ratio
4.4 nm
Polymerized Rod-like Nanoparticle
p-CnTVB
10-3
10-2
10-1
100
101
102
103
104
Scat
teri
ng I
nten
sity
(a.u
.)
4 5 60.1
2 3 4 5 61
2 3 4
q(nm-1
)
n=16D=3.9 nm
n=14D=3.4 nm
n=12D=3.0 nm
n=10D=2.6 nm
n=8D=2.1 nm
No. of CarbonDiameter
SANS Form Factor Analysis
p-CnTVB
+
Variation of Diameter Ratio at a Fixed Concentration
Without p-CnTVB
(F127/water=45/55)
With p-CnTVB (10/45/55)
0.910.800.680.59 Rrod/RC12E5
2017-07-04
38
10/45/55
Variation of p-CnTVB Concentration (for n=10)
p-C10TVB/C12E5/Water (X/45/55)
9/45/55
8/45/55
7/45/55
6/45/55
Diameter Ratio = 0.59
5/45/55
T= 4℃10/45/55 5/45/55
10/45/55
Variation of p-CnTVB Concentration (for n=10)
p-C10TVB/C12E5/Water (X/45/55)
9/45/55
8/45/55
7/45/55
6/45/55
Diameter Ratio = 0.59
5/45/55
T= 4℃10/45/55 5/45/55
deuterated p-C10TVB
+
CM C12E5/Water
Contrast matched SANS
2017-07-04
39
AB2 to AB3 Structural Transition
p-C10TVB/C12E5/Water (X/45/55)
Diameter Ratio = 0.59
T= 4℃
AB2-TypeHexagonal Array in “Honeycomb Lattice”
AB3-TypeHexagonal Array in “Kagome Lattice”
SH Lim, T. Lee, Y. Oh, T, Narayan, BJ Sung, and SM ChoiNature Communications (accepted)
SH Lim, T. Lee, Y. Oh, T, Narayan, BJ Sung, and SM ChoiNature Communications (accepted)
In collaboration with Prof. Bong Jun Sung
Theoretical Free Energy Calculations and Phase Diagram
Free energyTheoretical
Phase DiagramExperimental
Phase Diagram
p-CnTVBs and C12E5 are modeld as 2D hard discs A and B of different effective diameters.
2017-07-04
40
Summary
AB2-Type AB3-Type
• Small angle neutron and x-ray scattering techniques are very powerful for studying soft matter.
2
)()(
rderQd
d rQisld
1
Neutron Spin Echo Investigations of Lipid Membranes Interacting with Proteins
Sung-Min Choi
Neutron Scattering and Nanoscale Materials Lab.Dept. of Nuclear and Quantum Engineering
KAIST
Antonio Faraone (NIST) Philip A. Pincus (UCSB)Steven R. Kline (NIST)
Ji-Hwan LeeChangwoo Do (now at ORNL)Shin-Hyun KangMin-Jae Lee
Collaborators
D. M. Engelman, Nature, 438, 578 (2005)
Cell Membranes
2
Reynwar B.J. et al, Nature 447 (2007)
Interplay of Proteins and Lipidsdetermines the Shape and Function of Cell Membranes
• To create shapes and functions, membranes must undergo deformations • The work to deform membranes is determined by
the structure and elasticity of the membrane.
Zimmerberg J et al, Nature Rev.: Mol. Cell Bio. 7,9 (2006)
Antimicrobial Peptide Activity in Multicellular Organisms
E. coli treated with an Antimicrobial Cationic Peptideat low concentration (left) and at high concentration (right).
www.cmdr.ubc.ca/cool.html
• Provides the self-defense mechanism of plants and animals
3
Transmembrane Pore-Forming Models
K. A. Brogden, Nature Rev. MicroBiol. 3, 238 (2005)
L. Yang et al, Biophys. J. 21, 1475 (2001)
Antimicrobial Peptide-Membrane Interaction
P/L = Peptide-Lipid molar ratio
(P/L)*Pores are formed
H.W. Huang et al, Phys. Rev. Lett, 92, (2004)
Thermal Fluctuation of Membrane Interacting with Peptides
Add more peptides
How does the elasticity of membrane changes with P/L (pore formation) ?
???
M.T. Lee et al, Biochemistry 43, 3590 (2004)
4
Model System
Melittin• A cationic peptide from European bee venoms• Forms toroidal pores in membrane
~3.5 nmL. Yang et al, Biophys. J. 21, 1475 (2001)
Toroidal Pore
~4 nm
~7.7 nm
DOPC• 1,2-Dioleoyl-sn-glycero-3-phosphatidylcholine• low gel-to-fluid temperature (Tm = -20°C)
~ 3.7 nm
• Prepared by extrusion through polycarbonate filters (100 nm)
• 10 mM Hepes 150 mM NaCl 2 mM EDTA D2O buffer solution
(P/L)*= ~0.4 %
Unilamellar
ca.100 % Perforated
50
40
30
20
10
031P
-NM
R I
nte
nsi
ty R
atio
(%
)
2.01.51.00.50.0
(P/L ) (%)
When are the pores formed ? 31P-NMR Signal Intensity vs P/L
5
6 mM DOPC + Melittin in 10mM Hepes 150 mM NaCl 2 mM EDTA D2O buffer.
