wuolah-free-LESSON-7

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LESSON 7. STRUCTURE AND FUNCTION OF ORGANELLES

Eukaryotic cells have a diameter 10 times bigger and a volume 1000 times bigger than
prokaryotic cells, so they are organized in smaller compartments (organelles). Advantages of
compartmentalization:
 Incompatible reactions can be separated: fatty acids
 Efficiency of chemical reactions increases: localization and concentration of substrates,
faster diffusion for local reactions and enzyme clustering to increase efficiency
For all of this, there is a relation between structure and function.
The endomembrane system regulates protein traffic and performs metabolic functions in the
cell. Components of the endomembrane system are either continuous or connected via
transfer by vesicles:
- Endoplasmic reticulum
- Nuclear envelope
- Golgi apparatus
- Lysosomes and endosomes
- Peroxisomes
- Vacuoles
- Plasma membrane
There are three ways of transporting substances between cell components:
 Gated transport: between the nucleus and the cytosol
 Vesicular transport: between cell exterior, Golgi apparatus and Endoplasmic Reticulum
using vesicles, lysosomes and endosomes.
 Transmembrane transport: from cytosol to mitochondria, peroxisomes, plastids and ER.
Vesicle formation and transport
Vesicles continually bud off from one compartment and fuse with another, carrying
membrane components and soluble molecules known as cargo from the lumen and
membrane of the donor compartment to the lumen and membrane of the target
compartment . This membrane traffic allows the cell to secrete and eat.
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Proteins’ signal sequences

Proteins that are destined towards the secretory pathway and transported from one organelle
to another show signal sequences, with are interconexions of endomembrane system. They
are short peptides present at the N-terminus of the newly synthesized proteins and they are
different depending on the protein final destination.
The ENDOPLASMIC RETICULUM: biosynthetic factory
The endoplasmic reticulum (ER) accounts for more than half of the total membrane in many
eukaryotic cells. Its structure is a cisternae or tube network, delimited by a membrane
(continuous with the nuclear envelope).
ROUGH ENDOPLASMIC RETICULUM: with ribosomes studding its surface.

Proteins synthesis
Their destination is as a membrane of the endomembrane system or they are secreted.
Proteins are produced in rough EU, mitochondrion, chloroplasts or cytoplasm.
Proteins are produced by translating the genetic code of mRNA into a string of amino acids.
Ribosomes require tRNA to produce proteins: tRNA brings the amino acids that correspond
with the genetic code in mRNA to the ribosome. The tRNA molecule binds to the ribosome
and there is a site for the portion of the tRNA that binds to the next amino acid and a site for
the growing amino acid chain. The tRNA is released after the protein is completed.
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SRP cycle: translation begins with synthesis of a short signal peptide sequence: a signal
recognition particle. A protein complex binds to the signal peptide while translation continues.
The SRP then binds to its receptor in the ER membrane anchoring the ribosome. The ribosome
binds its receptor and the signal peptide meets the protein translocator. Translation continues
and translocation begins. The signal peptidase cleaves the signal peptide, displace and
recycled, leaving the new protein molecule in the lumen of the endoplasmic reticulum.
▪ Free ribosome cycle: free ribosomes are unattached to any membrane. They synthesize
proteins that are used by the cell (the metabolism of food).
▪ Membrane-bound ribosome cycle: synthesizes proteins that are transported out of the cell
and required for a specific function (digestive enzymes).
Type of proteins according to their final destination:

Free proteins in the ER lumen:
The signal sequence is guided to the ER membrane with a signal recognition particle which
binds the ER signal sequence in the new protein as it emerges from the ribosome. Protein
synthesis then slows down until the SRP ribosome complex binds to an SRP receptor in the ER
membrane. The SRP is then releases, passing the ribosome to a protein translocation channel
in the ER membrane. Thus the SRP and SRP receptor function as molecular matchmakers,
connecting ribosomes that are synthesizing proteins containing ER signal sequences to
available ER translocation channels. In addition to directing proteins to the ER, the signal
sequence functions to open the translocation channel. The signal peptide remains bound to
the channel while the rest of the protein chain is threaded though the membrane as a large
loop. Once the protein has passed through the membrane, it is releases into de ER lumen.
After the signal sequence has been cleaved off by a signal peptidases located on the luminal
side of the ER membrane, the signal peptide is then releases from the translocation channel
into the membrane and rapidly degraded. It is thought that a protein serving as a plug then
binds from the ER lumen to close the inactive channel.
Transmembrane – single domain:

