Chemistry of Life

ELEMENTS
 An element is a substance that cannot be broken down to other substances by chemical reactions.
 About 20 – 25% of the natural elements are essential for life.
 Around 96% of the living parts of organisms are made up of: O, H, N and C.

ISOTOPES
 Some atoms have more neutrons than other atoms of the same element.
 Carbon has three isotopes: Carbon-12, Carbon-13 and Carbon-14.

ELEMENTS
 An element is a substance that cannot be broken down to other substances by chemical reactions.
 About 20 – 25% of the natural elements are essential for life.
 Around 96% of the living parts of organisms are made up of: O, H, N and C.

ELECTRONS
 An orbital can contain no more than two electrons.
 The first shell has two electrons, the second has eight, the third had eighteen and the fourth has thirty two.

BONDING
 Covalent Bonds – Sharing of electrons in a covalent compound (Molecule). Can be polar and non-polar.
 Ionic Bonds – Anions and Cations bonded by opposite charges.
 Hydrogen Bonds - electrostatic attraction between polar molecules that occurs when a hydrogen (H) atom bound to
a highly electronegative atom such as N, O or F, and experiences attraction to some other nearby highly
electronegative atom – a particularly strong dipole-dipole attraction.

WATER: MEDIUM FOR LIFE

Specific Heat
Capacity

Heats of fusion
and evaportation
Hydrogen Bonding
Ice floats on water
Properties - density
Capillary Action
Cohesion /
Adhesion
Surface Tension

Solvent Properties
Polar Covalent
Hydrophillic
Bonds
Hydrophobic
 Cell Structure
MICROSCOPY
 The human eyes limit of resolution is about 200 µm, and most cells are 1 µm to 100 µm.
 Light Microscope: Uses glass lenses to focus visible light onto an object and collects the light that passes through it.
The lenses refract the light so the image is magnified as it is projected into the eye. Limited resolution of 250 nm –
100x objective lens, 8x ocular lens. Cells need to be fixed prior observation via rapid freezing or chemical fixations.
 Electron Microscope: Use magnets to focus a beam of electrons onto a preserved tissue. Its resolution is 2nm, a
hundred times better than the light microscope.
 Transmission Electron Microscope: Electrons pass through thin sections of preserved tissue and are projected on a
phosphorous screen. Usually stained with heavy metals that attach to certain organelles - enhances electron density.
 Scanning Electron Microscope: For detailed study of the surface of a specimen. The electron beam scans the surface
of the sample – usually coated in a thin film of gold. The cream excites the electrons on the surface – these
secondary electrons are detected by a device and translates the pattern of electrons to a 3D image.

PROKARYOTIC CELLS
 Lack a membrane bound nucleus. The DNA is in an unbounded region of the nucleoid.
 Are around 1 µm and include those of the Archaea and Bacteria. No membrane bound organelles.
 Cytoplasm bounded by the plasma membrane.
 All cells however contain: Plasma membrane, semifluid substance – cytosol, chromosomes and ribosomes.

EUKARYOTIC CELLS
 Contain a membrane bound nucleus and membrane bound organelles.
 The cytoplasm is in the region between the plasma membrane and nucleus.
 Organelles and other substances lie in the cytosol.
 Cytosol – is an aqueous solution of molecules with a gel like consistency.
 Cytoplasm – consists of cytosol and subcellular components; excluding the nucleus.
 Protoplasm – is the cytoplasm and the nucleus.

EUKARYOTES VS. PROKARYOTES

FEATURE EUKARYOTE PROKARYOTE
Size 10-100 µm 1-20 µm
NON Ribosomes All have All have
MEMBRANOUS Cytoskeleton All have Some have
Nucleus All have
Endoplasmic Recticulum All have
Golgi Bodies Some have
Lysosomes Some have
MEMBRANOUS Vacuoles Some have
Vesicles All have Absent in all
Mitochondria Present in most
Chloroplasts Plants, algae and some protozoa.
ANIMAL CELL VS. PLANT CELL
FEATURE ANIMAL CELL PLANT CELL
Cell wall X 
Chloroplast X 
Central Vacuole X 
Plasmodesmata X 
Lysosomes  X
Centrosomes with Centrioles  X
Flagella  X
Plasma Membrane  
Nucleus  
Ribosomes  
Endoplasmic Recticulum  
Golgi Apparatus  
Mitochondria  
Cytoskeleton  

NUCLEUS
 Surrounded by a double membrane – the nuclear envelope.
 Contains one or several nucleoli – contain high concentrations of RNA, protein and DNA.
 It is perforated at intervals – nuclear pores – channels that allow movement of certain molecules.
 In the nucleus, the DNA is organized in discrete units called chromosomes.

RIBOSOMES – PROTEIN FACTORIES

 Are complexes made up of rRNA and protein – cellular components that carry out protein synthesis.
 The cytoplasm of eukaryotic cells contain several million ribosomes.
 They are small granular structures of around 25 – 30 nm
 In Prokaryotes they are free and in Eukaryotes they are bound in the Endoplasmic Recticulum.
 Composed of two subunits that are assembled in the nucleolus from RNA and protein molecules. The subunits move
through the nuclear pore into the cytosol where it associates with mRNA to be a functional ribosome to undergo
protein synthesis.

ENDOMEMBRANE SYSTEM
 Endomembrane system regulates protein traffic and performs metabolic functions in the cell.
 Components of the system: nuclear envelope, endoplasmic recticulum, Golgi apparatus, lysosomes, vacuoles, plasma
membrane. These components are continuous or are connected via transport vesicles.

ENDOPLASMIC RECTICULUM – BIOSYNTHETIC FACTORY

 The ER is a network of membranous sacs (cisternae) that extends throughout the cytoplasm of eukaryotes.
 ER accounts for more than half of the total membrane in eukaryotes.
 ER membrane is continuous with the outer membrane of nuclear envelope.
 Two regions of the ER: Smooth ER which lacks ribosomes, and the Rough ER which has ribosomes.
SMOOTH ENDOPLASMIC RECTICULUM
 It synthesizes lipids, metabolizes carbohydrates and detoxifies drugs and poisons. Also stores calcium ions.

ROUGH ENDOPLASMIC RECTICULUM

 Has bound ribosomes giving it a ‘rough’ appearance.
 The ribosomes are involved in the synthesis of proteins.
 Proteins are glycosylated (covalently bonded to carbohydrates) in the ER lumen.
 It also distributes transport vesicles; proteins surrounded by membranes.
 It is a membrane ‘factory’ for the cell.

GOLGI APPARATUS
 The Golgi apparatus consists of flattened membranous sacs called
cisternae.
 The Golgi stack is usually surrounded by a cloud of small vesicles.
 Each stack has a distinct polarity
- Cis Face (Receiving): Faces the cisternae of the ER.
- Trans Face (Transporting): Opposite side of the Golgi stack
 Proteins & glycoproteins enter each Golgi stack at the cis face via
transport vesicles which bud off the ER membrane.
 These transport vesicles fuse with the cis cisterna.
 Functions of the Golgi apparatus:
- Modifies products of the ER
- Manufactures certain macromolecules (e.g. Pectin)
- Sorts and packages materials into transport vesicles

LYSOSOMES
 Is a membranous sac of hydrolytic enzymes than can digest macromolecules. Lysomal enzymes can hydrolyze

proteins (proteases), fats (lipases), polysaccharides (glycosidase) and nucleic acids (nucleases).

VACUOLES
 Not limited to plant cells. They are membrane bound vesicles and functions vary in different types of cells.
 In a plant cell: Contain hydrolytic enzymes, store nutrients, pigments or waste material, and are involved in the
maintenance of cell turgor pressure.
MITOCHONDRIA
 Not part of the endomembrane system.
 It has free ribosomes and circular DNA.
 Grow and reproduce somewhat independently in cells.
 Evolved from bacteria (prokaryotes) that were engulfed by
ancestral eukaryotic cells.
 Site for cellular respiration – release of energy during
oxidation of sugars and fats – also known as oxidative.
 Phosphorylation – released energy is stored as ATP.
 They are large organelles, 1-10 µm long, and can be seen in a light microscope. Can be spherical or elongated.
 It has a double membrane, the outer membrane is smooth but the inner membrane is convoluted with foldings
called cristae. The inner membrane is lined with numerous knob like structures – enzyme complexes responsible for
ATP synthesis.
 The core – matrix – contains mitochondrial ribosomes, mitochondrial DNA and structural proteins.

