RELEASE OF CONTENT THROUGH MECHANO- SENSITIVE GATES …€¦ · Sukharev et al.!1997" Annu Rev Physiol 59: 633$657 M. Louhivuori / ISSP workshop / Tokyo, 24.8.2010. non-selective

RELEASE OF CONTENT THROUGH MECHANO-

SENSITIVE GATES IN PRESSURISED LIPOSOMES

Martti LouhivuoriUniversity of Groningen

M. Louhivuori / ISSP workshop / Tokyo, 24.8.2010

MARTINI

coarse-grained model

www.cgmartini.nl


http://www.cgmartini.nl

http://www.cgmartini.nl

MARTINI CG modelinteraction sites

DPPC cholesterol peptide !ALYWK"

water

butane

C1

SC1

SC3

SP1QaQo

Na

C1

SC4

P4

C1

C5

SNd

SC4

SC4

Qd

SP1

C3No


parametrisation of MARTINI

experimental !thermodynamic" data

! non#bonded interactions

atomistic MD simulations

! bonded interactions


parametrisationbonded interactions

angle

dist

ribu

tion


parametrisationnon#bonded interactionsM. Louhivuori / ISSP workshop / Tokyo, 24.8.2010

Rhombohedral phase (experimentally observed for DOPC/DOPE 3:1 and 2:1 Lyan & Huang, 2002)

side view top view

THE VALIDATIONTHE VALIDATION comparing to experimental measurements

Reproduced in CG simulation (Marrink & Mark, Biophys. J., 2004)



bilayers rafts

sugars

membrane proteins

vesicles

vesicles w/ proteins


bilayers rafts

sugars

membrane proteins

vesicles

vesicles w/ proteins

MARTINI CG modelinteraction sites

DPPC cholesterol peptide !ALYWK"

water

butane

C1

SC1

SC3

SP1QaQo

Na

C1

SC4

P4

C1

C5

SNd

SC4

SC4

Qd

SP1

C3No

SPEEDshort#range interactions

large time#stepfew degrees of freedom

GENERALconsistent modeling

biomolecular systemseasily extended

EASE of USEbuilding#block approach

limited # of particlesphysical units

ACCURACYparametrisation based on

thermodynamic datamulti#level optimisation


drugs patient

targeted drug release.how?

mmm!ouch!


drug delivery vehicle“nanobot”

BOOM


drug delivery vehicle“nanobot”


mechano-sensitive channels

“safety valves” of cell

sense tension in the membrane

MscL, MscK, MscS, MscM

< 10 mN/m

PDB: 2VV5


MscLcontrollable activation & non!selective conductance

bottom

top


MscL activity

flickering conductivity

multiple levels

! subconductive states

activation < 1 ms

de#activation 1$100 msSukharev et al. !1997"

Annu Rev Physiol 59: 633$657


non-selective channel

no ion selectivity

even small proteins pass through! 15$20 Å

Cruickshank et al. !1997"Biophys J 73: 1925$1931


photosensitivivity

attached compound undergoes light induced charge separation

reversible

localised Koçer et al. !2005"Science 309: 755$758


photosensitivivity

photosensitive lipids used to transfer signal to mechanical stress

reversible

localised

Folgering et al. !2004"Langmuir 20: 6985$6987


nano-containeraka liposome

nano-particlesaka drugs


nano-transporter

tiny lipid vesiclesmembrane fusion

trans#membrane transport

drug delivery

curvature e%ects

mechano#sensitivepressure valves of cells

touch & hear

non#selective, large membrane channel

liposomes

MscL


game plan

BOOM


MscL saves the day


oh-oh! 660 kN/m·s ! lysis

140 kN/m·s ! ok


analysis


analysisclosed open 5 us


activation

mechanism


post-activation

67 mN/m

" 24 nm

1.04 H2O / ns


equilibrium?

0

20

40

60

80

100

120

140

0 10 20 30 40 50 60 70 80 90

tensionpressure

time (µs)

tensi

on

(mN

/m

)/

pre

ssure

(bar

)

(-0.54 ± 0.02) mNm

/µs

(-1.12 ± 0.05) bar/µs

(86 ± 4) µs

(93 ± 4) µs


-30

-20

-10

0

10

20

30

2 3 4 5 6

r [nm]

MFFA boundary potentials

mimic interactions with bulk solvent

0

0.5

1

1.5

2

2.5

3

3.5

0 1 2 3 4 5 6

r [nm]

RDF

MFFA

Risselada et al. !2008"J Phys Chem B 112: 7438$7447


pumping water into liposomes

additional mean#field potential inside the liposome

start with r = 0.01 nm

increase slowly for 20ns until r = 3.9 nm

fill the cavity with water, relax and repeat as needed


water-repellant lipid tails

modified Lennard#Jones potential against water

C5 wC5epsilon 2.0 2.0sigma 0.47 0.7

DOPC wDOPC

NC3

PO4

GL1

C1GL2

C2

D3

C4

C5

C1

C2

D3

C4

C5

NC3

PO4

GL1

C1GL2

C2

D3

C4

C1

C2

D3

C4C5wC5

V (r) = 4!!""

r

#12!

""

r

#6$

C5wC5

NC3

PO4

GL1

C1GL2

C2

D3

C4

C5

C1

C2

D3

C4

C5


3D pressure fields

divide system into a 3D grid

use local virial for each volume element

calculate averages

Plocal(r) =1V

!

"#

i

! (r ! ri)mivi " vi +12

#

i !=j

Fij

$

Cij!(r ! 1)dl

%

&

Ollila et al. !2009"Phys Rev Lett, 102: 078101



flow rate

R

0 5 10 15 20Box (nm)

0

500

1000

1500

2000

Den

sity

(k

g m

-3)

ProteinwDOPCWPO4

Partial densities

R

!

!

summarylarge#scale biological systems accessible to CG simulations

release of an osmotic shock via MscL activation achieved

MscL activation is indeed a last#ditch e%ort to prevent lysis

iris#like, non#symmetric opening

water flux OUT: !6.0 ± 1.3"& ions/ns

IN: !1.7 ± 0.3"& ions/ns

MODEL: 0.2$40& ions/ns& Steinbacher et al. !2007" & CurrTopicsMembranes 58:1#24

pore radius !11.6 ± 0.8" Å& !exp. 15$20 Å"

blocking of the channel by the cytoplasmic helices a first step in closure?


future

dye molecules to directly compare our release of nano#particles to experimental data

activation of the channel using lyso#lipids

nano#pores formed by other molecules, e.g. anti#microbial peptides



Siewert#Jan MarrinkErik van der GiessenJelger Risselada

UPPSALA UNIVERSITET

TAMPERE UNIVERSITY OF TECHNOLOGY

David van der Spoel

Samuli OllilaIlpo Vattulainen

Stockholm University

Erik Lindahl

Documents

RELEASE OF CONTENT THROUGH MECHANO- SENSITIVE GATES …€¦ · Sukharev et al.!1997" Annu Rev Physiol 59: 633$657 M. Louhivuori / ISSP workshop / Tokyo, 24.8.2010. non-selective