For P/L = 0/100 – 20/100 were measured at ~ 6hrs after melittin-mixing.
SANS Data were analyzed using the polydisperse spherical core-shell model.
Spherical Core-Shell model analysis
Vesicle Structure vs P/L : SANS
2
1 13 ( ) ( ) 3 ( ) ( ( )). .( ) c c s c s s solv s
s c s
V J qR V J q RVol fracP q
V qR qR
SANS Measurements
10-4
10-3
10-2
10-1
100
101
102
103
104
105
106
107
108
109
1010
1011
1012
I(q)
80.001
2 4 6 80.01
2 4 6 80.1
2 4
q(Å-1)
Melittin / 6mM DOPC
P/L=0/100P/L=1/100P/L=3/100P/L=5/100P/L=8/100P/L=12/100P/L=14/100P/L=16/100P/L=20/100
Sensitive to single membrane dynamics
(qR >> 1)
Measured at NG7 SANS NIST
Neutron Spin Echo Spectrometer (NG7 NSE, NIST)
6
3/2)(exp)0,(
),( q
qI
qI
Zilman and Granek , Phys. Rev. Lett., (1996)
32/1
025.0)( qTkTk
q BBk
Neutron-Spin Echo Measurements
• Intermediate dynamic structure factor of membrane at large Q which is sensitive to single membrane dynamics.
Zilman and Granek Model
(qR >> 1) = Bending modulus
)(qNeutron Spin-Echo Intensity
= Solvent viscosity
M.C. Watson and F.L.H. Brown, Biophys. J. (2010)
Elasticity of Lipid Bilayers Interacting with Melittins
32/1
~025.0)( qTkTk
q BBk
kd 2~
For a pure DOPC bilayer
J.-H. Lee, S.M. Choi, C. Doe, A. Faraone, P. A. Pincus, S.R. KlinePhys. Rev. Lett., 105, 038101 (2010)
Tkq
q
B
3
)(~
7
Change of Bending Modulus UponPerturbation of Local Chain Packing by Peptide Adsorption
Pure Membrane
6
8
10
12
Mixed Membrane (12 + 6)
Effect of Chain Packing Disordering on Bending Modulus
Szleifer et al, Phys. Rev. Lett. 60 (1988)
Surfactant Bilayer
Be
nd
ing
Mo
du
lus
Bending Modulus vs P/L
Region I : a) Membrane thinning (ca. -0.7 kBT)
b) Perturbation of hydrocarbon chain packing (dominant)
Region II : a) Membrane thickening (ca. +0.8 kBT)
b) High rigidity of pore forming melittins
Region III : ??
50
40
30
20Ben
din
g r
igid
ity
( k
BT
)
43210P/L (%)
P/L*
II IIII
8
Pore patchBound patch
or
or = Outer radius of a pore patch
= 3.9 nm
Yang et al, Biophy. J. (2001)Huang, J Phys II France (1995)
= Range of deformation
=
= 2.5 nm (DOPC)
(16h2κ/κa)
1/4
Membrane mediated interaction between pores
When the deformed membrane regions become overlapped, the membrane-mediated repulsive interaction between patches become significant.
50
40
30
20Ben
din
g r
igid
ity
( k
BT
)
43210P/L (%)
P/L*
Bending Modulus vs P/L
Region I : a) Membrane thinning (ca. -0.7 kBT)b) Hydrocarbon chain disordering (may be more dominant)
Region II : a) Membrane thickening (ca. +0.8 kBT)b) High rigidity of pore forming melittins
Non-interacting orweakly interacting
Interacting patches
Region III : Strong interaction between pore patches
7.7 nm
(P/L) Inter-pore distance
2 % 12 nm 4 % 7.9 nm
I II III
J.-H. Lee, S.M. Choi, C. Do, A. Faraone, P. A. Pincus, S.R. KlinePhys. Rev. Lett., 105, 038101 (2010)
9
Summary
• The thermal fluctuation and elasticity of DOPC vesicles interacting with pore-forming melittins has been measured by the Neutron Spin-Echo Spectroscopy,for the first time.
• The change of thermal fluctuation and elasticity with (P/L) was understood in terms of membrane thinning and thickening, chain packing perturbation and inter-pore interaction.
• The change of elasticity of DMPC-gD, DLPC-gD with (P/L), covering repulsive and attractive interaction ranges, has been investigated for the first time.