CHLOROPLASTS
 Member of a family of organelles called plastids.
 They contain the green pigment chlorophyll, as well as enzymes and
other molecule that function in photosynthesis.
 Highly developed internal membranes. Structure as follows:
- Thylakoids – membranous sacs, stacked to form a
granum.
- Stroma – internal fluid. Contains chloroplast DNA,
ribosomes and enzymes.
 Are mobile – move via the cytoskeleton.

PEROXISOMES: OXIDATION
 Are specialized metabolic compartments bounded by a single membrane. Perform reactions of many different
functions. Produce H2O2 and convert it to H20.

CYTOSKELETON
 Is a network of fibres that organize structures and activities in the cell.
 Components:
- Microtubules (Composed of tubulin) – flagella, cilia, centrosomes, centrioles, organelle movement.
- Microfilaments (Composed of actin) – smallest; support shape, muscle contraction.
- Intermediate filaments (Several types made up of a distinct protein) – medium size; anchor
nucleus/organelles, shape, nuclear lamina.
 Involved in maintenance and remodeling of cell shape.

CELL WALL
 Made up of cellulose fibres embedded in other polysaccharides and protein.
 It protects plant cell, maintains its shape and prevents excessive uptake of water.
ANIMAL CELL AND PLANT CELL

Proteins
PROTEIN STRUCTURE
 Amino Acid:

 Hydrophobic amino acids: non-polar side chains

 Hydrophilic amino acids: polar side chains
- Acidic: -vely charged
- Basic: +vely charged
 Polypeptide Formation:

 Primary Structure: Secondary Structure: Tertiary Structure: Quaternary Structure:

SICKLE HEMOGLOBIN VS. NORMAL HEMOGLOBIN
 Sickle Cell Anemia is caused by abnormal protein, haemoglobin, in red blood cells.
 Haemoglobin is globular, containing four polypeptide subunits.

Carbohydrates
IMPORTANCE OF CARBOHYDRATES
 Energy storage and distribution
 Structure components in plants (Cell wall) and arthropods (exoskeleton), animals (cartilage) and bacteria (cell wall).
 Biosynthetic precursors to amino acids and nucleic acids.
 Used on glycoproteins as addresses for intracellular trafficking.
 Antigenic (For example: ABO blood group specificity determinants are carbohydrates)

MONOSACCHARIDES
 Simplest carbohydrates consist of single molecules with formula (CH20)n. Transport form of energy.
 Classified based on number of C atoms: triose (3), tetrose (4), pentose (5), hexose (6) and heptose (7).
 Depending on the location of a carbonyl group: ALDOSE: Located at C1 and KETOSE: Located at C2
 Diversity attributed to the spatial arrangement around asymmetric carbons (Carbons attached to four different
atoms or groups of atoms).
 Readily soluble due to the ability to form H-bonds, through O atoms, to form H20 molecules.


DISACCHARIDES
 Consists of two monosaccharides joined by a glyosidic linkage; a covalent bond formed between the two by a
dehydration reaction.
 Used in transport and storage.

POLYSACCHARIDES
 A few hundred to a few thousand monosaccharides joined by
glycosidic links. Starch in Plants
 Major functions include being a structural component in the Storage Forms
Glycogen in
walls of plant, fungi and bacteria. In association with proteins – Animals
structural components of connective tissue and exoskeleton of Polysaccharides
animals. Is a storage reserve in all animals. Cellulose in Plants
 Can be Homo-polysaccharides or Hetero-polysaccharides in Structural Forms
regards to their monomers. Chitin in
Arthropods
 Can be Un-branched, linear, or it can be branched.

STARCH
 Primary product of photosynthesis: 3 carbon compound; glyceraldehyde – 3 – phosphate, converted to sucrose for
transport or starch for storage.
 Found in granules in plant chloroplasts – transient storage material.
 Also found in tubers, seed endosperms and cotyledons – long term energy storage.
 Starch is amylopectin (70-80%) and amylose (20-30%).

GLYCOGEN
 Polymer of glucose similar to amylopectin but more extensively branched.
 Each glycogen molecule is attached to a core protein – glycogenin. This is more compact, so it takes up less space for
storage which is an advantage for animals on the move.
 Present in all tissues but particularly more abundant in the liver and muscles.
CELLULOSE
 Unbranched β – glucose polymer. Parallel chains are held together by H bonds between the hydroxyl groups
attached to carbon atoms 3 and 6.
 Around 80 cellulose polymers associate to form a microfilm.
 Few organisms have cellulose to break it down. Cellulose abrades the wall of the digestive tract and stimulates lining
to form mucus which helps in the smooth passage of food.

CHITIN
 Exoskeleton of arthropods and cell wall of fundi. Is similar to cellulose but the glucose monomer has a nitrogen
containing appendage.
 Associates laterally to form microfibrils.

Lipids and Membranes

FATS/TRIGLYCERIDES
 Lipids are insoluble in water. Polar molecules = hydrophilic. Non-polar = hydrophobic.
 Fatty acids: are carboxylic acids with long hydrocarbon tails.
 Act as energy reserves – stored in adipose tissue (animals) and seeds/fruit (plants).

PHOSPHOLIPIDS AND GLYCOLIPIDS

 Composed of long chain fatty acids esterified to two of the three -OH groups of glycerol, with the third occupied by:
PHOSPHOLIPID: GLYCOLIPID:

CHOLESTEROL
 Composed of four rings of carbon with a hydroxyl group at one end
and a hydrocarbon tail at the other.
 Rigid Planar Steroid Ring structure.

MEMBRANES
 All membrane lipids (Phospholipids, cholesterol, glycolipids…) have non polar hydrophobic tail and a polar
hydrophilic head.
 Phospholipids are AMPHIPATHIC: hydrophobic tails and hydrophilic heads.
 When in water, these lipids spontaneously rearrange to form stable structures and a molecular bilayer.
 Membranes are fluid mosaics of lipids and proteins. Membrane proteins “float” in the lipid bilayer.
 Carbohydrates (sugars) attach to proteins or lipids at the outer surface (cell surface recognition).

MEMBRANE FLUIDITY

MEMBRANE PROTEINS
 Peripheral Proteins: Loosely attached by non-covalent interactions.
 Integral Proteins: Transmembrane – extending through membrane.
 Membrane Protein Functions: Transport, enzyme activity, signal transduction, cell-cell recognition, intercellular
joining, attachment to cytoskeleton and extracellular matrix.

MEMBRANE PERMEABILITY
 Controls passage of substance into and out of a cell – maintain stable conditions, homeostasis, movement in and out
of organelles.
 Permeability of molecules depend on: size, electrical charge, lipid solubility.
 H20, O2 and CO2 cross freely.
 Ions and other polar molecules cross only at selective pores formed by transmembrane proteins. Integral proteins
allow facilitate diffusion (channels, carriers) and active transport (pumps, co-transport).

DIFFUSION
 Is generally hydrophobic. Is the passive movement of molecules along a concentration gradient from high to low.
  Rate depends on permeability and magnitude of the concentration gradient. Substances in solution have net
movement down a concentration gradient to reach equilibrium.
 Osmosis: Movement of water by diffusion. Movement is towards low water concentrations.
- ISOTONIC: same [solute] and [H2O]
- HYPERTONIC: high [solute] and therefore lower [H2O]
- HYPOTONIC: low [solute] and therefore higher [H2O]

FACILITATED DIFFUSION
 Is less hydrophobic. Distinguished from simple diffusion by: faster, transport proteins become saturated as substrate
concentration increases, are particular for specific substrates, transport is inhibited by similar substrates that
compete binding sites.
 CHANNELS: act as pipes; allow direct passage from one side to the other. Movements are rapid, faster than carriers.
Most transport ions. High specifity.
 CARRIERS: bind to solute on one side of the membrane – which causes conformational change within the protein
which exposes the solute on the other side of the membrane where it is then released.