Some proteins that enter the ER remain embedded in the ER membrane as transmembrane
proteins. In the simplest case that of a transmembrane protein with a single membrane
spanning segment, the n-terminal signal sequence initiates translocation just as for a soluble
protein but the transfer process is halted by an additional sequence of hydrophobic amino
acids, a stop transfer sequence, further in the polypeptide chain. The stop transfer sequence
is releases laterally from the translocation channel and drifts into the plane at the lipid bilayer
where it forms a membrane spanning segment that anchors the protein in the membrane. As
a result, the translocated protein ends up as a transmembrane protein inserted in the
membrane with a defined orientation in some transmembrane proteins. An internal signal
sequence is used to start the protein transfer which continues until a stop transfer sequence
is reached. The two hydrophobic sequences are then released into the bilayer where they
remain anchored in complex multi pass proteins in which many hydrophobic regions span the
bilayer. Additional pairs of stop and start sequences come into play one sequence reinitiates
translocation further down the polypeptide chain and the other stops translocation and
causes polypeptide release and so on for subsequent starts and stops. Thus multipack
membrane proteins are stitched into the lipid bilayer as they are being synthesized by a
mechanism resembling a sewing machine.
Protein glycosylation:
Glycosylation is the process in which proteins are covalently bonded to carbohydrates. It
begins in the endoplasmic reticulum and continues in the Golgi apparatus. Enzymes in the ER
remove glucose and mannose residues from the oligosaccharide chains. Vesicles transport the
glycoprotein to the SIS Golgi network where one of two events occurs: either n-acetyl
glucosamine attached to the oligosaccharide and are transported to the lysosomes or
enzymes remove more mannose residues. Vesicles transport the modified glycoprotein to the
medial Golgi cisternae where n-acetyl glucosamine attaches and more mannose molecules
leave the molecule. Vesicles now transport the glycoprotein to the trans-Golgi cisternae.
There are galactose and sialic acid the chain. The glycoprotein is finally ready for transport to
the cells plasma membrane.
Glycosylation begins with the addition of a 14-sugar oligosaccharide precursor (3 glucose, 9

mannose and 2 n-acetylglucosamine molecules) to an asparagine amino acid. The link is
between the first sugar Acetylglucosamine and the group NH2 of the Asn lateral chain. This
entity is then transferred to the ER lumen. The oligosaccharyl transferase enzyme attaches the
oligosaccharide chain to asparagine. The oligosaccharide attached protein sequence now folds
correctly and is now translocated to the Golgi body where the mannose residue is removed.
Distribution of transport vesicles:
Vesicles are proteins surrounded by membranes. Each compartment encloses a space, called
a lumen, that is topologically equivalent to the outside of the cell, and all compartments
shown communicate with one another and the outside of the cell by means of transport
vesicles.
Exocytic-secretory way: protein molecules are transported from the ER to the plasma
membrane or (via late endosomes) to lysosomes.
Endocytic way: molecules are ingested in vesicles derived from the plasma membrane and
delivered to early endosomes and then (via late endosomes) to lysosomes.
Membrane recovery way: many endocytosed molecules are retrieved from early endosomes
and returned to the cell surface for reuse, some molecules are retrieved from the late
endosome and returned to the Golgi apparatus, and some are retrieved from the Golgi
apparatus and returned to the ER. All of these retrieval pathways recover membrane because
it can lose its surface during cell vesicles transport.
Transition Endoplasmic Reticulum: located between rough and smooth ER.
SMOOTH ENDOPLASMIC RETICULUM: lacks ribosomes.

- Synthesizes lipids: phospholipids of membrane and cholesterol.
- Detoxifies poison.
- Stores calcium: muscle cells.
Synthesis of phospholipids of membrane
It occurs in the outer endomembrane of the smooth ER.
1. A fatty acid is transported by a binding protein to the outer of the smooth ER membrane.
2. An enzyme called CoA transferase (it is in the outer membrane) activate fatty acids by
adding CoA.
3. The acyl transferase enzyme (it is in the membrane) transfers the glycerol 3 –phosphate to
a couple of activated fatty acids and removes CoA, forming phosphatidic acid.
4. A phosphatase (resident enzyme of the cytoplasm) removes the phosphate group from the
phosphatidic acid and forms diacylglycerol.
5. The choline phosphotransferase (it is in the membrane) transfers and activate the choline
with one phosphate to the diacylglycerol and forms phosphatidylcholine. In this process a
CMP is released.
Phospholipids synthesis adds to cytosolic half of the bilayer, so there are more phospholipids
in the outer endomembrane. The ER forms vesicles, whose bilayer has the same number of
phospholipids in both halves, but there is an asymmetric growth. In the cell exterior, there is
an asymmetric lipid bilayer of plasma membrane. Vesicles flip from ER membrane to cytosol
of plasma membrane, a reaction catalyses by scramblase: there is a delivery of new membrane
by exocytosis. Then, flippase catalyses flipping of specific phospholipids to cytoplasmic
monolayer.
Detoxifies poison (Cytochrome P450)
Detoxification is the removal of all the toxic materials such as metabolic wastes or drugs. The
smooth endoplasmic reticulum present in the liver contains enzymes that transform lipid-
soluble compounds into water-soluble ones and those can easily be expelled out of the body.
For example, the Heme group is oxidize using O2 and H2O, reduces O2 and gives it to the toxic
hydrophobic compound R-H, which becomes a soluble molecule R-OH.
Toxic compounds (Hydrophobic R-H)  Oxidation: Liver (Hydrophilic R-OH)  Elimination:
Kidneys
The nuclear envelope

