The subcellular organelles provide several benefi ts to the eukaryotic cell. They create compartments within the cell. Each compartment is specialized for a specifi c role.
Major advantages of having compartments are as below:
1. Enhances effi ciency of reactions: In the metabolic pathway, the reactions are sequential, which means that product of a reaction serves as substrate for the next reaction.
Such reactions work much more effi ciently if they are held in close proximity within a compartment.
2. Storage: Specifi c substance can be stored within a dis-tinct intracellular compartment, from where it can be released when the need arises. For example, calcium ions are stored in sarcoplasmic reticulum and released to promote muscle contraction.
3. Reciprocal regulation of opposing pathways: The path-ways that carry out opposing groups of reactions (such as fatty acid synthesis and degradation) do not occur simultaneously. A mechanism for their reciprocal regulation exists by which when one of the pathways is operative, the opposite pathway is inhibited. A smooth working of this arrangement is possible only when the opposite pathways are located in different com-partments. For instance, the pathway for fatty acid syn-thesis is located in cytosol, whereas the pathway for its degradation is located in the mitochondrial matrix.
4. The pH within an organelle can be maintained at a different level than the rest of the cell (e.g. lysosomes). Lysosomes are membrane-bound digestive vesicles of varying sizes and shapes (average diameter 0.5 m) that contain lytic enzymes (called hydrolase) at a relatively low pH (4.5–5.5).
The hydrolases cause intracellular digestion of mac-romolecules. Each lysosome contains over 30 hydro-lases, which can digest all types of biomolecules (e.g.
proteins, nucleic acids, carbohydrates, and lipids).
Table 7.2. Marker enzymes. A given marker enzyme is present within only one compartment of the cell
Subcellular organelle Marker enzyme
Mitochondria Inner membrane: ATP synthase
Lysosome Cathepsin
Golgi complex Galactosyl transferase–mannosidase I (cis); Fucosyl transferase (trans) Microsomes Glucose 6-phosphatase Cytoplasm Lactate dehydrogenase The following steps are involved in subcellular
fractionation:
The tissue is suspended in 0.25 M sucrose solution and the cells are disrupted by means of the shearing forces generated in a potter homogenizer.
The homogenate thus prepared is then subjected to high-speed centrifugation. This process brings about separation of various organelles because different organelles sediment at different speeds in a centrifugal fi eld, depending on their density and size. More the density and the size, more readily does the sedimen-tation take place. To some extent, shape of the subcel-lular organelle also determines its sedimentation.
Centrifugal forces of suffi cient magnitude and duration are used to produce separation of the organelles. Nucleus being the heaviest particle, sediments most readily.
Centrifugation at 104 g/min results in pelleting of nucleus.
Centrifugal force of 4 105 g/min results in sedimen-tation of mitochondria, lysosomes and peroxisomes (Fig. 7.2).
Separation of other organelles is then achieved in the same way, i.e. by increasing the centrifugal force.
At lower centrifugal forces, nuclei, mitochondria lyso-somes and peroxilyso-somes pellet to bottom of the centrifuge tube, whereas higher forces are required to pellet endo-plasmic reticulum, Golgi apparatus and membranes.
Some enzymes, called the marker enzymes, are pre-ferentially located in particular organelles, as shown in
104g/min
4 ×105g/min
6 ×106g/min
Nuclei and fragments of plasma membranes
Mitochondria, lysosomes, and peroxisomes
Microsomal fraction
Cytosol Centrifugal
force applied
Supernatant
Fig. 7.2. Separation of subcellular organelles by differential centrifugation.
(b) maintenance of the shape of cell, (c) cellular movement and
(d) controlling movements of molecules between the inside and the outside of the cell.
Eukaryotic, but not prokaryotic cells, possess intracel-lular membranes as well, which encompass various cell organelles, thus segregating one part of the cell from the other and also enabling each organelle to carry out its characteristic cellular function.
Membranes are very active metabolically and have sig-nifi cant infl uence on cell metabolism. Most membranes are around 50 Å (5 nm) thick. In general, the membranes of organelles are thinner than the plasma membrane. All membranes have similar structural organization, which may be pictured as a mosaic of globular proteins embedded in a fl uid-like phospholipid bilayer.
