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Lipids & Membranes
Prepared by LLT
Biochemistry of Metabolism
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Lipids are non-polar (hydrophobic) compounds,soluble in organic solvents.
Most membrane lipids are amphipathic, having a
non-polar end and a polar end.
Fatty acids consist of a hydrocarbon chain with a
carboxylic acid at one end.
A 16-C fatty acid: CH3(CH2)14-COO-
Non-polar polar
A 16-C fatty acid with one cis double bond between
C atoms 9-10 may be represented as 16:1 cis (9.
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Some fatty acids and their common names:
14:0 myristic acid; 16:0 palmitic acid; 18:0 stearic acid;
18:1 cis(9 oleic acid
18:2 cis(9,12 linoleic acid
18:3 cis(9,12,15 E-linonenic acid
20:4 cis(5,8,11,14 arachidonic acid
20:5 cis(5,8,11,14,17
eicosapentaenoic acid (an omega-3)
Double bonds in fattyacids usually have the
cis configuration.
Most naturallyoccurring fatty acidshave an even number
of carbon atoms.
C
O
O
1
2
34
EF
K
atty acid ith a cis-(9
double bond
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There is free rotation about C-Cbonds in the fatty acidhydrocarbon, except where there is a double bond.
Each cis double bond causes a kinkin the chain.
Rotation about otherC-Cbonds would permit a morelinear structure than shown, but there would be a kink.
C 1234
EF
fatty acid with a cis-(9
double bond
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Glycerophospholipids
Glycerophospholipids
(phosphoglycerides), are commonconstituents of cellular membranes.
They have a glycerolbackbone.Hydroxyls at C1 & C2 are esterifiedto fatty acids.
C HH
CH2 H
CH2 H
glycerol
An ester formswhen a hydroxylreacts with acarboxylic acid,with loss of H2O.
Formation of an ester:
O O
R'OH + HO-C-R" R'-O-C-R'' + H2O
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Phosphatidate
In phosphatidate:
fatty acids are esterified to hydroxyls on C1 & C2
the C3 hydroxyl is esterified to Pi.
O P O
O
O
H2C
CH
H2C
OCR1
O O C
O
R2
phosphatidate
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In most glycerophospholipids (phosphoglycerides),
Pi is in turn esterified to OH of a polar head group (X):serine, choline, ethanolamine, glycerol, or inositol.
The 2 fatty acids tend to be non-identical. They may differ
in length and/or the presence/absence of double bonds.
O P O
O
O
H2C
CH
H2C
OCR1
O O C
O
R2
X
glycerophospholipid
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Phosphatidylcholine, with choline as polar head
group, is an example of a glycerophospholipid.
It is a common membrane lipid.
O P O
O
O
H2C
CH
H2C
OCR1
O O C
O
R2
CH2 CH2 N CH3
CH3
CH3
+
phosphatidylcholine
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Phosphatidylinositol, with inositol as polar head group,is another glycerophospholipid.
In addition to being a membrane lipid,
phosphatidylinositol has roles in cell signaling.
O P
O
O
H2C
CH
H2C
OCR1
O O C
O
R2
OH
H
OH
H
H
OHH
OH
H
O
H OH
phosphatidyl-inositol
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polar
non-polar
"kink" due to
double bond
O P O
O
O
H2C
CH
H2C
OCR1
O O C
O
R2
X
glycerophospholipid
Each glycerophospholipidincludes
a polar region:
glycerol, carbonyl of
fatty acids, Pi, & thepolar head group (X)
2 non-polar hydrocarbon
tails of fatty acids (R1, R2).
Such an amphipathic lipid
may be represented as at
right.
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The amino group of sphingosine canform an amide bond with a fatty acid
carboxyl, to yield a ceramide.
Ceramides usually include a polarhead group, esterified to the terminalOH of the sphingosine.