Between the nuclear envelope and the chromatin there is the nuclear lamina, a net that covers
the chromatin. The nuclear envelope has the same structure as the ER: inner membrane, outer
membrane, ribosomes and nuclear pores. The number of nuclear pores changes from one
type of animals to another because it changes the number of nucleoporins, but the structure
is the same. Vertebrates have 3000-4000 pores. The nucleoporins are the proteins that form
the nuclear pore and controls molecules’ transport through the nuclear envelope.
- External ring: has a filament
component.
- Radial proteins: are present in the
pass between the nucleus and the
cytosol.
- Column nucleoporins: connect
radial proteins with anchoring
proteins.
- Anchoring proteins: (they support
the structure: the radial and the column nucleoporins to the nuclear envelope).
The inner ring proteins are similar to the external rings but their filaments are connected
forming the nuclear basket. In every nucleoporins’ family there is an octal symmetry: there
are eight nucleoporins. Many proteins are produced outside the nucleus but there are needed
inside it. There is a big molecules’ transport through the nuclear pore. Karyopherins:
molecules that work like passports:
- Exportins: from nucleus to cytoplasm (ribosomes, mRNA). The nuclear export receptor
pass to the nucleus, interacts with Ran-GTP and the cargo molecule and then goes back to the
cytosol, hydrolyses the Ran-GDP and a phosphate group and delivers the cargo molecule to
the cytosol. Just the plain exportin is allowed to go pass the nucleus.
- Importins: from cytoplasm to nucleus (proteins to DNA synthesis). The nuclear import
receptor links with the cargo molecule (a protein with nuclear localization signal) to pass from
the cytosol to the nucleus. The rand GTP binds to the import molecule, cargo molecule is
delivered to the nucleus and the importin goes back to the cytosol and hydrolyses the Ran-
GDP and a phosphate group.
GOLGI APPARATUS
Each cisternae is specialized in a specific biochemical trait. It is formed by three cisterna (cis,
medial and trans) and has two faces (cis Golgi network and trans Golgi network). Each
cisternae is specialized in a specific biochemical trait.
Modification of glycoproteins (N-glycosylation at RER)
It is the attachment of the sugar molecule oligosaccharide glycan to a nitrogen atom (the
amide nitrogen of an asparagine (Asn) residue of a protein).
- Proteins with complex oligosaccharides: modification of N-oligosaccharide fraction
(reduction) and addition of new sugars (O-glycoside).
- Proteins with mannose-rich oligosaccharides: modification of N-oligosaccharide fraction
(reduction) and addition of mannoses (O-glycoside).
Glycosylation of proteins (O-glycosylation)
O-glycosylation is the attachment of a sugar molecule to an oxygen atom in an amino acid
(serine or threonine) residue of a protein and GalNAc. Glycosil transferases enzymes have a
high substrate specify and justifies the high variability of glycoproteins.
Proteoglycans are main components of the extracellular matrix. They are formed by a back
bone of hyaluronic acid and branches of proteins with a high density of glycosylation. The
protein component of proteoglycans is synthesized by ribosomes and translocated into the
lumen of the rough endoplasmic reticulum. Glycosylation of the proteoglycan occurs in the
Golgi apparatus in multiple enzymatic steps. Firstly, a special link tetrasaccharide (xylose) is
attached to a serine side chain on the core protein to serve as a primer for polysaccharide
growth. Then sugars (dimers) are added one at a time by glycosyl transferase. The completed
proteoglycan is then exported in secretory vesicles to the extracellular matrix of the tissue.
One of the monomers is a NAc-sugar (N-acetylgalactosamine or N-acetylglucosamine). Roles:
extracellular matrix, external localization, main component of mucus.
Modification of lipids (glycosylation).

Glycolipids are lipids covalent bonded to oligosaccharides. They occur in the non-cytosolic
hemimembrane of the GA upon synthesis and in the external hemimembrane of plastic
membrane (5% of lipids). They have an asymmetric distribution in cells. In animal cells:
sphingosine and gangliosides. Their functions are the transmission of membrane potential
(change, nerve cells) and the identification of signals.
Biological relevance of protein and lipid glycosylation
 Regulates cell development
 Protection against proteases
 Protein tagging for target organelles
 Help on the folding process: it is used to fold completely proteins.
 Receptors: communication of cells.
 Cell identification
 Other (unknown): glycosylation has not been completely investigated.
Control intracellular vesicular traffic

