Acyl group: c16 or c18, 0, 1 or 2 double bond
choline head
PI
PL = glycerol attached to 2 FA phosphate and different side groups
(PE, PS, PC) SM = serine attached to 2FA phosphate and choline side
group PI = minor phospholipid critical for signaling; inositol ring
can be phosphorylated Cholesterol = complex hydrocarbon ring
structure
Due to the amphipathic nature of phospholipids, these molecules
spontaneously assemble to form
closed bilayers
Different cell type → different composition of lipids
4
Record
Fluorescent recovery after photobleaching
Thermal effect and dependent Two dimensional plane of a bilayer
move rotate “The Fluid Mosaic Model”
FRAP:Fluorescent Recovery After Photobleaching
Why does membrane need to be fluid?
Enables rapid diffusion of membrane proteins within plane of
bilayer and permits interaction (important for cell
signalling)
Facilitates distribution of membrane lipids and proteins from
insertion site (following synthesis) to other regions of cell
Allows membranes to fuse and mix molecules
Ensures even distribution of membrane molecules between daughter
cells following division
Lipid composition influence physical properties of membrane:
1. Different composition of organs
2. Specialized membrane function
sphingolipids: phosphoglycerides: cholesterol
basolateral 0.5 1 1
apical 1 1 1
3. Affects membrane fluity a. short C-H chain are more fluid b.
kinks in C-H: less stable
4. Influence thickness of membrane
5. Local curvature ()
the size of the molecule
its interactions with other molecules
temperature
Cholesterol →Membrane fluidity↓ In animal cells, cholesterol used
to modulate membrane fluidity - fills gaps between kinks of
unsaturated tails
Used particularly in plasma membrane ⇒ closer packing ⇒ less
fluidity/permeability
phospholipid cholesterol
Membrane fluidity
Membrane fluidity important for membrane function; determined by
phospholipid composition
Close packing of hydrocarbon tails ⇒ less fluidity (increased
viscosity)
Length and unsaturation (no of double bonds) determine closeness of
packing
Length varies from 14-24 C atoms; shorter chain length ⇒ less
interaction ⇒ increased fluidity
One tail of molecule has one or more double bonds - unsaturated (H
atoms); other tail has no double bonds - saturated
Double bonds ⇒ kinks () ⇒ less packing
double bond -kink
Two layers of bilayer have different compositions
- different phospholipid/glycolipid inside vs outside
- membrane proteins embedded into membrane with specific
orientation
Lipid bilayer asymmetry (mechanism?)
Phosphatidylcholine (PC) Sphingomyelin (SM)
Membrane synthesis occurs in endoplasmic reticulum (ER)
New membrane exported to other membranes by vesicles (budding and
fusion)
Lipid asymmetry occurs during manufacture To permit membrane
growth, newly synthesised membrane must be
evenly () distributed between both monolayers Asymmetry
distribution of bi-layer, Requires enzyme assistance -
flippases Flippases selectively transfer specific phospholipids ⇒
asymmetric
distribution in each monolayer
Flip-flop would require the polar head-group of a lipid to traverse
the hydrophobic core of the membrane. Need large energy
The two leaflets of a bilayer membrane tend to differ in their
lipid composition.
Flippases catalyze flip-flop in membranes where lipid synthesis
occurs.
Some membranes contain enzymes that actively transport particular
lipids from one monolayer to the other.
Flip Flop
Flip-flop of lipids (from one half of a bilayer to the other) is
normally very slow.
Membrane lipids are usually distributed unequally in the exoplasmic
an cytosolic leaflets
Membrane is an asymmetry in lipid composition across the
bilayer
But cholesterol is relative evenly distributed
Phospholipase can regulated the composition of phospholipid in
membrane. It can cut off the hydrophobic tail and can not across
membrane.
Membrane asymmetry Affects: Enzyme cleavage •phospholipase cleaves
phospholipids at exoplasmic sides
• cytosolic sides are resist to phospholipase cleavage
Specific of phospholipases
An membrane surface
Lipid rafts: Sphingolipids (particularly glycosphingolipids) in the
plasma membrane
outer leaflet tend to separate out from glycerophospholipids, &
co- localize with cholesterol in microdomains called lipid
rafts.
Lipid rafts are resistant to detergent solubilization, which has
facilitated their isolation and characterization.
Close packing of sphingolipids in association with cholesterol has
been attributed to lack of double bonds in sphingolipid hydrocarbon
chains. Glycerophospholipids often include at least one fatty acid
that is kinked, due to one or more double bonds.
Proteins with covalently attached lipid anchors (fatty acid or GPI)
tend to associate with raft domains.
Biomembranes: protein components and basic functions Proteins
interact with membrane in three different way:
Integral protein: three part and across membrane, has hydrophobic
and hydrophlic part
Lipid anchored: can not across membrane. It bound to one or more
lipid molecules.
Peripheral protein: can not interact with the hydrophobic core.