ACTIVE TRANSPORT
 Transport against a concentration gradient which requires
energy – normally ATP.
 PRIMARY ACTIVE TRANSPORT: involves direct coupling of
energy from ATP hydrolysis to the movement of solvent
against its electrochemical gradient.
 SECONDARY ACTIVE TRANSPORT – CO TRANSPORT:
1. Electrochemical gradient generate for an
ion against it’s concentration gradient is
used.
2. Downhill movement back across the
membrane can be used to drive the uphill
movement of another solute against its
electrochemical gradient – co transport.

Cell – Cell Interactions

EPITHELIA TISSUE
 Form continuous layers covering surfaces, cavities and tubes inside the body.
 They provide protection, regulate exchange of materials, secretion (glandular tissue).
 Bound together by tight anchoring junctions.
 Categorised by number of layers (1 = simple, 2+ = stratified), shape of cells (Squamous = flattened, cuboidal and
columnar)

CONNECTIVE TISSUE
 Provides basic structural, metabolic and defensive support. Includes bone, cartilage, blood, adipose tissue and
fibroblasts.
  Extracellular material – matric of polysaccharides and proteins.
 Fibres – collagen, reticulin and elastin.

MUSCLE TISSUE
 Are capable of contraction. Composed of actin and myosin filaments. Allows movement of animal and of internal
organs.
 Striated muscle – highly organized, skeletal and cardiac muscle.
 Smooth muscle – less regularly arranged than striated muscles; internal organs.

NERVOUS TISSUE
 Carry information via interconnecting network. Information
receive by dendrites transmitted along the axon.
 Supported by glial cells – which maintain composition of
extracellular environment which forms the myelin sheaths.

EXTRACELLULAR MATRIX
 Defining structure of animal cells. When plasma membranes are not in direct contact there is a fluid lattice network
of protein and polysaccharides between the separate membranes. This ECM is secreted by cells.

}
 INTERSTITIAL MATRIX – found in connective tissue Both forms consist of strong protein fibres in a hydrated polysaccharide gel.
 BASAL LAMINA – underlies epithelial tissue
 Three main components:
- Structural proteins (collagen, elastin)
- Adhesive proteins (fibronectin, liminin, others)
- Polysaccharides (glycosaminoglycans).

COMPONENTS OF THE EXTRACELLULAR MATRIX

 Structural Proteins:
- Collagen: three polypeptide chains that
entwine in a tight helix. In the Interstitial
Matrix with fibrils 30nm in diameter. In
the Basal Lamina it forms a strong sheet
like meshwork.
- Elastin: unfolded in a coiled configuration
it can be stretch extensively and once
released it recoils into original shape.
 Adhesive Proteins:

- Laminin (Basal Lamina) }

- Fibronectin (Interstitial Matrix) Both mediate attachment of cells to the matrix – affecting cell polarity, growth,
migration and differentiation.
 Polysaccharides:
- Glycosaminoglycans (GAGs) are usually linked to a protein core. They are long unbranched
polysaccharides. Consist of repeated disaccharide units. Have strong negative charge that attracts
cations, such as water.
CELL COMMUNICATION: DIRECT CONTACT – INTERCELLULAR JUNCTIONS
 Cells not separated by ECM are connected by complexes called junctions

TIGHT (Occluding) Junctions ANCHORING Junctions COMMUNICATING Junctions

- Prevent passage of molecules - Provide mechanical support. - Are gap junctions specialized for
through extracellular space. - Most common = desmosomes. electrochemical communication
- Prevent fluid from moving across a - On cytoplasmic face of each between cells.
layer of cells. plasma membrane is a thickening - Provide pathway of low electrical
- They link epithelial cells via a series called a plaque. resistance.
of integral membrane proteins that - Adjacent cells are held together by - Permit rapid current spread
fuse to form impermeable junctions. think linker glycoproteins between cells.
- Help stop molecules passing to (cadhenins) that link plaque to - Plasma membranes very close and
interstitial space, for example in the plaque. connect by hollow cylinders called
digestive tract stop enzymes seeping - Plaques inside cells are linked by connexons (composed of
out into blood stream. protein filaments of cytoskeleton. transmembrane proteins).

SIGNAL TRANSDUCTION PATHWAY

TYPES OF SIGNALS
 PARACRINE signaling (Local): secreting cell acts on nearby target cell by discharging molecules into extracellular fluid.
 SYNAPTIC signaling (Local): nerve cell releases neurotransmitter into a synapse.
 ENDOCRINE signaling (Long Distance): specialized endocrine cells secrete hormones.

Enzymes
ENZYMES
 Proteins (mostly) – 3D structure and biological catalyst.
 Enzymatic reactions stabilize transition state and reduce the activation energy require by reactants (substrates).
 Active site can lower the Ea / speed up the reaction by acting as a template for substrate orientation, stressing
substrates and stabilizing the transition state, providing favorable microenvironment and/or participating directly in
the catalytic reaction.
  Enzyme Specificity: Due to the 3D structure of the active site – a specialized region formed from the folding of
polypeptide chains. The site is lined by R – groups of catalytic amino acids. Substrates – binding amino acids;
arrangement determines specificity.
 Enzyme Activity:
- Affected by temperature and pH; outside the optimum range they lose secondary and tertiary
structure (denatured); loosing specificity and thereby loss of activity.
- Also affected by co-factors (helper molecules).
- Affected by inhibitors: Competitive Inhibition – mimics substrate; competition.
Non-competitive Inhibition – binds away from active site but it alters its shape
thereby reducing activity.
 Allosteric Modulation: Allosteric sites are ones other than active site. Molecule that can bind here can cause chain in
conformation of the enzyme which alters the active site. These interactions can inhibit enzyme by preventing
substrate binding or activate enzyme by allowing substrates to bind (Common ways of metabolic control).
 Enzyme activity also controlled by: enzyme production (transcription and translation), compartmentalized (occurring
in cellular compartments), inhibitors, activators, post translational modifications (folded or molded after creation).
 Types of Enzymes:
- Transferases and ligases: biosynthesis of cellular constituents
- Hydrolases: breakdown complex molecules
- Lyases & Isomerases: in pathways that transform compounds into substrates for oxidoreductases.
- Oxidoreductases: trap potential energy by coupling reactions with ATP formation.

Metabolism
METABOLISM
 Refers to all activities and chemical reactions in an organism’s body.
 ANABOLISM: biosynthesis of molecules – consumes energy.
 CATABOLISM: breakdown of molecules – releases energy.
 Metabolic Rate: the rate at which an organism uses fuel to supply ATP for these reactions – ‘The Cost of Living’. Rate
varies due to food & water availability, environmental temperature, energy expenditure, reproductive status & age.

ENERGY
 Potential energy is stored – such as chemical energy stored in bonds between atoms.
 Kinetic energy is expressed as motion and is sued to allow work to occur. Heat energy is associated with the random
movement of atoms or molecules.
 FIRST LAW OF THERMODYNAMICS: energy can neither be created nor destroyed. The total energy of the universe
remains constant.
 Bioenergetics: Much of work involves the transfer of one energy from one form to another. No energy transfer is a
100% efficient – some is always lost as heat; unavailable to do work.
 SECOND LAW OF THERMODYNAMICS: The entropy of the universe is always increasing.
 Energy converted in cells is only 30% efficient, the rest of the energy is lost as heat.

FREE ENERGY AND ENZYME REACTIONS

 The change in free energy, ΔG, is the chemical potential of a reaction.
  Energy from an exergonic reaction must be coupled to an endergonic reaction. The sum of these two ΔG’s must be
negative.
EXERGONIC REACTION ENDOGONIC REACTION

SPONTANEOUS REACTIONS
 During a reaction there is a transition state which generally has more
energy than the reactants and products.
 Energy required to reach this state is called the activation energy.
 This energy is reduced by enzymes.

