Membrane interactions of lipopeptides and saponins, two ... · PDF filelipopeptides and saponins, two groups of natural amphiphiles J. Lorent (public defence) Main supervisor: M-P.

Membrane interactions of lipopeptides and saponins, two

groups of natural

amphiphilesJ. Lorent

(public defence)

Main supervisor

: M-P. Mingeot-Leclercq (FACM)Co-supervisor

: J. Quetin-Leclercq (GNOS)

Introduction : physico-chemical

properties

of amphiphiles

Polar part(hydrophilic)

Apolar

part(hydrophobic)

S

O

O

ONa

Sodium dodecylsulfate

(detergent)

Introduction : selected

amphiphiles of natural

origin

surfactin hederagenin

Saponin Triterpenic

acidLipopeptide

α-hederin

C3C3

HN

HN

O

O

NH

HN

OOH

O

O

HN

HNO

O

NH

O

O

O

OH

O

CH2OH

COOH

O

HO

H

H

HO

HOH

HH

O

OH

H

OH

H

H

H

OCH3

H

O

H

CH2OH

COOH

HO

Isolated

from

Hedera helix

L. Isolated

from

Bacillus

subtilis

HHO H

HO H

HO H

HO H

HO

water (polar)

air (apolar)Interfacial

/ Surface tension or energyγ

= (F / l) or (E / A)

Introduction : amphiphiles (effect

on interfaces)

HHO H

HO H

HO H

HO H

HO

water

air

Interfacial

/surface tensionγ

= (F / l) or (E / A)


on interfaces)

HHO H

HO H

HO H

HO H

HO

water

air

Interfacial

/surface tensionγ

= (F / l) or (E / A)


on interfaces)

HHO H

HO H

HO H

HO H

HO

water

Interfacial

energy

air

Emulsions / suspensions


on interfaces)

• liquid

/ air (foam)

• liquid

(apolar) / liquid

(polar)

creams

Introduction : amphiphiles (self-aggregation)Polar part

(hydrophilic)Apolar

part(hydrophobic)

S

O

O

ONa

water


Apolar

part(hydrophobic)

S

O

O

ONa

[amphiphile]

Introduction : amphiphiles (self-aggregation)

water

S

O

O

ONa


Apolar

part(hydrophobic)

[amphiphile] > Critical

micellar

concentration (CMC)

Micelles or other

aggregates


on interfaces)

water

O

H

OH

H

H

O

H

P OO

O

R

O

HS OO

ONa

Introduction : amphiphiles (self-aggregation, intrinsic

curvature

and polymorphism)

Normal hexagonal HI MicellesCone

shape Cubic

(CI

)

ς

> 0Intrinsic

molecular

curvature

Detergents Lysophospholipids

OH

H

HH


curvature

and polymorphism)

Inverse hexagonal HII Inverted

micellesInverted

cone

shape Inverted

cubic

(CII

)

ς

< 0

Cholesterol

O

H

O

HH

O

H

P OOO

OO

H

N

OH

NH

H

H

O

H

P OO

OR

O

H


curvature

and polymorphism)

Lamellar

Lα

, Lβ

, Lo (bilayers

or lipid

membranes) Cylinder

shape

ς

= 0

Phosphatidylcholines Sphingomyelins

Introduction : bilayer

dynamics

and cholesterol effects

Lateral

diffusion (10-7s)

Protrusion

(10-9s)

Flip-flop (10-3-104s)

Rotational

diffusion (10-8s)

Trans-gauche isomerization(10-12s)

Bond oscillations (10-12s)

Ondulations (10-6-1s)

Yeagle

et al. (2005)

Gel state Liquid

crystalline

state

Liquid

ordered

state

T

> Tm

Lβ Lα

Lo

+ cholesterol + cholesterol

Introduction : the biological

membrane (lateral heterogeneity

and lipid

rafts)

• enriched

in cholesterol, glyco-sphingolipids, saturated

phospholipids• Variable size (20-200nm)• functional

platforms: receptor

Fas, DISC

assembly

(death

receptor)

