1. Lipids and Membranes

2. Membranes:
 They function to organize biological systems by forming
compartments within which biological processes take place.
 Many subcellular organelles are membrane bound, e.g.
nuclei, mitochondria, chloroplasts, endoplasmic reticulum…etc.
 They are organized assemblies of lipids and proteins with
small amount of carbohydrates.
 They regulate the composition of the intracellular medium by
controlling the flow of nutrients, waste products, ions…. etc.
into and out of the cell, through pumps and gates embedded
in the membrane, which allow the transport of substances
against or with electrochemical gradients, respectively.

3. Lipids:
 They are biological substances that are soluble in organic solvents such
as chloroform, methanol, acetone….etc. but are almost insoluble in water.
They are called neutral fats since they are uncharged molecules.
 They are classified as Simple, Complex or Derived lipids.
3. Simple lipids are esters of fatty acids and alcohols.
a. fats; esters of fatty acids with glycerol.
b. waxes; esters of fatty acids with higher mol wt. mono-
hydric alcohols.
4. Complex lipids are esters of fatty acids and glycerol, but they
contain groups other than the alcohol and the acid.
a. Phospholipids: FA + Alcohol + Phosphoric acid residue.
They may also contain a nitrogen-containing base.
b. Glycolipids: FA + Alcohol (Sphingosine) + Carbohydrate.
c. Lipoproteins: Lipid moiety + Protein part.
5. Derived lipids (Precursors): include FA, Alcohols, steroids,
fat-soluble vitamins, and hormones.

4. Fatty Acids are aliphatic carboxylic acids
 Fatty acids occur mainly as esters (as in fats and oils).
 They are usually straight chain aliphatic carboxylic acids that contain
even number of carbon atoms. They are either saturated (contain no double
bonds) or unsaturated (contain one or more double bonds).
Saturated fatty acids, are derivatives of acetic acid [CH3-COOH];
CH3-(CH2)n-COOH, where n is an even number, e.g. Palmitic
acid: CH3-(CH2)14-COOH and Stearic acid CH3-(CH2)16-COOH.
Unsaturated fatty acids are mono-unsaturated or poly-unsaturated,
e.g. Oleic, linoleic and linolenic acids are 18 C acids that have 1, 2 and 3
double bonds, respectively;
Oleic acid (18:1;9): CH3-(CH2)7-CH=CH-(CH2)7-COOH
Linoleic acid (18:2;9,12): CH3-(CH2)4-CH=CH-CH2-CH=CH-(CH2)7-COOH.
α-Linolenic acid (18:3;9,12,15): CH3-(CH2)2-CH=CH-CH2-CH=CH-CH2-CH=CH-
(CH2)7-COOH.
γ-Linolenic acid (18:3;6,9,12): CH3-(CH2)4-CH=CH-CH2-CH=CH-CH2-CH=CH-
(CH2)4-COOH.
Arachidonic acid (20:4;5,8,11,14):

5. Functional Roles of Fatty Acids Determine
their Structure and Degree of Unsaturation.
 Melting points of even-numbered-carbon fatty acids increase
with chain length and decrease with unsaturation, for
example, Triglycerides containing 3 saturated FAs of 12 C or
more are solid at body temperature, while if the FAs are 18:2 for
example, the triglyceride is liquid at temperatures below 0°C.
e.g. Membrane lipids, which must be fluid at all environmental
temperatures are more unsaturated than storage lipids, also lipids in tissues that
are exposed to cold temperatures such as in extremities or in animals that
hibernate are more unsaturated.
 Fats are solid, and oils are liquids at room temperature
(RT).
 Plants produce triacylglycerols that are rich in unsaturated
FAs and thus they are liquid at RT (oils), while Triacyl- glycerols
of animal origin are rich in saturated FAs (fats).

6. Simple lipids are FA esters with
Alcohols.
 Triacylglycerols (Triglycerides; Neutral fats).
They constitute most of plant and animal fats. They
are non-polar, water-insoluble TRIESTERS of
glycerol.
They may contain one (simple triacylglycerols), two
or three (mixed triacylglycerols) different types of
FAs,
e.g. Tristearin and Triolein have 3 residues of stearic and oleic
acids, respectively.
e.g. 1-palmitoyl-2-linoleoyl-3-stearoyl-glycerol, contains
palmitic, linoleic and stearic acids esterified with the OH
groups of C1, C2 and C3 of the glycerol moiety, respectively.