ENERGY COUPLING
 Endergonic reactions are often driven by exergonic reactions = energy coupling.
 Many endergonic reactions are power by the hydrolysis of ATP.
A + B  C + D + Energy } Exergonic
↓ Energy + E + F  G + H } Endergonic

ATP: ADENOSINE TRIPHOSPHATE

 The energy released by the hydrolysis of ATP is used to drive many cellular processes via energy coupling

HOW ANIMALS GET ENERGY

 Catabolism (breakdown) of fuel molecules by cellular respiration.
 Energy extracted from C – C, C – H and C- N bonds of fuel molecules. These bonds are in a reduced state because
electrons not shared with oxygen. They contain more extractable energy than oxidized bonds – redox.
 Energy from reactions in which reduced bonds in fuel molecules are oxidized is conserved in ATP.
 Cellular Respiration
CELLULAR RESPIRATION
 Cellular respiration is the metabolism of fuel molecules with the consumption of oxygen and production of carbon
dioxide and water.
 Main steps are:
1. GLYCOLYSIS: breakdown glucose into two molecules of pyruvate.
2. CITRIC ACID CYCLE: completes breakdown of glucose.
3. OXIDATIVE PHOSPHORYLATION: accounts for the most of the ATP synthesis.
 Storage polysaccharides (starch or glycogen) are hydrolyzed into glucose, then:

 Redox Reactions: Energy not harnessed from catabolism of fuel molecules in a single step but rather many with
electrons being stripped at key steps.

 Electrons often passed to storage/carrier/shuttle molecules as an intermediate to the production of ATP. This
includes: Nicotinamide adenine dinucleotide – NAD+ and Flavin adenine dinucleotide - FAD

GLYCOLYSIS
 Glucose  Pyruvate (6 Carbon  2 x 3 Carbon) [Net yield of 2ATP and 2NADH]
 Occurs in the cytosol where the reactants such as ADP, NAD+ and Phosphates, are readily available.
 Series of reactions catalyzed by specific enzymes. No oxygen is required.
FIRST PHASE:

SECOND PHASE:
ACETYL COA – PYRUVATE OXIDATION
 Pyruvate  Acetyl coenzyme A via oxidative
decarboxylation.
 Formation of 2C acetyla CoA and CO2 from 3C
pyruvate.
 One mole of NADH is produced per pyruvate
(Which = 2 per glucose).
 In the presence of O2 pyruvate enters the
mitochondrion and is converted to acetyl CoA.

CITRIC ACID CYCLE

 Eight steps – each catalyzed by a specific enzyme. Every turn of the cycle = the breakdown of acetyl (2C) and the
formation of 2CO2, 1ATP, 3NADH and 1FADH2 } Therefore for every glucose molecule: 4CO2, 2ATP, 6NADH, 2FADH2
 Occurs in the mitochondrial matrix.

OXIDATIVE PHOSPHORYLATION
 During glycolysis and the citric acid cycle, electrons are temporarily stored as NADH and FADH2
 This energy stored is converted to ATP via oxidative phosphorylation.
 FIRST STAGE: Electron Transport
NADH + FADH2 As electrons pass from one comples
O2 is the final
transfer electrons to Regenerate to another, protoons are pumped
electron acceptor -
carrier proteins of the s NAD+ and across the inner mitochondrial
which eventually
electron transport FAD membrane. Proton Gradient = Proton
forms H2O
system Motive Force

 STAGE TWO: Chemiosmosis

- Couples electron transport chain to ATP synthesis.
- Protons move back into the matrix through the ATP synthase complex.
- Drives the manufacture of ATP.
 MITOCHONDRIA:

FERMENTATION AND OTHER ENERGY SOURCES

 FERMENTATION: Anaerobic = no O2
- Stage 1. Glycolysis
- Stage 2. Pyruvate converted to either: Lactic acid (animals) or Ethanol (Plants and some microorganisms)
- Overall: Highly inefficient, only 2ATP formed from one molecule of glucose.
 OTHER SOURCES: Lipids – hydrolysed to glycerol and fatty acids; undergo β – oxidation.
Proteins – hydrolysed to amino acids that feed into pyruvate, acetyl CoA and citric acid cycle intermediates.
 Cell Division
PROKARYOTES: CELL DIVISION
 Bacteria mainly reproduce by binary fission.
 Single circular DNA molecule attached to plasma
membrane.
 DNA replicates, new molecule attached to
separate point of plasma membrane.
 Membrane between two molecules lengthens.
 Plasma membrane and cell wall grow inwards.
 Cell divides: two identical daughter cells.

EUKARYOTES: CELL DIVISION

 Complex due to multiple chromosomes, all must be equally
distributed. Synthesis of DNA and other molecules occur during
interphase. Interphase:
- G1 (First Gap) Phase: Cell growth, determines if
cell is ready for DNA synthesis – checkpoint.
- S (DNA Synthesis) phase
- G2 (Second Gap) phase: Cell continues to grow;
another checkpoint to ensure DNA synthesis is
complete and ready for mitosis.

CHROMOSOMES
 Each chromosome is one linear strand of DNA.
 Somatic cells have two sets of chromosomes. Gametes have half.
 In a normal resting cell, chromatin (DNA + Protein) is thinly strung out – not visible
in a light microscope.
 When a cell divides the chromatin becomes supercoiled and condensed – visible in
a light microscope.

SISTER CHROMATIDS
 Chromosomes distributed and daughter cells formed
during mitosis and meiosis.
 Two sister chromatids: joined copies of the original
chromosomes.

HUMAN CHROMOSOMES
 Karyotype is the term used to refer to the chromosomal constitution of an individual.
 Diploid = 23 pairs, Haploid = 46 in total. One chromosome in each pair comes from each parent chromosomes –
homologous. Twenty two pairs are autosomal, and one pair is sex determining.
CHROMATIN
 Eukaryotic chromatin is composed of a linear double stranded DNA wrapped around proteins called histones.
Supercoiling condenses DNA.
 Basic unit of chromatin is called nucleosome.
 DNA is twisted into helical strands by histone proteins that further condense or supercoil by continuing to wrap
around each other.
 During S phase, the chromosomes are duplicated but held together. During cell division, supercoiling continues until
the condensed chromosome can be seen.

MITOSIS
 Cell Cycle Control & Cancer
CELL CYCLE REGULATED BY MOLECULAR CONTROL SYSTEM
 The frequency of cell division varies with the type of
cell. These differences result from regulation at a
molecular level.
 Cancer cells manage to escape the usual controls on
the cell cycle. The cell cycle appears to be drive by
specific chemical signals present in the cytoplasm.
 The sequential events on the cell cycle are directed by a
distinct cell cycle control system, similar to a clock.
 The clock has specific checkpoints where the cell cycle
stops until a ‘go-ahead’ signal is received.
 Proteins called cyclins promote progress through the
cell cycle.
 Signals from cellular surveillance mechanisms report
whether crucial processes have occurred. Cell cycle arrest allows time for example, repairing of DNA.

MATURATION PROMOTING FACTOR

 Two types of regulatory proteins are involved in cell cycle control: cyclins and cyclin-dependent kinases (Cdks)
 MPF is a cyclin-Cdk complex that triggers a cell’s passage past the G2 checkpoint into the M phase.
 Most important is the G1 checkpoint – Continue to S, G2, M or exit to G0 (Non-dividing state)
 Growth factors (proteins released by some cells), activate cyclins, and stimulate cells to divide.

DEVELOPMENT OF SPECIALISED CELL TYPES

 Stem cell – Pluripotent: can become any type of cell.
 Progenitor Cell – Partially differentiaed cells, capable of producing multiple tissue types.
 Differenciated Cell – Specialised function. E.g. Neuron. Normally quiescent (G0) – exists the cell cycle.

APOPTOSIS: PROGRAMMED CELL DEATH

 Apoptosis is regulated by complex cell signals.
 Cell signals can activate or inhibit apoptosis.
 Cell signals may be extracellular or intracellular.
 Normal tissue development requires both cell division and apoptosis. During embryonic development, the hand first
appears as a paddle. The normal hand is formed by apoptosis of cells between the figures.