Lingwood

(2011)

Lo-phase (liquid

ordered) Lα-phase(liquid

crystalline)

Rafts (Lo

, liquid

ordered

phase)

Introduction : different

cell

deaths

induced

by saponins

cholesterol

mitochondriacaspase-3

condensation, fragmentation nuclei

death

receptor

MPT-pore

cytoplasm

necrosis

caspase-8

saponin

phospholipid

Ca2+

membraneinsertion

Apoptosis

membrane lysis

Ca2+

ROS

cyt

c

Mazzucchelli

et al., 2008

Trump

and Berezesky, 1995

Trump

and Berezesky, 1995

Choi et al., 2008

Jiang et al., 2010

Aim

of the study

HN

HN

O

O

NH

HN

OOH

O

O

HN

HNO

O

NH

O

O

O

OH

O

CH2OH

COOH

O

HO

H

H

HO

HOH

HH

O

OH

H

OH

H

H

H

OCH3

H

O

H CH2OH

COOH

HO

? Raft located

death

receptor

Materials

and methods

: Membrane models•

LUV (large unilamellar

vesicles)/ MLV (multilamellar

vesicles)–

Size = 100 nm-1 µm –

Easy

determination

of lipid

concentration Quantification

of effects

and amphiphilie/lipid

ratio

•

GUV (giant

unilamellar

vesicles)–

Size ~ 10-50 µm–

Difficult

determination

of lipid

concentration Easy

observation

of effects

•

SPB (supported

planar

bilayers)–

Difficult

determination

of lipid

concentration Nanoscopic

effects

•

Cancer cells

(monocytes from

leukemia, U937)–

Cytotoxicity, determination

of cell

death

Light microscopy

GUV

Cryo-TEM

LUV/MLV

AFM

SPB

Cells

Materials

and methods: phase separation

modelsDOPC : DPPC (1:1)

Lα

(liquid

crystalline) / Lβ

(gel state)

Tm

= -3°C Tm

= 42°CTR-DPPE (Lα

)labeled

GUVsDOPC,DMPC

DPPC

Chol

DMPC:Chol

(3:1)Tm

=24°C

Lo Lα Lo

liquid

ordered

/ liquid

crystalline

Interaction of α-hederin

and hederagenin

with

lipids

and

effects

on membrane models

CH2OH

COOH

O

HO

H

H

HO

HOH

HH

O

OH

H

OH

H

H

H

OCH3

H

O

H

CH2OH

COOH

HO

α-hederin hederagenin

CH2OH

COOH

O

HO

H

H

HO

HOH

HH

O

OH

H

OH

H

H

H

OCH3

H

O

H

Induction of Highly

Curved

Structures in Relation to Membrane Permeabilization

and Budding

by the Triterpenoid

Saponins, α-

and δ-

hederin

Joseph Lorent1,2, Cécile S. Le Duff3, Joelle

Quetin-Leclercq2

and Marie-Paule Mingeot-Leclercq1*

1Université

catholique de Louvain, Louvain Drug Research

Institute, Cellular and Molecular

Pharmacology, Avenue E. Mounier 73, UCL B1.73.05, B-1200 Bruxelles, Belgium.

2Université


Institute, Pharmacognosy, Avenue E. Mounier 73, UCL B1.72.03, B-1200 Bruxelles, Belgium.

3Université

catholique de Louvain, -

Institute of Condensed

Matter

and Nanosciences, Molecules, Solids

and Reactivity, Place Louis Pasteur 1, UCL L4.01.04, B-1348 Louvain-la-Neuve, Belgium.

Journal of Biological

Chemistry

2013, 288, pp. 14000-14017

Phase separation

and permeabilization

induced

by the saponin

α-hederin

and its

aglycone hederagenin

in a raft mimicking

bilayer

Langmuir, submitted

Joseph Lorent1,4, Laurence Lins2, Domenech

Òscar3, Joelle Quetin-Leclercq4, Robert Brasseur2

and Marie-Paule

Mingeot-Leclercq1

1Université

catholique

de Louvain, Louvain Drug Research Institute, Cellular and Molecular Pharmacology, UCL B1.73.05, Avenue E. Mounier

73, B-1200 Brussels, Belgium.