7. Simple lipids are FA esters with
Alcohols; cont.
Monoacylglycerol
Diacylglycerol Triacylglycerol / Triglycerides

8. Triacylglycerols function as energy
reservoirs.
 Triacylglycerols, although are not part of biological
membranes, they are a highly efficient form of
metabolic energy storage.
 This is because the main energy producing unit is a 2-
carbon atom molecule called acetyl-CoA, which is
produced from the oxidation of glucose and FAs. Acety-
CoA is also the building units of FAs.
 In glucose and Fat oxidation for energy production, each
molecule of glucose produces 2 acetyl-CoA molecules,
while each FA forming an ester in the triacylglycerol
produces one acetyl-CoA molecule for each 2-carbon
unit of the FA chain.

9. Triacylglycerols function as energy
reservoirs; cont.
 In animals, the cells that are specialized in synthesis and
storage of fats are called adipocytes, which form adipose
tissue that is most abundant in subcutaneous (under
skin) and abdominal cavity. The subcutaneous adipose
tissue serves as a thermal insulation in warm-blooded
aquatic and polar animals.
 Fat content of normal humans (21% for men and 26%
for women) enables them to survive starvation for 2-3
months, while glycogen that constitutes short-term
energy store, provides less than a day energy supply for
body’s metabolic needs.

10. Complex Lipids:
 Phospholipids are main lipid constituents of membrane.
 They are Diacylglycerols (esterified in glycerol’s C1 &
C2 with FAs), in which the OH group of glycerol’s C3 is
forming an ester with phosphoric acid (simplest form,
called phosphatidic acid) or a phosphoric acid derivative
(an X-group attached to phosphoric acid via one of its
OH groups),
e.g. choline, is an amino alcohol that bind with phosphatidic acid through
the (OH) group of the phosphate giving rise to phosphatidylcholines (also
called lecithins), the most abundant phospholipids of the cell membranes.
e.g. inositol is a cyclic alcohol that form phosphatidylinositol, an
important constituent of cell membrane phospholipids that act as a 2nd
messenger in cell signaling mechanisms.

11. Complex Lipids; cont.
Phosphatidylcholine
(Lecithin)
Phosphatidic acid
Phosphatidylinositol

12. Complex Lipids; cont.
 Sphingolipids are also major membrane
component.
 They are derivatives of the C18 amino
alcohols (sphingosine and
dihydrosphingosine), in which the amino
group is forming an ester with FA (i.e. amide
derivative, e.g. ceramide).
e.g. Sphingommyelins, are sphingolipids that are
present in brain and nerve tissues, and consists of
choline moiety attached to ceramide phosphate (i.e.
sphingosine, FA, phosphoric acid and choline).

13. Complex Lipids; cont.
Sphingosine Dihydrosphingosine
Ceramide
Sphingosine-1-phosphate
Sphingommyelin

14. Derived Lipids:
 Steroids play many physiologically important roles.
 Cholesterol, in addition to its association with
atherosclerosis, it is the precursor molecule of large
number of biologically important steroids, including
Vitamin D, steroid hormones, bile acids….etc.
 It contains the characteristic steroid nucleus.
 It is a major constituent of the cell membrane.
18
12 17
19 11 13
1 9 C D 16
2 14
10 8 15
3
A B
7
5
4 6
Steroid nucleus
C D
A B
HO 1, 25-Dihydroxy-
Cholesterol
cholecalciferol, Vitamin D

15. Membrane Structure
 The basic structural characteristic of membranes is due to the
physico-chemical properties of phospholipids and sphingolipids
that constitute biological membranes.
 These compounds with their hydrophilic (polar) head and
hydrophobic tail, interact in aqueous systems in vitro to form spheres,
called “Vesicles” or “Liposomes”, in which the polar heads are
directed to the outside (aqueous environment) while the hydrophobic
tails interact to exclude water (in the interior side) forming what is
called lipid bilayer.