CANCER
 Cancerous cells grow in an unregulated fashion and can invade and damage other tissue due to a loss of cell cycle
control, loss of apoptosis and a loss of cell differentiation.
 Cancer cells eventually form a mass of tissue called a tumour: Benign – growth is limited and cells stay together.
Malignant – cells can spread (metastasise). Cancer cells do not respond normally to the body’s control mechanisms
 Cancer cells may not need growth factors to grow and divide
 They may make their own growth factor
 They may convey a growth factor’s signal without the presence of the growth factor
 They may have an abnormal cell cycle control system

METASTASIS
 Spread of cancer cells to locations distant from their original site.
 Accumulation of genetic changes with time: Benign growth  Malignant, metastatic tumour.
 Tissue invasion can lead to circulation of tumour cells in lymphatic or blood vessels. Results in formation of multiple
secondary tumours, distant from the site of the primary tumour. Why early detection and treatment is critical.

ANCHORAGE DEPENDENCE
 Most animal cells exhibit anchorage dependence, in which they must be attached to a substratum in order to divide.
 Density dependence inhibition – in which crowded cells stop dividing.
 Cancer cells exhibit neither density-dependent inhibition nor anchorage dependence.

MUTATIONS CAUSE CANCER

 Contribution from genes and environment.
 DNA damage acquired throughout life leads to mutations in particular cells (Somatic mutations).
 Inherited mutations may predispose an individual to cancer.

MUTATIONS IN ONCOGENES AND TUMOUR SUPPRESSOR GENES CAUSE CANCER

 Two major classes of genes frequently mutated in cancer cells are:
- Proto-oncogenes: code for proteins that promote the cell cycle. Mutations can signal uncontrolled cell
division.
- Tumour Suppressor Genes: Code for proteins that inhibit the cell cycle or promote apoptosis.
Mutations can lead to loss of function.

CELL CYCLE STIMULATING PATHWAY

CELL CYCLE INHIBITING PATHWAY

TUMOUR SUPPRESSOR P53
 Critical for the response to DNA damage within the cell
 Activates DNA repair  Arrests the cell cycle to allow time for repair  If DNA cannot be repaired; promotes
apoptosis.

MULTISTEP MODEL OF CANCER DEVELOPMENT

 Cancer results from genetic changes that affect cell cycle control
 Many proto-oncogenes and tumour suppressor genes encode components of growth stimulating and growth-
inhibiting signalling pathways
 In the multistep model of cancer development, normal cells are converted to cancer cells by the accumulation of
mutations affecting proto-oncogenes and tumour suppressor genes
 Multiple mutations are generally needed for full-fledged cancer; thus the incidence increases with age
 At the DNA level, a cancerous cell is usually characterised by at least one active oncogene and the mutation of
several tumour-suppressor genes

TREATING BREST CANCER

 1/8 women will develop breast cancer but mortality rates are falling due to earlier detection & improved treatment.
 Genetic testing for mutations in susceptibility genes (BRCA1, BRCA2)
 Targeted chemotherapy. Tyrosine kinase receptors (stimulate cell division), eg HER2 (herceptin), ER tamoxifen)

Meiosis
HEREDITY
 Meiosis involves two successive divisions:
1. Reductional division - chromosome no. is halved
2. Equational division - sister chromatids separate
 Heredity is the transmission of traits from one generation to the next
 In asexual reproduction, a single individual passes genes to its offspring (clones are genetically identical individuals
from the same parent). In sexual reproduction, two parents give rise to offspring that have unique combinations of
genes inherited from the two parents
 SEX CHROMOSOMES: Human females have a homologous pair of X chromosomes (XX). Human males have one X and
one Y chromosome. The remaining 22 pairs of chromosomes are called autosomes.
 HOMOLOGOUS CHROMOSOMES: Each pair of homologous chromosomes includes one chromosome from each
parent. The 46 chromosomes in a human somatic cell are two sets of 23: one from the mother and one from the
father. A diploid cell (2n) has two sets of chromosomes. For humans, the diploid number is 46 (2n = 46).
MITOSIS VS. MEIOSIS
 As in mitosis, the chromosomes replicate before meiosis begins. Sister chromatids
form when DNA condenses.
 Meiosis is distinct from mitosis in the following ways:
- Meiosis takes place in two sets of cell divisions, called meiosis I and
meiosis II
- The two cell divisions result in 4 daughter cells rather than 2 in mitosis
- Each daughter cell has only half as many chromosomes as the parent
cell In meiosis there is pairing of homologous chromosomes to form a
tetrad (sister chromatids from paternal and maternal homologues).

TWO STAGES OF MEIOSIS

MEIOSIS I
 Interphase: Cell checks for complete DNA replication before proceeding into meiosis. Nuclear envelope, two
centrosomes (organize microtubules), chromosomes duplicated, uncondensed.

Prophase 1 Metaphase 1 Anaphase 1 Telophase 1 & Cytokinesis

MEIOSIS II

Prophase 2 Metaphase 2 Anaphase 2 Telophase 2 & Cytokinesis

GENETIC VARIATION
 Mutations (changes in an organism’s DNA) are the original source of genetic diversity
 Mutations create different versions of genes (alleles)
 Reshuffling of alleles during sexual reproduction produces genetic variation
 INDEPENDENT ASSORTMENT: Homologous pairs of chromosomes orient randomly at metaphase I of meiosis. Each
pair of chromosomes sorts maternal and paternal homologues into daughter cells independently of the other pairs.
For humans (n = 23), there are more than 8 million (223) possible combinations of chromosomes!
 CROSSING OVER DURING SYNAPSIS: Genetic material may be exchanged between homologous chromosomes via
crossing over. The point where crossing over occurs is a
chiasma (can be one or more points). X-shaped regions.
This is a mechanism for producing increased genetic
variation. Produces recombinant chromosomes, which
combine DNA inherited from each parent. Begins very
early in prophase I, as homologous chromosomes pair
up gene by gene. Homologous portions of two non-
sister chromatids trade places. Contributes to genetic
variation by combining DNA from two parents into a
single chromosome.
 RANDOM FERTILISATION: Any sperm can fuse with any ovum (unfertilised egg). The fusion of two gametes (each
with 8.4 million possible chromosome combinations from independent assortment) produces a zygote with any of
about 70 trillion diploid combinations
UNIQUE FEATURES OF MEIOSIS
 Synapsis Homologous pairing of replicated chromosomes physically connect.
 Genetic information can be exchanged between paired homologous chromosomes via crossing over.
 At the metaphase plate there are paired homologous chromosomes (tetrads) instead of individual replicated
chromosomes.
 At anaphase I it is homologous chromosomes instead of sister chromatids that separate
 Independent assortment of maternal and paternal chromosomes (in meiosis I).
 Reductive division at the end of meiosis, each cell contains only half the number of chromosomes.

OOGENESIS: MEIOSIS OF FEMALE GAMETES

SPERMATOGENESIS: MEIOSIS OF MALE GAMETES

Heredity and Genetic Disease

DEFINITIONS
 GENES: The basic unit of heredity. Each chromosome carries genes that express proteins responsible for specific
traits.
 ALLELLE: Are genes that occupy the corresponding loci on homologous chromosomes and encode the same protein.
However, the level of expression or the protein itself may not be identical.
 HEREDITY: Heredity describes the transmission of genetic information from parent to offspring. A trait is passed on
by the independent assortment of chromosomes and crossing over in meiosis I.

MENDEL & HEREDITY

 Mendel discovered the basic principles of heredity by breeding garden peas in carefully planned experiments.
 Mendel studied several traits of pea plants, each of which had two alternative forms.
  MENDELS FIRST LAW: Law of Segregation – Alleles are different variations of the same characteristic. You have two
copies of every gene, an allele from each parent. Individuals carry two alleles that code for a specific trait. The alleles
segregate during gamete formation such that any individual gamete contains only one allele of each pair.
 MENDEL’s SECOND LAW: Law of Independent Assortment – Each pair of genes behaves independently of every other
pair of genes. This principle only works if the two genes are on different chromosomes.
 MENDEL’S THIRD LAW: Law of Dominance – In every pair of alleles, one is more likely to be expressed than the other.
This allele is dominant over the other allele.
 THE MENDELIAN MODEL:
1. Alternative versions of genes account for variations in inherited characters
2. For each character, an organism inherits two alleles, one from each parent
3. If the two alleles at a locus differ, then one (the dominant allele) determines the organism’s
appearance, and the other (the recessive allele) has no noticeable effect on appearance
4. The two alleles for a heritable character separate (segregate) during gamete formation and end up
in different gametes
 Mendel chose to track only those characters that occurred in two distinct alternative forms He also used
varieties that were true-breeding (plants that produce offspring of the same variety when they self-pollinate)
 TEST CROSS: A cross with the homozygous recessive organism is called a testcross.