Indirectly bound to membrane, via integral protein connect to
membrane or cell. May has support
Hydrophobic interact with hydrophobic
1. Integral membrane protein( transmembrane protein) a. exoplasmic
domain
cytosolic domain hydrophilic
c. glycosylated
Membrane-embedded α helices are the primary secondary structures in
most transmembrane protein
Membrane protein: Helices can exposure the hydrophobic
residue
It can interact with membrane hydrophobic part
α-helices
α-helices:
Interact with fattyacyl of lipid by van- der-waals
Gly: most small Green par: hydrophobic amino acid Blue: hydropholic
amino acid
Structural model of bacteriorhodopsin, a multipass (7)
transmembrane protein that functions as a photoreceptor in certain
bacteria. Like G-coupled receptor
Retinal molecule
Multiple β strands in porins () form membrane-spanning
“barrels”()
Porins: Integral membrane protein Permit the uptake and disposal of
small
hydrophilic molecules, has regulation and prevent chemical
damage…
Inside is hydropholic part Outside is hydrophobic part All porins
are trimeric transmembrane
protein
Hydrophobic part
Can pass chemical Integrate to membrane
Modes of attachment to: Cytosolic leaflet: fatty acyl group (e.g.
myristate or palmitate) attached to the N-terminal glycine
residue unsaturated fatty acyl (farnesyl or geranylgeranyl) group
attached by thioether
bond to C-terminal cysteine. In some cases a second fatty acyl
group is linked to another cysteine.
Exoplasmic leaflet: Glycosylphosphatidylinositol (GPI) anchor:
phosphatydilinositol (PI): two fatty acid chains inserted in
membrane several sugar residues phosphoethanolamine: links to
C-terminus of protein
Lipid-anchored membrane proteins PE
Human ABO blood-group antigens
Human RBC type depend on the surfaces expressed different
glycoproteins and glycolipids
All human had enzyme for synthesis O antigen.
A type human: has GalNAc transferase
B type human: has Gal transferase AB type human: both enzyme
has
Peripheral membrane proteins
More loosely associated with the membrane. Usually non-
covalentlyattached to protruding portions of integral membrane
proteins.
Examples include signalling molecules such phospholipase C and
protein kinase B as that bind to the cytoplasmic membrane side to
PIP2 or PI3 via a specialised protein domain, the pleckstrin
homology domain
Interaction with cytoskeleton impede () the mobility of integral
membrane protein
Lipid-binding motifs help target peripheral protein to the membrane
Motility of membrane protein 1. Float freely 2. Immobile 3.
Anchored by cytoskeletal protein
11
Inter-facial binding surface and mechanism of action of
phospholipase A2 It can degradation of damage or aged cell
membrane
Blue: arginine and lysine (+), surrounding of the catalytic
site
Functions of the plasma membrane
Regulate transport of nutrients into the cell Regulate transport of
waste out of the cell Maintain “proper” () chemical conditions in
the cell Provide a site for chemical reactions not likely to occur
in
an aqueous environment Detect signals in the extracellular
environment Interact with other cells or the extracellular
matrix
(in multicellular organisms)
Effect of external ion concentration on water flow across the
plasma membrane of an animal cell
However, the pant cell has cellulose (cell wall) can prevented the
swelling or shrinking
Organelles of the eukaryotic cell
• Lysosomes • Peroxisomes • Mitochondria • Chloroplasts • the
Endoplasmic Reticulum (ER) • the Golgi complex • the Nucleus • the
Cytosol
12
Animal cell structure
Plant cell structure
13
Endosomes take up soluble macromolecules from the cell exterior
Lysosome are acids organelles that contain a battery of
degradative
enzyme Lysosome:
• Responsible for degrading certain cell components material
internalized from the extracellular environment
• Key Features – single membrane – pH of lumen ≅ 5 – acid
hydrolases carry out
degradation reactions
Vary in size and shape: primary lysosomes (about spherical and do
not contain oby), secondary lysosomes (large and irregularly
shaped)
Hydrolytic enzymes degrade proteins, nucleic and other large
molecules
Cellular structure that participate in delivering materials to
lysosome
1. Endocytic pathway: soluble macromolecules into the cell 2.
Phagocytosis:Whole cell and other large insoluble particles 3.
Autophagy: organelles and cytoplasmic substance.
Mammaliam cell Ovalbumin (black spot) is found in early endosome
(EE) and late endosome (LE), but very little present in
autophagosomes (AV)
Rat liver cell Second lysosome containing fragment of mitochondria
(M), and peroxisome (P)
14
Peroxisomes
• Key Features – single membrane – contain oxidases and
catalase
Oxidize toxic molecules, oxidase Catalase, degrade hydogen
peroxide. Oxidation of fatty acid, generate acetyl groups.