16. Membrane Permeability
 Permeability studies of lipid vesicles and electrical-conductance
measurements of planar bilayers have shown that lipid bilayer
membranes have a very low permeability for ions and most polar
molecules.
 Water is considered an exception of the above rule due to its small
size, high concentration and lack of a complete charge.
 Based on Permeability Coefficient (P) values of different ions
and molecules, ions such as Na+ and K+ traverse membranes ~ 109
fold less likely (slower) than does water. Similarly, Tryptophan,
which forms a zwitterion at pH 7.0, crosses the membrane 103 times
as slowly as does indole, the ring structure of the tryptophan that
lacks ionic groups (amino and carboxylic groups).

17. Membrane Proteins
 Membrane lipids form a permeability barrier that establish the
compartments, which constitute the nature of plasma membranes.
 Most molecules traverse membranes aided by proteins.
 Membranes differ in their protein content, where for example “Myelin”,
which serves as an electrical insulator around certain nerve fibers contains
~ 18% proteins (pure lipids serve well for insulation) as compared to 50%
and 75% protein contents of plasma membrane of other cells and the
inner membranes of mitochondria and chloroplasts, respectively.
 The higher protein contents of other structures (compared to myelin) are
mainly because these are metabolically active structures that have proteins
embedded in the membrane structure, which serve as channels, receptors,
pumps, and enzymes that are required for transport of molecules and
signals across the membranes.
 Membrane proteins are either “Integral Membrane Proteins”, which
mainly span the membrane lipid bilayer and can be released only by a
detergent or organic solvent (to solubilize the lipid bilayer and thus release
the spanning proteins) or “Peripheral Membrane Proteins”, which are
bound to membranes primarily by electrostatic and hydrogen bonds
interactions with the polar heads of the lipid bilayer or through interaction
with integral protein. The association of the peripheral proteins to
membranes can be disrupted by increasing the ionic strength (increasing
salt concentration) or by changing the pH.

18. Integral and Peripheral
Membrane Proteins
Extracellular
Peripheral
Proteins
Lipid
Bilayer
Integral Intracellular
Proteins

19. Membrane Proteins,
Interaction with membrane structures
 Proteins may span (traverse) membrane with α-helices.
 In archaea, there is an integral membrane protein called
Bacteriorhodopsin, which acts as a proton pump, where it
captures light energy and uses it to move protons across the
membrane out of the cell. The resulting proton gradient is
subsequently converted into chemical energy.
 Bacteriorhodopsin is built of 7 closely packed α-helices, arranged
almost perpendicularly to the plane of the cell membrane, spanning
its entire width (~45 Ao).
 Most of the amino acid residues in these membrane-spanning α-
helices are non polar, where they interact with the hydrophobic
hydrocarbon core of the cell membrane or with other hydrophobic
residues in adjacent α-helices.
 The tertiary structure of bacteriorhodopsin is similar to that of
rhodopsin, which senses light in the retina of vertebrate animals.
Both proteins belong to the 7-transmembrane receptor family.
However, their functions are different and there is only slight
conservation of the amino acid sequences.

20. Bacteriorhodopsin, 3-D-Structure
Lipid 7-transmembrane,
7 α-helices
Bilayer
structure of
bacteriorhodopsin

21. Membrane Proteins,
Interaction with membrane structures
 Proteins may span (traverse) membrane with β-sheets.
 Gram negative bacteria such as E. coli and some gram positive bacteria
contain an outer membrane channel protein called Porin.
 Porin is formed of β-strands in an anti-parallel arrangement that is
forming a single β-sheet that curls up to form a hollow cylinder, which acts
a s a pore (hence the name porin), or a channel.
 Unlike other membrane transport proteins, porins are large enough to
allow passive diffusion, i.e. they act as channels that are specific to
different types of molecules.
 Porins are also present in the mitochondrial and chloroplast membranes.
 Similar to bacteriorhodopsin, porins have hydrophobic outer surface to
allow the interaction with the hydrocarbon core of the cell membrane or
with other hydrophobic residues in adjacent β-strands.
 In contrast, the interior of the channel is quite hydrophilic and filled with
water to permit the diffusion of the solutes.
 This characteristic structure of non-polar surface with polar interior of the
channel is achieved by a tandem alteration of hydrophobic and hydrophilic
amino acid residues along the β-strands.