SEX LINKED TRAITS

 The Y chromosome carries very little genetic information. Hence, genes on the X chromosome typically have no
corresponding allele on the Y chromosome.
 Males have only a single allele and are referred to as hemizygous for X-linked genes.
 Oocytes (egg cell) always pass on an X chromosome.
 Males only have one X chromosome that is always derived from the oocyte (always maternal).
 In inheritance males are hemizygous (A -, or a -) and females heterozygous or homozygous for genes on the X
chromosome (AA, Aa or aa).
 Red-green colour blindness is the most common X-linked trait in humans (8% Caucasian males). For sex-linked
inheritance the sex of the offspring matters. Males inherit their X chromosome only from their mother. Females
inherit X chromosomes from both parents. Mother is the carrier for colour blindness in males.

Genes
GENES
 Chromosomes composed of chromatin. A nucleosome is the basic unit of
chromatin and is composed of DNA wrapped around histones. Supercoiled
into visible chromosomes during mitosis.
 A gene is a hereditary unit that contributes to our traits or characteristics. It
is an organised sequence of DNA. DNA is a nucleic acid made of monomers
called nucleotides. DNA is organised into chromosomes. A gene is found in a
fixed position on a chromosome.
 The sum of all genetic material of a cell or organelle is a “genome”.
 The complete set of chromosomes in an organism is a “karyotype”.
NUCLEIC ACIDS
 Nucleic acids are polymers called polynucleotides
 Polynucleotides is made up of monomers: nucleotides. The three components of a nucleotide are the nitrogenous
base, sugar (pentose) and a phosphate group.
 A nucleoside is the same but doesn’t include the phosphate group.
 Nitrogenous bases can be pyrimidines (six membered ring) – Cytosine, Uracil and Thymine.
 They can be Purines (six membered ring fused to a five membered ring) – Adenine and Guanine.
 In DNA – Deoxyribose sugar, in RNA – Ribose sugar.

POLYNUCLEOTIDES
 Adjacent nucleotides are joined by covalent bonds: phosphodiester bonds. These bonds form between the –OH
group on the 3’ carbon of one nucleotide and the –OH on the phosphate on the 5’carbon of the next. It is a
condensation reaction releasing water as a byproduct.
 Phosphodiester bonds – these links create a backbone of
sugar-phosphate units with nitrogenous bases as
appendages. The sequence of bases along a DNA or mRNA
polymer is unique for each gene.
 DNA molecules have two polynucleotides spiraling around
an imaginary axis, forming a double helix.
 Nucleic acids have distinct ends
– the 3’ end has a free hydroxyl group on the 3’
carbon of a sugar
– the 5’ end has a free phosphate group at the 5’
carbon of the sugar
 In the DNA double helix, the two backbones run in opposite
5’→ 3’ directions from each other, an arrangement referred
to as antiparallel: the 5’ end of one strand is directly
opposite the 3’ end of the other strand.
 One DNA molecule includes many genes.

HYDROGEN BONDING
 Adenine (A) always with Thymine via 2 H-bonds.
 Guanine (G) always with Cytosine via 3 H-Bonds. }
Complementary base pairing.

 In RNA, thymine is replaced by Uracil (U) so A and U pair.

GENETIC CODE
 Via the transcription of mRNA from DNA. There
is a 3 letter nucleotide code for each amino acid
– Codon. This is the genetic code. There are 64
possible codon sequences, but only 20 amino
acids. Hence the genetic code is degenerate- as
more than one codon specifies one amino acid.
 The genetic code contains start and stop
codons.
 DNA Synthesis
DNA REPLICATION
 Occurs during the S (DNA Synthesis) phase of the cell cycle.
 ENZYMES IN REPLICATION: Topoisomerase – relaxes the
super coiling of DNA. Helicase – unwinds double strand of
DNA. Primase – synthesizes primers. DNA polymerase –
adds and proofread new bases. Ligase – links added bases.
 Nucleotides are added sequentially to a growing DNA
molecule by building blocks known as deoxyribonucleoside
triphosphates (dNTPs) – dATP, dCTP, dGTP and dTTP.
 Synthesis of DNA is always in a 5’ – 3’ direction, relative to
the strand being synthesized.
 DNA is replicated semi-conservatively: meaning that each
separate strand provides the template for building the new
strand by the base pairing rules.

REPLICATION IN PROKARYOTES AND EUKARYOTES

 Bacteria have a single circular chromosome. Replication begins at a single origin
of replication where unwinding begins. A replication fork is formed on each side
of the origin as small lengths of DNA separate for synthesis of new strands. Each
replication fork contains a leading strand growing toward the fork and a lagging
strand growing away from the fork. The two replication forks eventually meet at
the terminus.
 Eukaryotic cells contain much more DNA, so multiple origins of replication are
needed. Two replication forks are formed at each origin = bidirectional.
Synthesis proceeds 5’ to 3’ at each unit of replication (replicon), with leading and lagging strands. Replication forks
extend until they join with other replication forks or reach the end of the linear chromosome.
 A variety of enzymes and proteins are needed during the process of DNA replication to unwind the supercoiled
helices, separate the strands and keep them apart. New-strand synthesis is performed by DNA polymerases.
Synthesis always proceeds 5’- 3’ on the strand being produced, therefore:one strand is synthesised continuously; the
leading strand. The other (lagging strand) is synthesized by: DNA polymerase. It must work discontinuously in the
direction away from the replication fork and is synthesised as a series of small segments. These small segments of
DNA called Okazaki fragments are created. The Okazaki fragments are joined together by DNA ligase.
ERRORS IN REPLICATION: HUNTINGTON’S DISEASE
 DNA synthesis occurs at a rate of ~ 3000 nucleotides/min. On average, only 1 mistake occurs per billion bases.
 Replication errors will accumulate over time (this accounts for some of the differences between identical twins).
 HUNTINGTON’S DISEASE: Part of this gene has a repeated section called a trinucleotide repeat (3 nucleotides) – CAG,
which encodes for the amino acid glutamine (Q). This repeat region varies in length between individuals. When the
length reaches a certain threshold, it produces an altered form of the protein, called mutant Huntington protein
(mhtt). The resulting change in functions of these proteins leads to the disease symptoms, as mhtt increases the
decay rate of certain neurons.
 Polyglutamine stretch probably facilitates binding of a number of proteins. Extended polyglutamine tracts misfold
and form aggregates (inclusions) – possibly neurotoxic. Inclusions may also inhibit proteolytic degradation, inhibit
apoptosis and cause over production of reactive oxygen species.
 ANTICIPATION: HD can become more severe in successive generations. HD can also begin earlier and earlier in
successive generations. This phenomenon is called Anticipation. This commonly occurs in many triplet repeat
expansion diseases. The longer the length of trinucleotide repeats (within the disease range) the more severe and
earlier onset the disease.

DNA to Protein
TRANSCRIPTION
 Transcription is the production of messenger RNA (mRNA) from a
specific DNA sequence.
 It requires the enzyme RNA polymerase, nucleoside triphosphates
(ATP, GTP, CTP and UTP), and the DNA template.
 In Eukaryotic cells it occurs in the Nucleus.
 Only one strand of the double stranded DNA is ‘read’ during
transcription, this is called the template strand.
 Promoter is the initiation site, and the termination site is called a
terminator (in prokaryotes).
 Double stranded DNA unwinds, exposing the bases of the
template strand.
 RNA polymerase catalysis the incorporation of bases into a
growing RNA strand.
 The synthesis is in the 5’ to 3’ direction – which is the RNA.
 Same base pairs except Thymine is replaced with Uracil.

