Biochemie 2

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Biochemie 2 Prof. Gerrit van Meer Membrane
Enzymology Bijvoet Center, Institute of
Biomembranes Dept. of Chemistry College
1 "Stryer" Chapter 12 Lipids and Cell Membranes
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All living cells protect themselves from the
environment by a wall This allows them to do
their chemistry in a secluded environment The
wall consists of lipids and proteins In all
kingdoms of life, eubacteria, archaeae and
eukaryotes, this wall is a thin layer of oil a
lipid membrane This layer is amazingly stable
considering that it is only 4 nm thick compared
to a cell diameter of 20 µm, 20 x 103 nm 0.2 mm
on 1 m
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Cytosol Cytoplasm Nucleoplasm
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Dynamics Secretion Endocytosis
G
Lysosome
ER
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Bidirectional vesicular traffic between organelles
TGN
E
G
L
ER
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Hydrocarbons are very simple molecules
H
3
C4 Butane, gas C8 Octane, liquid C20 and
higher Paraffins, solid. Because these molecules
aggregate in water (aqueous environment), they
are relatively useless for life They were present
early in evolution
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Hydrophobic
C18
C36
C36
Hydrophilic
C18
C18
Hydrocarbon with terminal carboxy-group
Molecules with a polar end AND a hydrophobic end
are termed "amphipathic". In water they will
orient their polar part towards water, their
hydrophobic part towards eachother.
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The basic structure of biomembranes, the lipid
bilayer
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The folded protein has a lower entropy (is more
ordered). Still, the reaction runs
spontaneously. But according to the second law
of thermodynamics the entropy for a spontaneous
reaction must increase!
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Proteins contain hydrophobic side chains. Water
molecules cant interact with the hydrophobic
surface and will adopt an ordered orientation
around the hydrophobic moiety (caged water).
When the hydrophobic surfaces bind to each
other, the ordered water is released and the
total entropy increases!
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The hydrophobic effect plays an important role in
protein- protein interactions. In addition, the
strength of a polar bond is greater in a
hydrophobic environment (water has a dielectric
constant of 80 a polar bond is 80 x weaker in
water than in vacuum)
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Glycerolipids
glycerophospholipids phosphatidylcholine
choline (PC) ethanolamine (PE) serine, inositol
(PS, PI)
P
phosphate
O
O
glycerol
O
O
O
O
fatty acid
C16 C181 C204 palmitic oleic acid
arachidonate solid fluid very fluid
65 mol
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Glycerolipids
Sterols
Sphingolipids
sphingomyelin glycosphingolipids
glycerophospholipids PC, PE, PS, PI
P
G
glucose
O
O
glycerol
sphingosine
cholesterol
O
O
O
OH
O
N
OH
OH
O
fatty acid
fatty acid
65 mol 10 25 cholesterol makes
all membranes fluid
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Fatty acids organize themselves as micelles at
the critical micellar concentration
Phospholipids organize themselves as bilayers
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P
Phospholipid
Triacylglycerol
Cholesterolester
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P
20 nm
Apo-A1
HDL
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OH
PC PE SM chol
van Meer EMBO J., 2005
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2 nm
LPC
PE
PC
60-100 nm
4 nm
Lipids have a molecular shape that suits specific
processes, for example vesicle budding
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The potential transport routes of a lipid between
two membranes
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Physical properties of some lipids
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Fluorescent lipids / lipid analogs
N-Rhodamine-PE
C12-NBD-SM
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Kuerschner et al., Nature Methods 2005 van Meer
and Liskamp, Nature Methods 2005 (comment)
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In Eukaryotes and Bacteria the membrane is a
lipid bilayer
Cytosol
Cytosol
Cellular membranes have a cytosolic side and
a non-cytosolic side. They are asymmetric
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Living cells protect themselves from the
environment by a thin layer of oil a lipid
membrane, the lipid bilayer
Cytosolic side
Intracellular membranes originate from an
invagination Cellular membranes have a cytosolic
side and a non-cytosolic side They are
inherently asymmetric
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Bidirectional vesicular traffic between organelles
TGN
E
G
L
ER
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Adaptor proteins and coats help to include
specific proteins into specific budding
vesicles. Where does the energy for budding come
from?