H2CHC
OH
CH
N+ CH
C
CH2
CH3
H
H3
OH
( )12
sphingosine
H2CHC
OH
CH
NH CH
C
CH2
CH3
H
OH
( )12
C
R
O
ceramide
Sphingolipids are derivatives of thelipid sphingosine, which has a long
hydrocarbon tail, and a polar domainthat includes an amino group.
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Sphingomyelin, with a phosphocholine head group, is
similar in size and shape to the glycerophospholipid
phosphatidyl choline.
Sphingomyelin, a
ceramide with a
phosphocholine or
phosphethanolamine
head group, is a
common constituent
of plasma membranes
H2CHC
O
CH
NH CH
C
CH2
CH3
H
OH
( )12
C
R
O
PO O
O
H2C
H2CN
+
CH3
H3C
CH3
Sphi li
phosphocholi
sphi osi
tt ci
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head group that is a complex oligosaccharide, including
the acidic sugar derivative sialic acid.Cerebrosides and gangliosides, collectively calledglycosphingolipids, are commonly found in the outerleaflet of the plasma membrane bilayer, with their sugarchains extending out from the cell surface.
cerebroside withF-galactose head group
H2CHC CH
NH CH
C
CH2
CH3
OH
C
R
O
OH O
H H
H
OHH
OH
CH2OH
HO
H
( )12
A cerebroside is asphingolipid
(ceramide) with amonosaccharide
such as glucose orgalactose as polar
head group.A ganglioside is aceramide with a polar
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liquid crystal crystal
In the liquid crystal state, hydrocarbon chains of
phospholipids are disordered and in constant motion.At lower temperature, a membrane containing a singlephospholipid type undergoes transition to a crystallinestate in which fatty acid tails are fully extended, packingis highly ordered, & van der Waals interactions betweenadjacent chains are maximal.
Kinks in fatty acid chains, due to cis double bonds,interfere with packing in the crystalline state, and lowerthe phase transition temperature.
Membrane fluidity:
The interior of a lipid bilayeris normally highly fluid.
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Cholesterol is largely hydrophobic.But it has one polar group, a hydroxyl, making itamphipathic.
CholesterolHO
Cholesterol, animportant constituentof cell membranes,has a rigid ring
system and a shortbranchedhydrocarbon tail.
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Cholesterolin membrane
Cholesterol inserts into bilayer
membranes with its OH oriented
toward the aqueous phase & its
hydrophobic ring system adjacent tofatty acid chains of phospholipids.
The hydroxyl group of cholesterol
forms hydrogen bonds with polarphospholipid head groups.
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The presence ofcholesterol in a phospholipid
membrane inhibits transition to the crystalline state.
However interaction with the relatively rigidcholesterol decreases the mobility of hydrocarbon tails
of phospholipids.
Phospholipid membranes that include cholesterol have
a fluidity intermediatebetween the liquid crystal and
crystal states.
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Two strategiesby which phase changes of membranelipids are avoided:
Cholesterol is abundant in membranes, such asplasma membranes, that include many lipids with
long-chain saturated fatty acids.In the absence of cholesterol, such membranes wouldcrystallize at physiological temperatures.
The inner mitochondrial membrane lacks cholesterol,
but includes many phospholipids whose fatty acidshave one or more double bonds, which lower themelting point to below physiological temperature.
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Lateral mobility of a lipid, within the plane of a
membrane, is depicted above and in an animation.
Lateral diffusion of membrane lipids and proteins isassayed by the FRAP technique:
Fluorescence Recovery AfterPhotobleaching.
Lateral obility
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FRAP technique:
A membrane lipid or protein is tagged with afluorescent dye. Proteins may be labeled withfluorescent antibodies.
A laser bleaches (destroys fluorescence of) the dye
in a small region of membrane.
Fluorescence recovers as undamaged label diffusesinto the region.
Generally lipids diffuse faster than proteins.Lateral diffusion of some proteins is constrainedby protein-protein interactions, e.g. formation oflarge complexes or association with the cytoskeleton.
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Flip-flop of lipids (from one half of a bilayer to the other)is very slow.