Exocytic secretory pathway
1. Formation of vesicles on donor membrane.
It is carried out by coating proteins, which select different cargo. They can be modified by
forming complex with additional proteins. There are three types:
- Clathrin: endocytic vesicles (outside into
inside) from Golgi apparatus and plasma
membrane.
- COP I: coated vesicles bud from Golgi
apparatus
- COPII: coated vesicles bud from ER.
In the plasma membrane there are some cargo receptors that activate when they link to some
molecules. After activating, they link to adaptins, some proteins that link to clathrin in the
other side of the membrane. As more cargo receptors activate, more clathrins bind to the
other side of the membrane forming buds. When there is a high number of complex, a
membrane itsmus is formed. Then, a protein called dynamin (it produces ATP in order to
move) cuts and separates the vesicle. Finally, proteins are removed and a naked transport
vesicle is formed, which will later be transported by kinesins.
2. Transport from donor to target membrane. It is carried out by microtubules cytoskeleton

and motor proteins, which allow movement.
Kinesins are motor proteins that move along microtubules usually carrying cargo such as
organelles and vesicles from the centre of a cell to its periphery (exocytic pathway). Dyneins
are cytoskeletal motor proteins that also move along microtubules in cells and transport
vesicles in the endocytic pathway. They generate force and movement on microtubules in
biological processes (cell division).
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3. Arrival to target membrane and fusion. Vesicles go to different proteins.

SNARE proteins are a large protein complex that mediate vesicle fusion, that is, the fusion of
vesicles with their target membrane bound compartment. The v-SNARE are in the vesicle
membrane and the t-SNARE in the target membrane. Rab GTPases are proteins that regulate
many steps of membrane trafficking, including vesicle formation, vesicle movement along
actin and tubulin networks, and membrane fusion. They have a partner called rab effector
present in the target membrane. There are specific rab proteins for specific destinations.
The vesicle with cargo molecules and cargo receptors have around many v-SNARE and rab
proteins. Firstly, there rab effector identifies the rab protein and determinates its final
destination: there is a first check-point. Then, the rab protein hydrolyses the GTP and changes
rab effector shape. The v-SNARE and the t-SNARE interact with each other (there is a second
check-point) and that produces the movement of the vesicle, which fusions with the
membrane and liberate the cargo molecules into the membrane. Finally, there is a complex of
t-SNARE and v-SNARE proteins left that is expulsed.
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LYSOSOMES
They are digestive compartments, membranous sacs loaded with hydrolytic enzymes.
Lysosomal enzymes can hydrolyse proteins, fats, polysaccharides and nuclei acids in an acid
medium. Lysosomes are related with other cytoplasmic vesicular structure: endosomes, which
are linked to endomembrane system and endocytosis and exocytosis processes.
Proton pumps transfer protons from the cytosol to the inner lysosomes using ATP energy.
Lysosomes are bigger than the vesicles involved in other processes. The lysosomal hydrolase
precursor has to be a glycoprotein to keep the precursor inactivated and to identify the
protein that must be transported to the lysosome.
Origin of hydrolases
1. Each primary lysosome is a vesicle that comes from the Golgi apparatus and has hydrolases.
Hydrolases are synthetized in the ER and are transported to the Golgi network, where they
are linked to an oligosaccharide (mannose).
2. The lysosomal hydrolase precursor binds to a phosphate and to N-Acetylglucosamine.
3. The N-Acetylglucosamine is removed in order to uncover the mannose 6 phosphate signal
and let it be recognised.
4. In the trans Golgi network there are mannose 6 phosphate receptors that bind to the signal
and form clathrin coats.
5. There is a glycoprotein vesicle transport, hydrolases arrive to endosomes and they fusion
with their membrane.
6. Due to the endosome acidic pH, there is a dissociation and the receptor is removed.
7. Then, the phosphate is remove in order to avoid another interaction with the receptor and
keep them inactivate until the lysosome formation.
8. The early endosome is the young form of a lysosome, formed by the fusion of vesicles. It
turns into a lysosome when they have the proper pH and the necessary amount of
hydrolases. Mannose 6 phosphate receptors go back in budding vesicle to Golgi apparatus
following the exocytic pathway and helped by retromer coat proteins. At the end of the
cycle there is a receptor recycling.
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Digestion
Lysosomes carry out the digestion of material taken up from
outside the cell by endocytosis. They also digest material
derived from phagocytosis and autophagy. In phagocytosis,
phagosome vesicles are bigger than endocytosis vesicles,
where they are formed by coating proteins and have the
same size. In digestion processes there is a pH change, from
6 in late endosomes to 5 in lysosomes.
1. Autophagy: is the major intracellular degradation system by which cytoplasmic
materials are delivered to and degraded in the lysosome. It is the digestion of old organelles
or those with some sort of degradation that are no longer functional. Some basic components
can be recovered and used again. There are some vesicles that engulf cytosol and organelles,
forming autophagosomes (they have double membrane). They fusion with lysosomes and
their hydrolytic enzymes carry out the digestion, which forms lysosomes.
2. Phagocytosis begins by interaction of phagocytic receptors with ligands on the surface