A number of diverse functions are performed by bio-logical membranes, as outlined below:
Membrane lipids—Diffusion barrier, controlling movement of specifi c molecules, maintain shape of cell.
Membrane proteins—Enzymes, carrier activities, signal transduction, link between cytoskeleton and extracellu-lar matrix.
Carbohydrates—Cell-cell recognition, adhesion and receptor action.
Therefore, lysosomes are referred to as the potential lethal bags of the cell. Since the lysosomal membrane separates these enzymes from the cytosol, the cyto-solic contents cannot be digested by them under nor-mal circumstances. Even when the acid hydrolases leak out of the organelle because of membrane damage, they can cause only minimal damage to the cytosolic contents. This is because their optimum pH is low, and so they are readily inactivated at the relatively high pH prevailing in the cytosol.
Cytoskeleton
Electron microscopic studies have shown that a complex network of protein fi laments is present in the cytoplasm. The fi laments form a fl exible, framework within the cell, called cyto-skeleton. Pivotal role is played by the cytoskeleton in a variety of cellular functions, such as intracellular transport, maintenance of shape of the cell, motility and cell division.
Three types of fi laments are usually found in the mammalian cell: (a) microfi laments of 5 nm diameter, (b) microtubules of 25 nm diameter, mainly present in the long cells of the nervous system, and (c) myosin, which is mainly present in the muscle cells.
Eukaryotic cells have internal scaffold, the cytoskeleton, that control shape and movement of the cell.
Cytoplasm
The cellular matrix in which the subcellular organelles are embedded is known as cytoplasm. It accounts for approx-imately 50% of the cell volume. Cytoplasm was previously believed to be an inert jelly. However, it is now known that cytoplasm contains a variety of enzymes that participate in a number of metabolic reactions. The other components of the cytoplasm are glycogen granules and fat droplets.
Role played by various organelles is given in Table 7.3.
For details the student is advised to refer to a textbook of cell biology.
Cytosol is soluble part of cytoplasm, wherein numerous metabolic reactions take place.
II. Biological Membranes
Biological membranes are large fl exible sheets that are universal elements of cell structure. Each cell is surrounded by a plasma membrane, which forms a boundary between the cell and its environment. It plays important role in:
(a) cell-cell recognition and communication,
Table 7.3. Functions of subcellular organelles Organelle Function
Nucleus Contains chromatin composed of DNA and proteins, RNA synthetic apparatus, and a fi brous matrix
Nucleolus A nuclear subcompartment where most of the cell’s rRNA is synthesized
Endoplasmic reticulum (smooth)
Site of biosynthesis of several biomolecules, detoxifi es certain hydrophobic compounds
Ribosome Protein synthesis
Golgi apparatus Functions in the synthesis, processing and sorting out of secreted proteins, lysosomal proteins and certain membranes
Mitochondrion Principal site of oxidative pathways (e.g.
TCA, -oxidation), ATP production, urea cycle (partly) and haem synthesis (partly) Lysosome Acidic organelle, contains a battery of lytic
enzymes that degrade material internalized by the cell, and worn-out cellular membranes and organelles
Peroxisome Oxidation of long chain fatty acids, D-amino acids and -hydroxy fatty acids
Cytoplasm Site of several metabolic pathways
biological membranes. In 1935, Davson and Danielle sug-gested another model in which phospholipids were the major constituents that formed the matrix or continuous part of the membrane. Few globular proteins were also present, but only towards the polar exterior. A major question with these earlier models was as to how the membrane proteins interacted with the lipid bilayer.
No satisfactory explanation was in sight, which casted doubt on general validity of these models.