TRANSLATION
 mRNA moves from the nucleus to the cytoplasm in Eukaryotic cells. Translation requires: Ribosomes, mRNA, amino
acids, tRNA (adaptor molecules) that transfer amino acids to the growing polypeptide in a ribosome. It occurs in
three stages – initiation (UAG start codon), elongation and termination (UAA, UAG or UGA sop codons).
 RIBOSOMES: Ribosomes are large RNA/protein complexes. Present in the cytoplasm. Comprised of a large and a
small subunit. In the presence of mRNA, the two subunits join.
  Stages of Translation:
1. INITIATION: Once the mRNA/ribosome complex is formed,
the mRNA threads through the ribosome until a start
codon (AUG) is found. This initiates protein synthesis. AUG
codes for methionine (or fMet in prokaryotes), so Met is
the first amino acid in virtually all eukaryotic proteins.
2. ELONGATION: A second amino acid is then added to
methionine, by the formation of a covalent peptide bond.
The amino acid added is determined by the next 3 bases in
the mRNA. Many more amino acids are added, forming a
polypeptide chain.
3. TERMINATION: When the ribosome moves to a stop codon
on the mRNA (UAA, UAG or UGA), translation stalls, the
ribosome separates (into large and small subunits), and
protein synthesis is terminated.
 tRNA: tRNA molecules are short RNA molecules, that form the crucial
link between DNA and protein. The anticodon recognises the
sequence of bases in the mRNA molecule, while an amino acid is
attached at the 3’ end. As the mRNA threads through the ribosome,
tRNA molecules attach to the mRNA, along with their amino acids.
There are 2 sites within a ribosome, such that 2 tRNA molecules can
attach at the same time. The amino acids are brought into close
proximity. A peptide bond is formed between the two amino acids.

SUMMARY

Genetic Engineering
BACTERIAL PLASMIDS
 BACTERIAL PLASMIDS: Only present in
bacterial cells. Bacteria contain (usually)
1 main chromosome, 1 or more
plasmids. A plasmid is a double-
stranded, circular DNA molecule.
Capable of replication. Dispensable! Can
be taken up from the environment!
  How are Plasmids used in Genetic Engineering?
1. PLASMIDS ARE ISOLATED: Inoculate bacteria into liquid culture; grow overnight to produce large
amounts of the bacteria and hence the plasmid. Break open the bacteria to release the plasmid and
purify the plasmid from remainder of bacterium.
2. FOREIGN GENE IS INSERTED INTO PLASMID = ENGINEERED PLASMID: Plasmid + Double-stranded
linear DNA, containing foreign gene  Plasmid containing foreign gene. This process is called
ligation.
3. ENGINEERED PLASMID PLACED BACK INTO BACTERIUM: Some bacteria taken up engineered plasmid,
this process is called transformation.

HOW ARE FOREIGN GENES ISOLATED?

 In order to insert a foreign gene into a plasmid, we must first isolate it from the rest of the genome. This is done by
cutting the foreign gene from the genome. DNA is cut using restriction endonucleases. Restriction endonucleases
(restriction enzymes) are bacterial enzymes that cut DNA at specific sequences. Endonucleases cut DNA at internal
sites.
 Example: The restriction enzyme EcoRI recognizes the specific sequence GAATTC. If EcoRI is incubated with a
genome, it will cut that genome at every place where the sequence GAATTC exists. GAATTC is a genetic palindrome –
the sequence reads the same on both strands of DNA, in the 5’ to 3’ direction. This means that both strands will be
cut.
 What are Sticky Ends? In solution, they will tend to stick to each other, because of complementary base pairing. But
this is not a covalent (or permanent) attachment. Blunt ends are when the restriction enzyme cuts at the same place
on both strands; AluI is an example. Both sticky ends and blunt ends can anneal (H-bond) to each other. The pieces of
DNA are held together only by H-bonding between the complementary bases. The sugar-phosphate backbone
remains broken.
 Inserting the Foreign Gene: This is easily achieved by cutting the plasmid with the same restriction enzyme that was
used to isolate the foreign gene. The sticky ends on the plasmid will be compatible (form complementary base pairs)
with the sticky ends of the foreign gene. But requires an additional enzyme, DNA ligase, to covalently link the sugar-
phosphate backbone = ligation.
 How do we isolate the bacteria that contain a plasmid with the foreign gene? We have a reaction tube containing a
mixture of plasmid, bacteria that don’t contain plasmid and bacteria that do contain plasmid. Most plasmids used for
genetic engineering already have an antibiotic resistance gene. If the mixture of bacteria are placed onto an agar
plate containing antibiotic, only those bacteria that contain the plasmid can grow.
 Can Distinguish Two Type of Bacteria by their LacZ Activity: IF LacZ is functional – bacterial colonies are Blue. IF LacZ
has been disrupted (by the presence of the foreign gene) – bacterial colonies are white.

Viruses
VIRUS CHARACTERISTICS
 Are small infectious particles consisting of little more than genes packaged in a protein coat.
 Size: Typically less than 1 µm
 Viruses are obligate intracellular parasites – lack metabolic enzymes, DNA replication enzymes, ribosomes etc… and
require a host to survive!
 Viruses have either a DNA or RNA genome – single- or double-stranded (basis of animal virus classification)
 Surrounded by a protein coat and in some cases an envelope
 Can infect all types of organisms from animals and plants to bacteria and archaea
VIRUS STRUCTURE
 The nucleic acid (DNA or RNA) of a virus is surrounded by a protein coat – CAPSID
 Each capsid is composed of protein subunits – CAPSOMERES
 The capsid with its enclosed genome is referred to as – NUCLEOCAPSID
 In some viruses the capsid is covered by an – ENVELOPE – composed of lipids, proteins and carbohydrates
 A completely assembled and infectious virus – VIRION
 Can be helical, polyhedral, enveloped and complex, shapes.

VIRAL REPLICATION
 Viruses replicate within host cells and generally use the host cell’s DNA, RNA and protein synthesis machinery -
obligate parasites.
 Viruses are specialized at entering and replicating within a specific target host.
 Attachment: Viruses attach to cell membrane
 Penetration: by endocytosis or fusion
 Uncoating: by viral or host enzymes
 Biosynthesis: Production of nucleic acid and proteins
 Maturation: Nucleic acid and capsid proteins assemble
 Release: by budding (enveloped viruses) or rupture (lysis of host cell)

INFLUENZA VIRUS
 Influenza virus infects cells attaching via haemagglutinin (HA), replicates within the infected cell, and is then released
by the action of neuraminidase (NA). Influenza viruses infect host cells turning them into Influenza Virus “flu
factories”.
 Relenza is an influenza virus NA inhibitor. Relenza sits inside the neuraminidase “enzymatic pocket” blocking
enzymatic activity.
 Vaccines: Harmless preparations (usually proteins or polysaccharides) derived from a pathogen. When introduced
into humans or animals, an immune response is stimulated which prevents subsequent infection by the pathogen.
 Variation in the genes encoding HA and NA results in new strains of the virus which are not recognized by the human
immune system, which leads to new epidemics and pandemics = virulence determinants.

Bacteria
BACTERIA
 REPRODUCTION: Bacteria replicate by binary fission -
DNA replicates then bacteria split down the middle to
produce identical daughter cells. Endospores are
produced by some bacteria - defensive strategy against
hostile or unfavorable conditions (endosporulation).
 STRUCTURE: Can be spheres, rods or spirals.
  CELL WALL: The Bacterial Cell Wall is a semi rigid
structure present in all bacteria. It determines the
shape of the bacteria. It provides mechanical support
so that the cell does not burst when exposed to
conditions of osmotic pressure. Bacteria can be
classified via their cell wall as either gram negative and
gram positive.
 BACTERIAL CELL SURFACE:
- Capsule: A polysaccharide or protein layer. Capsules can allow cells to adhere to their substrate or to
other individuals in a colony. Some capsules protect against dehydration and some may increase
resistance to host defenses.
- Fimbriae: also called attachment pili, which allow them to stick to their substrate or other individuals
in a colony. Sex pili are longer than fimbriae and allow bacteria to exchange DNA from donor bacterial
cell to a recipient bacterial cell – conjugation.
- Flagella: The beating of flagella scattered over the entire surface or concentrated at one or both ends
is the most common method of movement.