2ULg, Centre de Biophysique Moléculaire Numérique, Agro-BioTech

Gembloux, Passage des Déportés, 2, B-

5030 Gembloux, Belgium.

3University of Barcelona, Departament

de Fisicoquímica, Facultat

de Farmacia, UB and Institut de Nanocie ncia

i Nanotecnologia

IN, 08028 Barcelona, Spain.

4Université


Institute, Pharmacognosy, B1.72.03, Avenue E. Mounier 72, B-1200 Brussels, Belgium.

Interaction with

dehydroergosterol

(DHE) in aqueous

solution : DHE spectroscopy

Both

molecules

bind

to DHE forming

some

type of mixed aggregates

(micelles) I50

(inflexion point) of α-hederin

corresponds to its

CMC (13 µM)

I1

= monomeric

peak(=monomeric

DHE)I2

,I3

= structured

emission(=crystalline

DHE)

340 365 390 415 4400

100

200

300

400

500

600

700increasing [-hederin]

Emission wavelength (nm)

Fluo

resc

ence

inte

nsity

(nm

)

1 10 1000.4

0.6

0.8

1.0

1.2

Log [-hederin] (µM)

Inte

nsity

rat

io (I

1/I 2

)

1 10 1000.4

0.6

0.8

1.0

1.2

Log [hederagenin] (µM)

340 365 390 415 4400

100

200

300

400

500

600

700increasing [hederagenin]

Emission wavelength (nm)


I50

= 12.3 ±

3.2

µM I50

= 14.3 ±

8.5 µM

402 396 nm 402 391 nm

I1I2

I2I1

I3I3

TRIS.HCl

(10 mM, pH=7,4)

Permeabilization

of LUV :

cholesterol

dependencePC/SM/PI/Chol

(4:4:3:x)

15 % cholesterol

30 % cholesterol

18h30min

p<0.05, ** p<0.01, *** p<0.001 two-way

ANOVA (18h vs 30min)

-hederin -hederin hederagenin0

25

50

75

100 4

Max

% c

alce

in r

elea

sed

0 10 20 30 40 50

0

25

50

75

100***

***

***

***

**

[-hederin]:[cholesterol]

% c

alce

in r

elea

sed

Permeabilization

depends

on the cholesterol

content and the saponin

sugar

chain

DMPC/Chol

(3:1) [compound] / [lipid] ratio

2 sugars 1 sugar no sugarNumber

of sugars

at

C3:

Calcein

(self-quenching

concentration)

Calcein

assay

LUV

fluorescence

permeabilization

Permeabilization

of LUV :

cholesterol

dependencePC/SM/PI/Chol

(4:4:3:x)

15 % cholesterol

30 % cholesterol

18h30min

p<0.05, ** p<0.01, *** p<0.001 two-way

ANOVA (18h vs 30min)

-hederin -hederin hederagenin0

25

50

75

100 4

Max

% c

alce

in r

elea

sed

0 10 20 30 40 50

0

25

50

75

100***

***

***

***

**

[-hederin]:[cholesterol]

% c

alce

in r

elea

sed

Permeabilization

depends

on the cholesterol

content and the saponin

sugar

chain

DMPC/Chol

(3:1) [compound] / [lipid] ratio

2 sugars 1 sugar no sugarNumber

of sugars

at

C3:

Reorganization

of MLV : cholesterol

dependence

Control

Disruption of bilayer

structure depends

on sugar

chain

and cholesterol

(not shown) Increase

of isotropic

motion and appearance

of hexagonal pattern with

α-hederin

10% α-hederin

20% α-hederin

DMPC/Chol

(3:1)

and macroscopic

phase separation

Malcolm et al. 2003Chemical

shift tensor

of the phosphore atom

31P-NMRPic plus large vesicule

plus grand

micelles

MLV (low

curvature)