In 1972, Jonathan Singer and Garth Nicolson proposed the fl uid mosaic model (Fig. 7.4), in which some pro-teins called intrinsic propro-teins are actually immersed in the lipid bilayer, while others are loosely attached to the sur-face of the membrane, i.e. extrinsic proteins. The impor-tant features of the fl uid-mosaic model, the most accepted model for the overall structure of biological membranes, and supported by a wide variety of experi-mental observations, are excerpted as below:
The bilayer organization of lipids in membranes can be viewed as two dimensional solutions of oriented lipids;
and there are proteins embedded in it. The proteins may be intrinsic (tightly associated with hydrophobic core) or extrinsic (loosely bound to membrane), as discussed.
The integral membrane proteins can be considered as
‘icebergs’ fl oating in two dimensional lipid ‘sea’. The bilayer organization of the lipids acts both as solvent for the amphipathic integral membrane proteins and as a permeability barrier.
Membrane proteins are free to diffuse laterally in the plane of the bilayer unless restricted in some way, but would not be able to fl ip from one side of the bilayer to the other.
Some lipids may interact with certain membrane pro-teins. These interactions are essential for the normal functioning of the protein.
Biological membranes are diffusion barriers and sites of specifi c regulated transport around cells and intra-cellular compartments. They are also involved in cell recognition and intercellular communication, and have many associated enzymes.
A. Chemical Composition of Membranes
Lipids and proteins are the two major components of all biological membranes, as mentioned earlier. Relative pro-portions of the two vary greatly amongst different types of membranes. For example, lipids account for about 20% of the total weight of the rat liver cell membrane to over 54%
in myelin sheath. In most membranes, 50–65% of the total membrane mass is accounted by proteins. Membranes of organelles have a higher percentage of proteins because of their greater participation in enzymatic and carrier (i.e. transport) activities. Inner mitochondrial membrane (IMM) contains highest proportion of proteins (75%).
Other membrane components, present in smaller quantities (5–8%) are glycolipids and glycoproteins.
Carbohydrates do not exist in free form in membranes.
Small quantities of free and esterifi ed cholesterol are also present in most membranes.
The membrane lipids: phospholipids, glycolipids and choles-terol, are amphipathic molecules with hydrophilic heads on the outside and hydrophobic tails oriented inside.
B. The Lipid Bilayer
The bilayer structure of membranes is due to amphipathic nature of the major membrane lipids, i.e. the phospho-lipids. A phospholipid molecule is oriented in such a way that its polar head group is exposed on the external sur-face of the membrane, and the fatty acyl chain is oriented to the inside of the membrane. This forms a sheet-like phospholipid bilayer, which is two molecules thick (Fig. 7.3). The fatty acyl chains of the phospholipids in each layer, or leafl et, form a hydrophobic core that is 2–3 nm thick in most membranes. The diffusion barrier mentioned above is because of this hydrophobic core, which is highly impermeable to polar molecules and ions.
Understanding of the membrane structure has evolved gradually over a period of several years and still new infor-mation is being added. As a result, the proposed model for membrane structure continues to undergo modifi ca-tions and refi nements. It was in 1925 that Gorter and Grendel fi rst proposed the lipid bilayer structure for the
50 Å
Fig. 7.3. The bimolecular leaf arrangement of phospholipids in biological membranes.
actually amphipathic because its hydroxyl group can inter-act with water. It is oriented in such a way in the mem-brane that its hydrophilic hydroxyl group faces the exterior and the cyclopentanophenanthrene ring fi ts into the hydrophobic lipid phase of the membrane.
In membranes three major classes of lipids are phos-pholipids, sphingolipids and cholesterol. The (glycero-) phospholipids have glycerol backbone and the sphingo-lipids are based on sphingosine, an amino-alcohol.
Cholesterol is most rigid lipid in animal cells; in plants
-sitosterol is present instead.
The proportion of lipid component of different membranes vary. Neutral lipids and sphingolipids occur in high con-centration in plasma membranes. The inner mitochon-drial membrane (IMM) is especially rich in cardiolipins and phosphatidylethanolamine. High concentration of sphingolipids is present in myelin sheaths of axons of neural tissue, whereas the intracellular membranes pri-marily contain phospholipids with relatively smaller amount of sphingolipids or cholesterol. The cholesterol content of a given membrane may vary with the nutri-tional status of an individual.