22. Porin, Membrane Channel Protein
Tertiary Structure Top view of the channel
Lipid
Bilayer
Hydrophobic Hydrophilic
amino acid amino acid
residues residues
(Yellow) (White)
Secondary Structure

23. Membrane Proteins,
Interaction with membrane structures
 Proteins may be partially embedded in the membrane, i.e. not
spanning through the membrane.
 The membrane-bound enzyme prostaglandin H2 synthase-1
(bound to endoplasmic reticulum), which catalyzes the synthesis of
prostaglandin H2 (a pain and inflammation mediator and a
modulator of gastric acid secretion) from arachidonic acid (20:4;
5,8,11,14), is a homodimer that firmly attaches to the membrane
through the interaction of a set of α-helices with hydrophobic
surfaces with the membrane.
 This association is sufficiently strong to the extent it requires the
action of a detergent(s) to be disrupted and thus it’s classified as an
integral membrane protein.
 Since the substrate of this enzyme, the arachidonic acid, is
hydrophobic molecule that is generated by the hydrolysis of the
membrane lipids, the association of this enzyme to the membrane is
crucial to its function, where the substrate reaches the active site of
the enzyme from the membrane without entering the aqueous
environment of the cytoplasm. This is achieved by travelling of the
substrate through a hydrophobic channel in the protein.

24. Prostaglandin H2 Synthase-1,
Catalyzed reaction & Membrane association

25. Cox-Inhibitors,
Blocking substrate channel to active site
 Drugs that are called COX-inhibitors (cyclooxygenase inhibitors)
such as aspirin and ibuprofen act by blocking this channel and
prevent prostaglandin synthesis by inhibiting the cyclooxygenase
activity of the synthase enzyme.
 Aspirin, for example (acetyl salicylic acid), acts by acetylating a
serine residue in position 530 of the synthase enzyme (Ser530),
which lies along the channel that leads the substrate to the active
site.

26. Membrane Proteins,
Interaction with membrane structures
 Soluble proteins may associate with the membranes through enzyme-
catalyzed attachment of a hydrophobic group to the protein.
 Examples of such groups include:
– Palmitoyl group attached to a cysteine residue by thioester bond.
– Prenyl group (farnesyl or geranylgeranyl) attached to a cysteine residue at a C-
terminal end of the protein, e.g. Ras attachment to the membrane.
– Glycosylphosphatidylinositol (GPI) anchor attached to the C-terminus.

27. Prediction of Transmembrane Helices
• To identify transmembrane helices is by evaluating the stability of a
postulated helical segment and see whether it’s most stable in a
hydrocarbon environment or in water, i.e. by estimating the free-energy
change when this helical segment is transferred from the interior of a
membrane to water.
• The sum of the Free-energy changes for the transfer of individual amino
acids from a hydrophobic to an aqueous environment, determine whether a
segment composed of such amino acids is likely to span the membrane or
not.
• For example, an α-helix formed of L-Arg (positively charged) from the interior of the membrane
to water would be highly favorable (-51.5 kJ mol-1 OR -12.3 kcal mol-1/ residue). In contrast, the
transfer of an α-helix formed of L-Phe (hydrophobic) from the interior of the membrane to
water would be unfavorable producing an energy change of +15.5 kJ mol-1 OR +3.7 kcal mol-1/
residue.
• The hydrocarbon core of a membrane is typically 30Ao, a length of a 20-
amino acid residue-α-helix.
• Thus transmembrane domain/region of a protein can be identified by
estimating the free-energy change that takes place when a hypothetical α-
helix formed of any 20-amino acid residues is transferred from the
membrane interior to water.
• The free-energy change of each set (called window) is plotted to create a
“Hydropathy PLOT”.
PLOT
• A peak of +84 kJ mol-1 (+20 kcal mol-1) or more indicates that the tested
poly-peptide segment (20 amino acid residues) could be a membrane-
spanning α-helix.

28. Prediction of Transmembrane Helices

29. Prediction of Transmembrane Helices
(A possible
transmembrane helix)
No predicted tansmembrane
helices could be found