(high

curvature)

C1

= highC2

= 0

Reorganization


dependence


structure depends

on sugar

chain

and cholesterol


of isotropic



α-hederin

10% α-hederin

20% α-hederin

Control

DMPC/Chol

(3:1)

+

Reorganization


dependence


structure depends

on sugar

chain

and cholesterol


of isotropic



α-hederin

10% hederagenin

20% hederagenin

Control

DMPC/Chol

(3:1)

>< 100 nm

Sterol/phospholipid

phase separation

in MLV

Decrease

of FRET efficiency

is

significant

only

with

α-hederin

at

high

compound/lipid

ratios

D

0 0.2 0.6 0.8 1.2

F

1

2

3

**

C

I 430/I

371

0 0.25 0.6 0.8 1.21

2

3

*

E

I 430/I

371

0 2500 5000 7500 100001

2

3A

time (s)

I 430/I

371

0 2500 5000 7500 10000

B Control0.250.60.81.2

time (s)

-hederin hederagenin

3h

24h

-hederin/lipids hederagenin/lipids

*p < 0.05, one-way

ANOVA (vs. Control)

[compound]/[lipids]DMPC/Chol

(3:1)

Low

energy

transfert

High energy

transfert

OH

OH

Förster

energy

resonance

transfert (FRET)

DHEDPH-PC

High FRET ratio

Low

FRET ratio

Sterol/phospholipid

phase separation

in MLV

Decrease

of FRET efficiency

is

significant

only

with

α-hederin

at

high

compound/lipid

ratios

D

0 0.2 0.6 0.8 1.2

F

1

2

3

**

C

I 430/I

371

0 0.25 0.6 0.8 1.21

2

3

*

E

I 430/I

371

0 2500 5000 7500 100001

2

3A

time (s)

I 430/I

371

0 2500 5000 7500 10000

B Control0.250.60.81.2

time (s)


3h

24h

-hederin/lipids hederagenin/lipids

*p < 0.05, one-way

ANOVA (vs. Control)

[compound]/[lipids]

DMPC/Chol

(3:1)


10 µM

budding

wrinkled

phase separation

spherical

Incubationtime

Effects

on GUV

< CMC(α-hederin)

2h

48hComplete

transformation of GUV

DMPC/Chol

(3:1)

1hTR-DPPE NBD-DPPE

Fluorescence microscopy


10 µM

budding

wrinkled

phase separation

spherical

Incubationtime

Effects

on GUV

< CMC(α-hederin)

1h

2h

48hComplete

transformation of GUV

DMPC/Chol

(3:1)

Phase separation

in GUV

NBD-DPPE (green)TR-DPPE (red)

Confocal

(1 slide)

Confocal

(profile, circumference)

Confocal

(3D)

α-hederin

(10 µM)

Accumulation of mainly

TR-DPPE into

worm-like

domains At

later

stages : separation

of TR-DPPE and NBD-DPPE

TR-DPPE(Lα

)

NBD-DPPE(Lo

)Confocal

slide

Profile (cirumference)

3D reconstruction (all slides)

Confocal

microscopyGUV (DOPC:DPPC:Chol)

Phase separation

in GUV


Confocal

(1 slide)

Confocal


Confocal

(3D)

α-hederin

(10 µM)

Accumulation of mainly

TR-DPPE into

worm-like

domains At

later

stages : separation

of TR-DPPE and NBD-DPPE


40 µM30 min

1h

budding

wrinkled

pore

Incubationtime

> CMC(α-hederin)

spherical

Effects

on GUV

Effects

on GUVα-hederin hederagenin

40 µM30 min

1h

4h

48h

N.D.