Flip-flop would require the polar head group of a lipid totraverse the hydrophobic core of the membrane.
Flippases catalyze flip-flop in membranes where lipidsynthesis occurs. Otherwise, flip-flop of lipids is rare.
Consistent with low rates of flip-flop, the 2 leaflets of abilayer tend to differ in lipid composition.
Flip Flop
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Lipid rafts:
Biological membranes typically include a mix of lipids.
Sphingolipids in the plasma membrane outer leaflet tendto separate out from glycerophospholipids, & co-localizewith 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 bondsin sphingolipid hydrocarbon chains.
Glycerophospholipids often include at least one fatty acidthat is kinked, due to having one or more double bonds.
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Caveolae are invaginated lipid raft domains of theplasma membrane.
Caveolin, a protein associated with the cytosolic
leaflet of the plasma membrane in caveolae, interactswith cholesterol.
Electron micrograph of caveolae (on home page of D. Brown)
caveolae
cytosol
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Peripheral proteins are on the membrane surface.
They are water-soluble, with mostly hydrophilic surfaces.
Often peripheral proteins can be dislodged by conditionsthat disrupt ionic & H-bond interactions, e.g., extractionwith solutions containing high concentrations of salts,change of pH, and/or chelators that bind divalent cations.
Membrane
proteins may beclassified as:
peripheral
integral
having alipid anchor
integral
lipid
anchor
peripheral
lipid bilayer
embraneroteins
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Some cytosolic proteins have domains that bind to polarhead groups oflipids that transiently exist in a membrane.
The enzymes that create or degrade these lipids are subjectto signal-mediated regulation, providing a mechanism formodulating affinity of a protein for a membrane surface.
O P
O
O
H2C
CH
H2C
OCR1
O O C
O
R2
OH
H
OPO32
H
H
OPO32
H
OH
H
O
H OH
1 6
5
43
2
PIP2
phosphatidylinositol-
4,5-bisphosphate
E.g., pleckstrinhomology (PH)domains bind tophosphorylated
derivatives ofphosphatidylinositol.
Some PH domainsbind PIP2 (PI-4,5-P2).
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Otherpleckstrin homology domains recognize and bindphosphatidylinositol derivatives with Pi esterified at the
3' OH of inositol. E.g., PI-3-P, PI-3,4-P2, PI-3,4,5-P3.
O P
O
O
H2C
CH
H2C
OCR1
O O C
O
R2
OH
H
OH
H
H
OHH
OPO32
H
O
H OH
1 6
52
3 4
phosphatidyl-inositol-3-phosphate
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Some proteins bind to membranes via a covalentlyattached lipid anchor, that inserts into the bilayer.
The attached lipid may be a fatty acid such as palmitate
ormyristate.
Palmitate is usually attached via an ester linkage to thethiol of a cysteine residue.
A protein may be released from plasma membrane tocytosol via depalmitoylation, hydrolysis of the ester link.
lipid
anchor
membrane
H3C (CH2)14 C
O
S CH2 CH
C
NH
O
palmitate
cysteineresidue
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An isoprenoid such as a farnesyl residue, is attached tosome proteins via a thioether linkage to a cysteine thiol.
Fatty acid orisoprenoid chains link proteins to thecytosolic surface of the plasma membrane.
Many signal proteinsbind via lipid anchors and/orpleckstrin homology domains to the cytosolic surface ofthe plasma membrane. They often bind in regions of lipidrafts, where they cooperate in signal cascades.
CH CH2CH3C
CH3
CH CH2CCH2
CH3
CH CH2 S ProteinCCH2
CH3
farnesyl residue linked to protein via cysteine S
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Glycosylphosphatidylinositols (GPI) are complex
glycolipids that attach some proteins to the outer surfaceof the plasma membrane.