of target particles. Then, receptors cooperate for recognition and ingestion of the particle.
They bind to pathogen-associated molecular patterns (PAMPs) and prepare the cell for
phagocytosis by inducing inside-out activation of phagocytic integrin. Important changes in
membrane remodelling and the actin cytoskeleton take place leading to the formation of
pseudopods that cover the particle. At the point of contact, a depression of the membrane
(the phagocytic cup) is formed. Then, the membrane surrounds the target particle and it closes
at the distal end, leaving a new phagosome. The phagosome changes its membrane
composition and its contents, to turn into a phagolysosome, a vesicle that can destroy the
particle ingested. This transformation is known as phagosome maturation and consists of
successive fusion and fission interactions between the new phagosome and lysosomes. At the
end, the mature phagosome called phagolysosome has a different membrane composition,
which allows it to contain a very acidic and degradative environment. They are finally
degraded and expulsed by exocytosis to the cytosol.
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Recycling
It is the partial degradation of cholesterol (absorption). Liver produces low density
lipoproteins that transport cholesterol molecules to the cells. Cholesterol in hydrophobic, so
it needs to be transported by LDL particles. High levels of cholesterol in blood is due to low
density of LDL receptors in plasma membrane that cannot take so much cholesterol levels
from the blood.
LDL particles interact with LDL receptors of the plasma membrane, enter into the cell and form
coated vesicles. Then, vesicles are uncoated and they fusion with endosomes forming early
endosomes. LDL receptors are budding off of transport vesicles that are recycling endosomes
and they return to plasma membrane. Finally, there is a partial hydrolyse. Vesicles fusion
lysosomes and hydrolytic enzymes free cholesterol.
Transcytosis
In new-borns, antibodies are transferred from mother. The bottom part of antibodies are Fc
receptors. Antibodies are transported in vesicles from the intestinal lumen to the early
endosome, where antibodies are kept and recycling transport vesicles transport empty
vesicles to the intestinal lumen. Transport vesicles take antibodies to recycling endosomes
and at the end, transcytosis transport vesicles carry antibodies to extracellular fluid.
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Storage of ready-to-use transporters of plasma membrane

Insulin is produced by the pancreas. It interacts with glucose to allow the cells of the body to
use glucose as energy. When there is excess glucose in the bloodstream, insulin encourages
the storage of glucose as glycogen in the liver, muscle and fat cells. These stores can then be
used at a later date when energy requirements are higher. As a result of this, there is less
insulin in the bloodstream, and normal blood glucose levels are restored.
PEROXISOMES
They are specialized metabolic compartments bounded by a single membrane with oxidative
enzymes, which come from ER and Golgi apparatus or from cytosol. Its main function is the
oxidation of molecules: peroxisomes produce hydrogen peroxide and convert it into water.
- Oxidases use molecular O2 to remove Hydrogen atoms from specific organic substrates. In
this process H2O2 is produced and it will be deleted by catalases. RH2 + O2  R + H2O2
- Catalases use H2O2 to oxidize various substances (phenols, formic acid, formaldehyde,
alcohol). They are especially important in liver and kidney, where peroxisomes detoxify a
wide variety of toxic molecules. H2O2 + R’H2  R’ + 2 H2O
When an excess of H2O2 is accumulated in the cell the catalase transform it in water by the
reaction: H2O2  2H2O + O2.
Fatty acids beta oxidation
Β-oxidation of fatty acids occurs in mitochondrion in animals and in peroxisomes in yeasts and
plants. Four steps:
 A long-chain fatty acid is dehydrogenated to create a trans double bond between C2 and C3
(α and β carbon). This is catalysed by acyl CoA dehydrogenase to produce trans-delta 2-
enoyl CoA. It uses FAD as an electron acceptor and it is reduced to FADH2.
 Hydration of the double bond.
 Dehydrogenation of the β-hydroxyl group to a ketone. This enzyme uses NAD as an electron
acceptor and it is reduced to NADH + H+.
 Acylation: Acetyl-CoA is added and the process continues until all of the carbons in the fatty
acid are turned into acetyl CoA.
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Glyoxysomes
Glyoxysomes are special peroxisomes. In germinating seeds, they transform the stored fatty
acids from seed lipids into sugars needed for the young plant growth. The glyoxylate cycle is
an anabolic pathway occurring in plants, bacteria, protists and fungi. It carries out the
conversion of acetyl-CoA to succinate for the synthesis of carbohydrates.
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MITOCHONDRION
It is a double membrane organelle that does not belong to endomembrane system. Its main
function is cell energy production (ATP), but actually is the ATP regeneration. Its origin comes
from the endosymbiotic theory.
Outer membrane
It is formed by 40% lipids and 60% proteins (similar that plasma membrane). It has a high
amount of unsaturated fatty acids that produce high permeability, and a low amount of
cholesterol. Besides, porines carry out the pass of molecules smaller than 10 KD by facilitated
diffusion. Monoamine oxidases are a family of enzymes bound to the outer membrane
of mitochondria that catalyse the oxidation of monoamines, employing oxygen to clip off
their amine group. Acyl–CoA–sintetase is an enzyme involved in the metabolism of acetate,
it supports the reaction that using a substrate of pyruvate produces Acyl–CoA. The enzyme’s
activity takes place in the mitochondrial matrix.
Intermembrane space
It has a chemical composition like cytoplasm. Adenylkinase is used to produce 2 ADP from
AMP and ATP. It is a specific transporter that exchanges ATP from the matrix with ADP and Pi
from intermembrane space.
Inner membrane
It is formed by 20% lipids, 80% proteins and has no cholesterol. Besides, it also has the
cardiolipin molecule, which represents the 20% of the lipids and is like a double phospholipid
that makes highly impermeable the membrane. Proteins are essential and there is a huge
variety of them on the inner membrane:
Transport proteins (also at outer membrane)
Some proteins are synthesized in mitochondria or chloroplasts from its DNA but other proteins
are formed in the nucleus by genomic DNA. Proteins can have their functions in the outer
membrane or inside the mitochondria. Those proteins present specific signal sequences that
determinate the protein final destination. Set of transport proteins of proteins on
mitochondrion membranes. SAM COMPLEX (Sorting Assemble Machine) is a protein channel
of the outer membrane that never works alone. TOM COMPLEX (Transport Outer Membrane)
have protein receptors that interact with the proteins and pass them from the cytosol to the
inner membrane.
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TIM 23 COMPLEX is connected by two expansions to the outer membrane and has a
chaperonin called Hsp70 import ATPase. TIM 22 COMPLEX Transport proteins use different
complex to transport proteins to four destinations: outer membrane, intermembrane space,
inner membrane and matrix.
- Final destination: matrix. The precursor protein shows a signal sequence that is recognised
by the TOM COMPLEX receptor. When the signal sequence interacts with the signal
receptor, chaperonins change its shape and it inserts into the membrane through TOM
COMPLEX and TIM 23 COMPLEX. Then, it is translocated into the matrix, separated from
the signal peptide by the peptidase and a mature mitochondrial protein is formed.
- Final destination: inner membrane. The first paths are the same as the previous way.
However, when the signal sequence is removed, the protein realises that its final
destination is the inner membrane. Therefore, it links to a second signal sequence and
passes through the OXA COMPLEX to the inner space.
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Other transport proteins