N.D.

phase separation

spherical

Incubationtime

Phase separation

in GUV


Confocal

(1 slide)

Confocal


Confocal

(3D)

Formation of small

circular

domains

with

hederagenin

hederagenin

(40 µM)

Permeation

of GUVs

to FITC-dextran

α-hederin

4 kDa10 µM

Budding(20s)

Gradual

permeation

to FITC-dextran

of 4 kDa

Phase separationControl (90 min)< CMC

(α-hederin)

Dextran-FITC solution

Permeabilization

Permeation

of GUVs

to FITC-dextran

α-hederin

4 kDa10 µM

Budding(20s)

Gradual

permeation

to FITC-dextran

of 4 kDa

Phase separation

(30, 60, 90 min)Control (90 min)< CMC

(α-hederin)

Permeation

of GUVs

to FITC-dextranα-hederin

Immediate

permeation

to FITC-dextran

of 4 kDa

4 kDa

4 kDa

40 µM

10 µM

Budding(20s)

Gradual

permeation

to FITC-dextran

of 4 kDa

Phase separation

No wrinkling(20s) Increasing

wrinkling

Control (90 min)

Control (90 min)> CMC

(α-hederin)

< CMC(α-hederin)

hederagenin

Permeation

of GUVs

to FITC-dextran

Control (48h)

4 kDa40 µM

Permeation

to FITC-dextran

of 4 kDa

after

48h

Phase separation

(48h)

Nanoscopic

effects

of α-hederin

on supported

planar

bilayer

Accumulation of membrane material

into

worm-like

structures (new mesophase) Formation of holes

with

increasing

size upon

time

t = 0 t = 6 min t = 15 min t = 24 min

40 µM

DMPC/Chol

(3:1)α-hederinAtomic

force microcopy

Profile (supported

planar

bilayer)

Nanoscopic

effects

of α-hederin

on supported

planar

bilayer

Accumulation of membrane material

into

worm-like

structures (new mesophase) Formation of holes

with

increasing

size upon

time

t = 0 t = 6 min t = 15 min t = 24 min

40 µM

DMPC/Chol

(3:1)

α-hederin

Conclusions : Membrane interactions of α-hederin

Internal

leaflet

External

leaflet

10 µM < CMC

Budding

Phase separation Transformation intoanother

mesophase

Positive curvature

Hydrophilic

sugar-sugar

interactions

Monomers

Pore formation ?

Transient

defects

?

DMPC

Cholesterol

immediate

later

Internal

leaflet

External

leaflet

40 µM

> CMC

toroidal

macroscopic

pore

immediate

permeabilization

without

transformationinto

new mesophase

(at

least at

short incubation periods)

Conclusions: Membrane interactions of α-hederin

Aggregates

and monomers

DMPC

Cholesterol

Conclusions : Membrane interactions of hederagenin40 µM

> CMC ??

accumulation into

round spots transformation into

inverse structures ? transient

defects

permeabilization

?

Inverse aggregates

?

Investigations on cellsα-hederin

and hederagenin

induce apoptosis and

non-apoptotic cell death in U937 and THP-1 cells in a cholesterol-dependent manner

Joseph Lorent1,2, Joelle Quetin-Leclercq2

and Marie-Paule

Mingeot-Leclercq1*

1Université

catholique

de Louvain, Louvain Drug Research Institute, Cellular and Molecular Pharmacology, UCL B1.73.05, avenue E. Mounier

73, B-1200 Bruxelles, Belgium.2Université


Institute, Pharmacognosy, UCL B1.72.03, avenue E. Mounier 73, B-1200 Bruxelles, Belgium.

0 20 40 600

25

50

75

100C

time (h)

% c

ell d

eath

0

10

20

30

40A

% fr

agm

ente

d nu

clei

0 20 40 60

D

time (h)

B CTL10 µM15 µM20 µM40 µM


Cell

death

: Acridine orange/Ethidium

bromide

in U937 cells

Apoptosis

induction = faster

with

α-hederin

than

hederagenin

+ Q-VD-O-Ph

(general

caspase

inhibitor) inhibition of apoptosis

for both

compounds no inhibition of non-apoptotic

cell

death

Non-apoptotic

cell

death

(very

important at

high

α-hederin

concentrations)

Apoptotic(fragmented

nuclei)