The oxidation of fatty acids in Mitochondria: produceds CO2 and ATP
Peroxisomes: no ATP
Tansport to cytosol and the synthesis of cholesterol
H2O2 → catalase → H2O + O2
The endoplasmic reticulum (ER)
Responsible for most lipid synthesis most membrane protein
synthesis Ca++ ion storage detoxification
Key Features – network of interconnected
closed membrane tubules and vesicles
– composed of smooth and rough regions
Smooth ER: synthesis of fatty acids and phospholipids; no ribosome;
detoxify and modify chemistry
Rough ER: ribosome bound, synthesis of secreted and membrane
proteins
Cisternae: an extensive network of closed, flatted membrane-
bounded sacs
• Modifies and sorts most ER products
• Key Features – series of flattened
compartments & vesicles – composed of 3 regions:
cis (entry), medial, trans (exit) – each region contains
different
set of modifying enzymes
The golgi complex processes and sorts secreted and membrane
protein
1. Transport vesicles to golgi complex from RER 2. Concentrated and
packaged into immature secretory 3. Vesicles accumulate 4.
exocytosis
15
Model of the Golgi complex based on three-dimensional
reconstruction of electron microscopy. White: transport vesicle;
orange and red: trans-Golgi membrane; blue: vesicles have budded
off the RER fuse with cis-membrane.
• Site of photosynthesis in plants and green algae
• Key Features – outer membrane – intermembrane space – inner
membrane – stroma – thylakoid membrane – thylakoid lumen
Bone marrow stem cell
• Separates – DNA from cytosol – transcription from
translation
• Key Features – outer membrane – inner membrane – nuclear pores –
Nucleolus – nucleoplasm
The nucleus contains the DNA genome, RNA synthetic apparatus and a
fibrous matrix
nucleus
16
Occupy up to 25% of the volume of cytoplasm
The two membrane has different composition and function; out
membrane composed of about half lipid and half protein and has
porins. Inner membrane less permeable, about 20% lipid and 80%
protein, has cristae to increase the area.
Key Features – outer membrane – intermembrane space – inner
membrane – matrix
Mitochondria are the principal sites of ATP production in aerobic
cells
Site of photosynthesis in plants and green algae
Key Features – outer membrane – intermembrane space – inner
membrane – stroma
Chloroplasts contain internal compartments in which
photosynthesis
Thylakoids are flattened to form disks → formed stacks (called
grana) → embedded in matrix → stroma; contain green pigments,
generated ATP during photosynthesis
The apical part of a detergent extracted intestinal epithelial
cell
The cytoskeleton: components and structural functions
About ¼- ½ protein in cytosol Most water soluble protein
bound
to filament or localized in specific regions
Triton X-100 can not extract the cytoskeleton and organelles.
Three types of filaments compose the cytoskeleton
Microfilaments (actin filaments;): diameter 8-9nm; actin assemble
into Microtubules (MT): diameter 24nm; α and β-tubulin
polymerization Intermediate filaments (IF): diameter 10nm
17
Actin filament: microfilament, 8~9nm, two strains of actin
Intermediate filament: 10nm, keratin, desmin, vimentin, lamin
Microtubule: α and β tubulin, protofilament, 24nm hollow tube
A mixture of actin filaments , microtubules and vimentin
intermediate filaments
Cytoskeletal filaments are organized into bundles and
networks
Bundles and networks are the most common arrangements of
cytoskeletal filament in a cell
Bundles, the filaments are closely packed in parallel arrays;
network, the filament crisscross
Cortical cytoskeleton supporting the plasma membrane in human
RBC
Association of the Erythrocyte Cortical Cytoskeleton with the
Plasma Membrane
the ERM proteins
The major protein that provides the structural basis for the
cortical cytoskeleton in erythrocyts
Figure legend: The plasma membrane is associated with a network of
spectrin tetramers corsslinked by short actin filaments in
association with protein 4.1. The spectrin- actin network is linked
to the membrane by ankyrin (major), which binds to both spectrin
and band 3. An additional link is provided by the binding of
protein 4.1 to glycophorin (secondly). The major protein that
provides the structural basis for the cortical cytoskeleton in
erythrocyts is the actin-binding protein spectrin (spectrinbind
actin). Spectrin is a member of the large calponin family of acting
binding proteins, which includes a-actinin and fimbrin.
Intermediate filaments support the nuclear membrane and help
connect into tissue
Desmosome: between cells Hemidesmosome: cell and matrix Nuclear
lamina: laminin, anchored to the inner nuclear membrane
IF crisscross the cytosol from the nuclear envelop to the plasma
membrane; It provides mechanical support.
Nuclear lamina: lamin A and lamin C filaments and associated with
lamin B
Fluorescence micrograph of a PtK2 fibroblast cell stained to reeal
keratin intermediate filament
18
MTOC: microtubule organizing center Associated with ER and other
organelles Formation of mitotic apparatus
Flow cytometry separates different cell types
By cell marker DNA dye
Flow Cytometry and Cell Sorting
‘FACS’ has become a generic term for ALL flow cytometry FACS :
Fluorescence Activated Cell Sorter Is actually a trade name of
Becton Dickinson (BD).
FACS is one version of Flow Cytometry, which can sort cells by
their surface markers
Individual cell is positively or negatively charged based on their
fluorescence color
When charged cells pass through an electric field, they are
deflected and hence separated
0.1 1 10 100 1000 0.1
1
10
100
1000
Double Negative
die