ADP must be transported from the cytosol to the matrix to produce ATP. Phosphate groups
and carboxylic acids transport proteins. Fatty acids are transported from cytosol to the matrix,
where the beta oxidation takes place. The Acyl CoA-sintetase is a resident enzyme in the outer
membrane that makes interact the fatty acid with CoA to form the Acyl CoA complex. The
carnitine plamiltranserase I transports the Acyl CoA to the intermembrane space and makes
it interact with the carnitine. CoASH is removed and recycled. A translocase is a protein that
transports the complex from the intermembrane to the carnitin palmitiltransferase II.
Carnitine is produced and can be used again. Carnitine palmitiltransferase II uses CoASH to
form SCoA complex.
Cellular respiration proteins

Four different complex formed by various peptides that cannot move are essential in the
electron transport chain, they can accept and lose electrons, which produce the release of
protons to the intermembrane space. In the pyruvate oxidation, each pyruvate from glycolysis
goes into the mitochondrial matrix, where it is converted into a two-carbon molecule bound
to Coenzyme A, known as acetyl CoA.
Citric acid cycle: the acetyl CoA combines with a four-carbon molecule and goes through a
cycle of reactions, ultimately regenerating the four-carbon starting molecule. ATP, NADH and
FADH2 are produced, and carbon dioxide is released.
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Electron transport chain
Oxidative phosphorylation (ATP synthase) proteins
Oxidative phosphorylation is powered by the movement of electrons through the electron
transport chain, a series of proteins embedded in the inner membrane of the mitochondrion.
There are four complex that produce the gradient of protons.
The NADH and FADH2 deposit their electrons in the electron transport chain, turning back into
NAD+ and FAD. As electrons move down the chain, energy is released and used to pump
protons out of the matrix, forming a gradient. Protons flow back into the matrix through an
enzyme called ATP synthase, making ATP. At the end of the electron transport chain, oxygen
accepts electrons and takes up protons to form water.
Mitochondrion structure
In the matrix there are
 A high amount of proteins:
- Related to catabolism: enzymes of Krebs cycle and beta oxidation.
 Enzymes which transforms pyruvic acid to acetyl-CoA, coming from glycolysis, fatty
acids or some amino acids.
 Enzymes which oxidize citric acid to acetyl-CoA (Krebs’ cycle = citric acid cycle = tri-
carboxylic acids cycle).
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- Related to replication of mtDNA, transcription of mtDNA and translation of mtRNAm. Most