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Specificity in vesicular transport requires
specific inclusion of cargo with correct SNARE
SNARES provide specificity in targeting, docking
and fusion
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cytosol
In the cell fusion machinery cytosolic Virus
fusion non-cytosolic side
cytosol
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PC
SM
GlcCer
cholesterol
Plasma membrane
Golgi
PS
PE
ER
PC
PE
Cellular membranes differ in 1. lipid composition
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Surface polarity of membrane lipids in
epithelial cells
Apical
GSL PL Chol 33 33 33 8 66 26
Basolateral
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Sphingolipid polarity in epithelial cells Tight
junctions barrier in exoplasmic leaflet Lipid
differences in exoplasmic leaflet
Apical
Golgi
endoplasmic reticulum
Basolateral
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Apical
Golgi
Basolateral
Hypothesis (van Meer and Simons)
Glycosphingolipids aggregate and separate from
the glycerophospholipid PC How do the correct
Snares recognize the two different lipid phases?
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Phase diagram de Almeida et al. (2003) Biophys.
J. 85, 2406
Cholesterol
Coexistence of two different liquid phases Liq.
ordered Liq. disordered at equimolar concentration
lo
C
C
P
P
O
O
O
O
O
O
N
OH
O
ld lo
ld lo so
ld
ld so
POPC
PSM
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10 µm
This happens but om a scale much larger than
vesicle budding (100nm Ø) Baumgart, Hess and
Webb, Nature 425 (2003) 821
Lipowsky and Dimova (2003) J. Phys. Condens.
Matter 15, S31 A system with 2 phases will try to
shorten the phase boundary in order to reduce
the line tension
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Roux et al., EMBO J. 2005 40 nm tubes drawn
from homogeneous SM/chol/PC (111) are enriched
in PC
1 sphingomyelin SM 2 Cholesterol 3
Phosphatidylcholine PC
van Meer
EMBO Reports, May 2005
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Detergent-resistant membranes
Brown and Rose, Cell 1992
Sphingolipids Disaturated phospholipids Cholestero
l GPI-anchored proteins 140/160 acylated
proteins
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G
Endosome
Lysosome
ER
N
Endocytosis results in the uptake of membrane
lipids
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Segregation of Tfn (green back to the cell
surface) and EGF (red to the lysosomes) after
endocytosis yellow colocalization. Sharma et
al (2003) JBC 278, 7564
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The fluorescence emission of this analog shifts
from green to red at high concentrations
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Concentration of Bodipy-LacCer in endosome
subdomains (green low concentration red high
concentration) Sharma et al (2003) JBC 278, 7564
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Membranes can contain lateral domains
GPI-anchored proteins for example prion
protein 140/160 acylated for example
signaling kinases
15-17 20-23 aa
Domains can be recognized by proteins
The physical basis for the lateral heterogeneity
lies in the preferential interactions between
the sphingolipids. Due to their long saturated
fatty acid chains they form a thicker
membrane. Some proteins prefer the thicker part
of the membrane because they have longer
transmembrane domains consisting of amino acids
with hydrophobic side chains. For unknown
reasons, also proteins with lipid tails have a
preference for ordered domains......
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After stimulation receptors oligomerize and lipid
rafts are stabilized
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Lipid rafts in biomembranes Lipid-protein
aggregates that depend on lipid-lipid
interactions Rafts appear small and transient
unless stabilized Rafts in transport
induced/stabilized by an increase in order
by curvature (Roux et al. 2005, EMBO J) by
caveolin, ceramide How many types? Rafts on the
cytosolic surface? How are rafts involved in
protein sorting/signaling? How do proteins
recognize rafts? Lipid tails, membrane
thickness, other physical principles?
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Jacobson et al. (2007) Nature Cell Biol. 9, 7. Do
lipid rafts form around proteins? Answer This is
possible. However, lipid rafts depend on
preferential lipid-lipid interactions
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Takamori et al.(2006) Cell 127, 831.
40 nm