Membrane

damage(Ethidium

bromide)

No membrane Damage

(acridine orange)

Non-apoptoticAO/EB staining

0 20 40 600

25

50

75

100C

time (h)

% c

ell d

eath

0

10

20

30

40A

% fr

agm

ente

d nu

clei

0 20 40 60

D

time (h)

B CTL10 µM15 µM20 µM40 µM


Cell

death

: Acridine orange/Ethidium

bromide

in U937 cells

Apoptosis

induction = faster

with

α-hederin

than

hederagenin

+ Q-VD-O-Ph

(general

caspase

inhibitor) inhibition of apoptosis

for both

compounds no inhibition of non-apoptotic

cell

death

Non-apoptotic

cell

death

(very

important at

high

α-hederin

concentrations)

0 5 7.5 10 12.5 15 17.5 200

10

20

30

***

***

% f

ragm

ente

d nu

clei

0.0 5.0 7.5 10.0 12.5 15.0 17.5 20.0

* *** ******

******

**

Non-depletedCholesterol-depleted24h 48h

Cholesterol

dependence

of total cell

death

/ apoptosis

in U937

Cholesterol

dependence

of cell

death

: decrease

of α-hederin

induced

total celldeath

and apoptosis

in depleted

cells

α-hederin

Non-depleted cells

0 5 10 15 20 25 300

25

50

75

100

******

******

*** *** ***

time (h)

% c

ell d

eath

Cholesterol depleted cells

0 5 10 15 20 25 30

control10 µM15 µM20 µM25 µM

time (h)

two-way

ANOVA*p<0.05, **p<0.01, ***p<0.001vs. depleted

cells

Trypan

blue

staining

(cell

death

assay)

Microscopedeath

cell

Fluorescent DNA intercalator

DAPI (apoptosis

assay)

apoptoticnormal

Cell

nuclei

(fluorescence microscopy)

accumulates

only

in death

cells

0 5 7.5 10 12.5 15 17.5 200

10

20

30

***

***

% f

ragm

ente

d nu

clei

0.0 5.0 7.5 10.0 12.5 15.0 17.5 20.0

* *** ******

******

**

Non-depletedCholesterol-depleted24h 48h

Cholesterol

dependence

of total cell

death

/ apoptosis

in U937

Cholesterol

dependence

of cell

death

: decrease

of α-hederin

induced

total celldeath

and apoptosis

in depleted

cells

α-hederin

Non-depleted cells

0 5 10 15 20 25 300

25

50

75

100

******

******

*** *** ***

time (h)

% c

ell d

eath

Cholesterol depleted cells

0 5 10 15 20 25 30

control10 µM15 µM20 µM25 µM

time (h)

two-way

ANOVA*p<0.05, **p<0.01, ***p<0.001 vs. depleted

cells

Conclusions : Potential

interactions

α-hédcholestérol

mitochondria

cyt

c

Caspase-3

apoptosisDR

MPT

cellnecrosis

Ca2+(*)

*Choi, J.H. et al. 2008**Swamy

and Huat

2003

Caspase-8(*)

membrane lysis

(*)

(*,**)

(*)

Effect of surfactin

on membrane models displaying lipid phase separation

Magali Deleu1°*, Joseph Lorent2°, Laurence Lins3, Robert Brasseur3, Nathalie Braun4, Karim El Kirat4, Tommy Nylander5, Yves F. Dufrêne4

and Marie-

Paule Mingeot-Leclercq2

1Université

de Liège Gembloux Agro-Bio Tech, Unité

de Chimie Biologique Industrielle, Passage des Déportés, 2, B-5030 Gembloux, Belgium

2Université


Institute, Cellular and Molecular

Pharmacology, Avenue E. Mounier 73, B1.73.05, B-1200, Brussels, Belgium

3Université

de Liège Gembloux Agro-Bio Tech, Centre de Biophysique Moléculaire Numérique, Passage des Déportés, 2, B-5030 Gembloux, Belgium