mitochondrion proteins are produced at cytoplasm (95%).
 Mitochondrion DNA
 Mitorribosomes: ribosomes that support the transcription and translation processes.
Mitochondrion biogenesis
They way that mitochondria produces more mitochondria proves that the endosymbiotic
theory is true. Small mitochondria can fuse in order to produce big mitochondria and big
mitochondria fission to form small mitochondria.
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CYTOSKELETON
It is a network of protein fibres extending throughout the cytoplasm and some in nucleus that
organizes structures and activities in the cell. Functions:
- Structural support: shape and size.
- Coordination of intracellular motility.
- Regulation of some biochemical activities.
- Mitosis.
- Structural support of organelles.
Cytoskeleton is composed of three types of molecular structures:
1. Structural proteins:
- Microtubules are the thickest of the three components of the cytoskeleton. Subunits:
tubulin.
- Microfilaments are also called actin filaments and are the thinnest components.
Subunits: actin.
- Intermediate filaments are fibres with diameters in a middle range. Subunits: keratin-
like proteins.
2. Motor proteins: myosin, dynein and kinesin.
3. Accessory proteins: cross linking proteins, membrane binding proteins and
polymerization controlling proteins.
MICROTUBULES OR TUBULIN POLYMERS
They are hollow tubes whose walls consist of 13 columns of tubulin molecules. The diameter
is 25 nm with 15 nm lumen. Protein subunits are tubulins, consisting of α-tubulin and β-
tubulin. The microtubule is formed by 13 protofilaments and has two parts: the plus end is the
place where the microtubule grows and the minus end.
22
Polymerization and depolymerization

A GTP-tubulin dimer may yield a straight protofilaments and GTP hydrolysis changes subunit
conformation and weakens bond in the polymer, there is now a curved protofilaments. Then,
depolymerisation would break the chain and form a GDP-tubulin dimer. Finally, there is a GDP-
GTP exchange and the process starts again.
Motor proteins
Motor proteins are a class of molecular motors that can move along
the cytoplasm of animal cells and convert chemical energy into
mechanical work by the hydrolysis of ATP. Motor proteins are the
driving force behind most active transport of proteins and vesicles in
the cytoplasm.
Kinesins and cytoplasmic dynein play essential roles in intracellular
transport such as axonal transport and in the formation of the spindle
apparatus and the separation of the chromosomes during mitosis and
meiosis. Both are composed by many peptides and have two head
domains, a coiled coil and a light chain. Kinesin heads, lagging head
(ATP) and leading head (ADP), interact with the microtubule surface.
Hydrolysis of the ATP produces the movement of the lagging head.
Then, the ADP of the leading head takes the free phosphate, changing
to ATP and moving. This process is repeated
and produces the kinesin movement.
23
Accessory proteins
Polymerization / depolymerisation controlling proteins
Centrioles show a huge quantity of gamma tubulin dimers.
1. Some proteins nucleate the assembly of gamma tubulins and remain associated with the
minus end.
2. Some proteins bind tubulins subunits and prevent assembly.
3. Some proteins remain associated with growing plus ends and can link them to other
structures, such as membranes.
4. Some proteins enhances catastrophic disassembly at plus end.
5. Some proteins stabilize tubules by binding along sides.
Cross linking proteins

Some proteins maintain parallel microtubules: some produce cross-linking between them and
other link them to intermediate filaments.
24
Microtubules presence on cell structures and processes

Cilia and flagella are composed of microtubules that are connected to the plasma membrane
and arranged in as a 9 + 2 pattern. The pattern consists of a ring of nine microtubule paired
sets that encircle two singular microtubules. The base of cilia and flagella is connected to the
cell by modified centriole structures called basal bodies. Movement is produced when the
nine paired microtubule sets slide against one another causing cilia and flagella to bend. The
motor protein dynein is responsible for generating the force required for movement.
Mitotic spindle is formed by the attachment of microtubules to chromosomes` kinetochores.

It is formed in metaphase and it is important in mitosis.
Centrosome is an organelle that serves as the main microtubule organizing centre of the
animal cell. Each centriole is based on a nine triplet microtubule assembled in a cartwheel
structure.
25
MICROFILAMENTS (ACTIN FILAMENTS)

They are two intertwined strands of actin, each a polymer of actin subunits. They have a
diameter of 7 nm and actin protein subunits. Functions:
- Maintenance of cell shape (tension-bearing elements).
- Changes in cell shape.
- Muscle contraction.
- Cytoplasmic streaming.
- Cell motility (as in pseudopodia).
- Cell division (cleavage furrow formation).
Structural actin proteins
The monomer is a molecular protein called Actin which has a pocket with ATP units. Various
monomers assemble forming polymers, which will link and form an actin filament. The minus
end is the place where the filament loses proteins and the plus end the place where they are
added. The ATP activates the monomers in order to make a filament.
Polymerization and depolymerization

The actin filaments are very dynamic. Many new actin molecules are added in the plus end
and a few of them are lose. Many actin molecules are lost in the minus end and a few of new
molecules are added. Treadmilling occurs when one end of a filament grows in length while
the other end shrinks resulting in a section of filament seemingly “moving” across a stratum
or the cytosol.
26
Motor proteins
Myosin I motors have a short tail domain that can bind an actin filament or a membrane.
Membrane binding, endocytic vesicles.
Myosin II filaments pull actin filaments with opposite polarity past one another, mediating
local contraction. Filament sliding is the mechanism of muscle contraction based on muscle
proteins that slide past each other to generate movement.
Accessory proteins
Polymerization and depolymerization controlling proteins
1. A complex of proteins nucleate assembly to form a web and remains associated with the
minus end.
2. Bind subunits, speed elongation and the filaments’ growth.
3. Stabilize filament.
4. Prevent assembly and disassembly at plus end.
5. Bind ADP-actin filaments and accelerate disassembly.
6. Bind subunits, prevents assembly and slow down filaments’ growth.
27
Cross linking proteins. Membrane binding proteins
Presence on cell structures and processes