The linkage is similar to the following, although theoligosaccharide composition may vary:
protein (C-term.) - phosphoethanolamine mannose - mannose -
mannose - N-acetylglucosamine inositol (of PI in membrane)
The protein is tethered some distance out from the
membrane surface by the long oligosaccharide chain.GPI-linked proteins may be released from the outercell surface by phospholipases.
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Hydrophobic domains of detergents substitute forlipids, coating hydrophobic surfaces of integral proteins.
Polar domains of detergents interact with water.If detergents are removed, purified integral proteins tendto aggregate & come out of solution. Their hydrophobicsurfaces associate to minimize contact with water.
Amphipathic
detergents arerequired forsolubilization ofintegral proteins
from membranes.
detergent
solubilization
rotein with
bound detergent
polarnon-polar
membrane
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A membrane-spanning E-helix isthe most common structural motiffound in integral proteins.
membrane
N
C
Integral protein structure
Atomic-resolution structures have been determinedfor a small (but growing) number of integral membraneproteins.
Integral proteins are difficult to crystallize for X-ray
analysis.Because of theirhydrophobictransmembrane domains,detergents must be present
during crystallization.
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In an E-helix, amino acid R-groups protrude out from thehelically coiled polypeptide backbone.
The largely hydrophobic R-groups of a membrane-spanning E-helix contact the hydrophobic membrane core,while the more polar peptide backbone is buried.
Colors: C N O R-group (H atoms not shown).
E-helix
R-groups in magenta
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Particularamino acids tend to occur at different
positions relative to the surface or interior of the bilayer
in transmembrane segments of integral proteins.
Residues with aliphatic side-chains (leucine, isoleucine,
alanine, valine) predominate in the middle of the bilayer.
H3N+ C COO
CH3
H
H3N+ C COO
CH CH3
CH2
CH3
H
H3N+ C COO
CH2
CH CH3
CH3
H
H3N+ C COO
CH CH3
CH3
H
alanine (Ala, A) isoleucine (Ile, I) leucine (Leu, L) valine (Val, V)
aminoacids:non-polaraliphatic R-groups
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It has been suggested that the polar character of thetryptophan amide group and the tyrosine hydroxyl, alongwith their hydrophobic ring structures, suit them forlocalization at the polar/apolar interface.
tryptophan tyrosine
H2N C C
CH2
HN
H
H N+
C C
CH2
H
H
Tyrosine and
tryptophan arecommon near themembrane surface.
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Cytochrome oxidase is an integral protein whose intra-membrane domains are mainly transmembrane E-helices.
Explore with Chime the E-helix colored green at far left.
membrane
Cytochrome oxidase dimer (PDB file 1 CC)
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If a hydropathy plot indicates one 20-amino acidhydrophobic stretch (1 putative transmembrane E-helix),topology studies are expected to confirm location of
N & C termini on opposite sides of membrane. Iftwo transmembrane E-helices are predicted, N & C
termini should be on the same side. The segmentbetween the E-helices should be on the other side.
N
membrane
N
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An E-helix lining a water-filled channel might have
polar amino acid R-groups facing the lumen, & non-polarR-groups facing lipids or other hydrophobic E-helices.
Such mixed polarity would prevent detection by ahydropathy plot.
A helicalwheel looksdown the axisof an E-helix,
projecting side-chains onto aplane.
Simpli ied helical heel diagram o ourE-helices lining the lumen o an ion channel.
Polar amino acid -group
on-polar amino acid -group
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Porin F-barrel
While transmembrane E-helices are the most commonstructural motif for integralproteins, a family of bacterialouter envelope channelproteins called porins haveinsteadF barrel structures.
AFbarrel is a F sheet rolledup to form a cylindrical pore.
At right is shown one channelof a trimeric porin complex.
Porin onomer
PDB 1A S
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In aF-sheet, amino acid R-groups alternately point above& below the sheet.
Much of porin primary structure consists ofalternating
polar & non-polar amino acids. Polar residues face the aqueous lumen.
Non-polar residues are in contact with membrane lipids.
Explore an example of a bacterial porin with Chime.
!polar R group, !non-polar R group