4Université

catholique de Louvain, Institute of Condensed

Matter

and Nanosciences, Bio and Soft Matter, Croix du Sud 1, L7.04.01, B-1348 Louvain-la-

Neuve, Belgium

5Lund University, Center

for Chemistry and Chemical Engineering, Physical Chemistry, 1S-221 00 Lund, Sweden°Equal First author

BBA, Biomembranes, 2013, 1828, pp. 801-815

Molecular

structure, CMC and activity

of surfactin

CMC = 12.85 ±

0.05 µM

(pH = 8.5)

HN

HN

O

O

NH

HN

OOH

O

O

HN

HNO

O

NH

O

O

O

OH

O

(determined

by isothermal

titration

calorimetry)

C13

, C14

, C15

linear

acyl

chain

Tested

on DOPC/DPPC

(1:1) bilayer

at

pH =8,5

0 min 22 min 28 min

42 min 68 min

3µM

180 min

120 min

60 min20 min0 min

15µM

0 min 12 min 31 min 60 min 120 min

1mM

15 x 15 µm 20 x 20 µm

z-range10 nm

Integrity

of SPB : AFMSupported

planar

bilayer

: DOPC/DPPC (1:1) (25°C)

~ CMC

> CMC

< CMC

DPPC

DOPC

Permeabilization

and remodelization

of LUV: calcein

release, R18

dequenching

and dynamic light scattering

(DLS)

3 µM15 µM25 µM100 µM

)exp1()exp1(% 21 2max

1max

tktkreleased yycalcein CMC

Calcein

release R18 dequenching

Dynamic

light scattering

Permeabilization

and remodelization

of LUV: calcein

release, R18

dequenching


(DLS)



1max

tktkreleased yycalcein

< CMC

CMC

Permeabilization

and remodelization

of LUV: calcein

release, R18

dequenching


(DLS)



1max


~ CMC

CMC

Permeabilization

and remodelization

of LUV: calcein

release, R18

dequenching


(DLS)



1max


Two

release mechanisms

> CMC

CMC

Conclusions : Surfactin

/ membrane interaction

[surfactin] < CMC

[surfactin] ~ CMCSolubilization

of DOPC, pore formation

Erosion of DPPC domains

and transient

defects

Red

= surfactin

[surfactin] >> CMC

Transformation into

micelles, Fusion ?

Conclusions : Surfactin

/ membrane interaction

General conclusions

HN

HN

O

O

NH

HN

OOH

O

O

HN

HNO

O

NH

O

O

O

OH

OCH2OH

COOH

O

HO

H

H

HO

HOH

HH

O

OH

H

OH

H

H

H

OCH3

H

O

HCH2OH

COOH

HO

Membrane raft receptors

(Fas,.) ?

Other

receptor

dependentactivities

General conclusions

HN

HN

O

O

NH

HN

OOH

O

O

HN

HNO

O

NH

O

O

O

OH

OCH2OH

COOH

O

HO

H

H

HO

HOH

HH

O

OH

H

OH

H

H

H

OCH3

H

O

HCH2OH

COOH

HO

Perspectives

Short term-perspectives

Cell

staining

(McIntosh 2008, Garvik

2009)GUV : DOPC:DPPC:DHE (1:1:1)

Biphoton

microscopy

of DHE

Tof-SIMS

SPB (Lorent)Cells(Mazzucchelli

2008)

Perspectives

Long term-perspectives

CH2OH

COOH

O

HO

H

H

HO

HOH

HH

O

OH

H

OH

H

H

H

OCH3

H

O

H

Hemisynthesis

HemolysisActivity

on cancer cells

New anticancer

drugs

?

HN

HN

O

O

NH

HN

OOH

O

O

HN

HNO

O

NH

O

O

O

OH

O

CH2OH

COOH

HO (QSAR)

New nanoparticles

Membrane interactions of lipopeptides and saponins, two ... · PDF filelipopeptides and saponins, two groups of natural amphiphiles J. Lorent (public defence) Main supervisor: M-P.

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