Muscle cell contraction is produced by myofibrils, which consist of specific proteins (actin and
myosin).
Amoeboid movement is the most common mode of locomotion in eukaryotic cells. The
formation in a specific direction of pseudopod is due to some filaments of the external and
the internal cytoplasm. The interaction of myosin with the actin network generates contractile
forces for forward motion.
Cytoplasmic streaming
It is when there is flow inside the cytoplasm: it is moving inside the cell. The flow is driven by
forces from the cytoskeleton, and it is likely that the function of flow is at least partly to speed
up transport of molecules and organelles around the cell.
28
INTERMEDIATE FILAMENTS
They are fibrous proteins supercoiled into thicker cables. Its proteins subunits are one of
several different proteins of the keratin family, depending on cell type. Main functions:
maintenance of cells shape (tension-bearing elements), anchorage of nucleus and certain
other organelles, formation of nuclear lamina.
Monomers and assembly
The first stage of filament assembly is the formation of dimers in which the central rod
domains of two polypeptide chains are wound around each other in a coiled-coil structure.
The dimers then associate in a staggered antiparallel fashion to form tetramers. The four
peptides (two carbons and two nitrogen) of the tetramer link with the ones of other tetramer,
forming protofilaments. The final intermediate filament contains approximately eight
protofilaments wound around each other in a rope-like structure. Because they are assembled
from antiparallel tetramers, both ends of intermediate filaments are equivalent.
Consequently, in contrast to actin filaments and microtubules, intermediate filaments are
nonpolar and they do not have distinct plus and minus ends.
29
EXTRACELLULAR COMPONENTS AND CONNECTIONS

Extracellular components and connections between cells help coordinate cellular activities.
Most cells synthesize and secrete materials that are external to the plasma membrane. These
extracellular structures include cell walls of plants, the extracellular matrix (ECM) of animal
cells and intercellular junctions.
The extracellular matrix (ECM) of animal cells
Animal cells lack cell walls but are covered by an elaborate extracellular matrix (ECM). The
ECM is made up of glycoproteins and other macromolecules. Functions: support, adhesion,
movement, regulation. The integrin of the plasma membrane are connected by extrinsic
proteins with the cytoplasm microfilaments. They are also connected by the fibronectin to the
extracellular matrix, formed of collagen and proteoglycan complexes (hyaluronic acid and
proteoglycans).
Intercellular junctions
Neighbouring cells in tissues, organs or organ systems often adhere, interact and
communicate through direct physical contact. This contact is facilitated by intercellular
junctions.
30
Tight junctions are membranes of neighbouring cells that are pressed together, preventing
leakage of extracellular fluid. The membranes are sewn together. There is no space between
cells.
Desmosomes or anchoring junctions are fasten cells together into strong sheets. The plaque
is a structure inside the cell connected with the intermediate filaments that helps to keep cells
linked. However, it is not a total linkage and any fluid can pass through the intercellular space.
Gap junctions or communicating junctions provide cytoplasmic channels between adjacent

cells. They connect physically the cells in order to help in transport processes.
31

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LESSON 7. STRUCTURE AND FUNCTION OF ORGANELLES

Proteins’ signal sequences

ROUGH ENDOPLASMIC RETICULUM: with ribosomes studding its surface.

Type of proteins according to their final destination:

Transmembrane – single domain:

Glycosylation begins with the addition of a 14-sugar oligosaccharide precursor (3 glucose, 9

SMOOTH ENDOPLASMIC RETICULUM: lacks ribosomes.

The nuclear envelope

Modification of lipids (glycosylation).

Control intracellular vesicular traffic

2. Transport from donor to target membrane. It is carried out by microtubules cytoskeleton

3. Arrival to target membrane and fusion. Vesicles go to different proteins.

2. Phagocytosis begins by interaction of phagocytic receptors with ligands on the surface

Storage of ready-to-use transporters of plasma membrane

Other transport proteins

Cellular respiration proteins

Electron transport chain

- Related to replication of mtDNA, transcription of mtDNA and translation of mtRNAm. Most

Polymerization and depolymerization

Cross linking proteins

Microtubules presence on cell structures and processes

Mitotic spindle is formed by the attachment of microtubules to chromosomes` kinetochores.

MICROFILAMENTS (ACTIN FILAMENTS)

Polymerization and depolymerization

Cross linking proteins. Membrane binding proteins

Presence on cell structures and processes

EXTRACELLULAR COMPONENTS AND CONNECTIONS

Gap junctions or communicating junctions provide cytoplasmic channels between adjacent

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