BENEFICIAL BACTERIA
 Ecosystems: Cyanobacteria fixes Carbon and Nitrogen. Mutualistic Relationships with both plants and animals:
Rhizobia fixes nitrogen for legumes through an association with their roots.
 In Humans: Bacteria in the gut and on the skin is an important aspect of good human health. Human skin has
bacteria which prevents colonization of pathogenic bacteria. In our guts bacteria aids digestion, such as breaking
down foods that out intestines cannot, and provide vitamins that we cannot synthesize.
 Commercially: Aid in making cheese, plastics and drugs. Also are used to reduce pollution in a process called
bioremediation.

PATHOGENIC BACTERIA
 Virulence determinants: genetic or biochemical or structural features that enable a bacteria to produce disease in a
host. There are two types of Virulence determinants:
- Those that Promote Colonization of the Host:
 Fimbriae (also known as pili): allow them to stick to their substrate or other individuals in a
colony.
 Capsule (Slime Cover): A polysaccharide or protein layer. Capsules can allow cells to adhere to
their substrate or to other individuals in a colony. Some capsules protect against dehydration
and some may increase resistance to host defenses.
 Afimbrial adhesions: are surface proteins of pathogenic bacteria which promote attachment
to host structures, but are not organised into fimbrial structures.
- Those that Damage the Host: Bacteria produce toxins to disrupt the cells of the immune system,
disrupt epithelial cells and therefore gain access to the body, disrupt host cells to promote nutrient
release.
 Exotoxins: produced inside mostly gram positive bacteria as part of their growth and
metabolism. They are then secreted or released following lysis into the surrounding medium.
 Endotoxins: Part of the outer portion of the cell wall of gram negative bacteria. They are
liberated when the bacteria die and the cell wall breaks apart.
 Bacterial Disease Prevention: Disinfectants kill microorganisms found on the surface of skin, medical instruments
etc. Good general hygiene practices. Probiotics are live microbial ingredients in foods or supplements that compete
for resources with disease causing microorganisms and produce toxins that kill potential "invaders".
  Antibiotics: are natural organic substances of microbial origin which is either toxic or growth inhibitory to other
microorganisms, such as penicillin. However they do not distinguish between good and bad bacteria and so can
result in opportunistic disease and can also promote antibiotic resistance if used incorrectly.
 Mode of Action of Antimicrobials:
- Inhibition of pathogen’s attachment to, or recognition of, host.
- Inhibition of DNA and RNA synthesis
- Inhibition of general metabolic pathways.
- Inhibition of protein synthesis.
- Inhibition of cell wall synthesis.
- Disruption of cytoplasmic membrane.
 GROUP A STREPTOCOCCI:
- Also known as Streptococcus pyogenes and the "flesh eating bacterium“. Spherical gram-positive
bacterium that grows in long chains. Bacterial pathogen that commonly colonises the throat and skin
of the human host. Cause a wide variety of endemic human diseases.
- The most common disease caused by Streptococcus pyogenes is pharyngitis which is also known as
strep throat".
- Virulence Factors of group A Strep:
 M protein – major surface protein – antiphagocytic
 Streptokinase - enzymatically activates plasminogen, proteolytic enzyme, into plasmin which
in turn digests fibrin and other proteins
 Hyaluronidase - breaks down hyaluronic acid, an important component of the connective
tissue
 Streptodornase - protect the bacteria from being trapped and killed by neutrophils
- Treatment of streptococcal infections requires antibiotic treatment – Penicillin. Vaccines are currently
under development – a “one shot vaccine” that prevent S. pyogenes infection and DOES NOT cause
post streptococcal sequelae!

Protista
GENERAL CHARACTERISTICS
 Protista are a diverse, mostly cellular groups of eukaryotes residing primarily in aquatic habitats.
 Eukaryotic: Typical but plus some additional and unusual organelles – such as micronucleus, macronucleus,
contractile vacuole and pigment spot.
 Diversity: exhibit more structural and functional diversity than any other eukaryotic group.
 Mostly unicellular: may also be filamentous, colonial or multicellular.
 Mostly aquatic: occupy a vast array of habitats.
 Ecological Importance: Protists supply nearly half of the world’s oxygen, largely as a result of photosynthetic
unicellular algae. Photosynthetic protists are responsible for a large part of the primary production in oceans. Non-
photosynthetic protists, protozoa, play critical roles in transferring energy to higher levels in the food chain. Role in
decomposition and nutrient recycling. Chemicals made from some protists are used in treatment of ulcers, high
blood pressure, and arthritis. Euglena are used to catalyze treatment of raw sewage. Trichonympha live in the
digestive tracts of termites and enable them to digest wood.
 Parasitic: some protista cause human disease.
 Protist Nutrition:
- Photoautotrophs: contain chloroplasts.
- Heterotrophs: absorb organic molecules or ingest large food particles.
- Mixotrophs: combine photosynthetic and heterotrophic nutrition.
  Protist Reproduction: Most reproduce asexually only via Binary fission. Few reproduce sexually (meiosis and micro
nuclear fusion or fertilization) and some utilize a process called conjugation.
 Protist Locomotion: Cilia – hair like projections extending from the surface of the cell. Flagella – long thread like
appendage found singly or in pairs. Pseudopodia – temporary projection of the cytoplasm of certain cells.

PROTOZOAN PARASITES
 Parasitic protozoans based primarily on their mode of locomotion.
 Protozoa that enter the body via ingestion have two morphological forms:
– Trophozoite (all) - Feeding and reproducing stage that lives within the host.
– Cyst (some) - Infective form that survives in the environment. Parasite stage responsible for transmission.
 CILIATE: Balantidium coli (Balantidiasis) - Only ciliate known to cause disease in humans. Intestinal protozoan
parasite -commonly found in animal intestinal tracts. Transmitted via the faecal-oral route via cyst. Trophozoites
attach to mucosal epithelium lining the intestine.
 AMOEBAE: Acanthamoeba and Naegleria- Cause rare and usually fatal brain infections. Common inhabitants of
natural and artificial water systems.
 FLAGELLATES: Trypanosoma brucei – Causes African sleeping sickness (trypanosomiasis). The insect vector is the
tsetse fly found in vegetation along watercourses and lakes, forest edges, and areas of scrub savanna in Africa.
Humans usually infected when bitten by infected tsetse flies.
 APICOMPLEXANS: Infective form characterized by ornate complex of organelles at their apical end involved in
attachment to host. Parasites of animals. Life cycles involve at least two types of hosts. E.g. Toxoplasma gondii.
 Definitive host = is a host in which the parasite reaches maturity and, if possible, reproduces sexually.
 Intermediate host = is a host that harbors the parasite only for a short transition period, during which (usually) some
developmental stage is completed.

Fungi
CHARACTERISTICS OF FUNGI
 Eukaryotes: can have multiple nuclei. Some are single celled but most are filamentous and have complex
multicellular bodies. Cell walls are typically composed of chitin.
 Are heterotrophs: secrete enzymes and other substance and absorb the digested nutrients. Do not perform
photosynthesis.
 The morphology of multicellular fungi enhances their ability to absorb nutrient. They consist of mycelia – networks of
branched hyphae adapted for absorption. Some fungi have hyphae divided into cells by septa, with pores allowing
cell to cell movement of organelles. This is called Septate hypha. Coenocytic fungi lack septa.
 Specialised hyphae: some unique fungi have hyphae that allow them to feed on animals. Others have hyphae called
haustoria that allow them to penetrate the tissues of their host to extract or exchange nutrients.
 Fungal Nutrition: Digest nearly any form of organic carbon, and use glucose as a source of carbon. Fungi can be
Saprophytic – rhizoids anchor hyphae and absorb nutrients, are decomposes, releasing enzymes onto substrate and
absorb nutrients. Can be Parasitic with highly specialised hyphae with haustoria that absorbs nutrients from host
cells.
 Reproduction: Many reproduce both sexually or asexually. Others only sexually or asexually. Most propagate by
producing large numbers of spores.
BENEFICIAL FUNGI
 Decomposes organic material including cellulose and lignin of plant cell walls. Nutrient cycling. Return inorganic
nutrients to soil.
 Mycorrhizal fungi – provide nutrients to host plants allowing plants to grow in low nutrient soils. Absorbs and
transfers mineral nutrients such as phosphorous, zinc and copper to the plant. Protects plant against toxic levels of
some minerals.
 Endophytes – Living in plant tissues, can be parasitic or symbiotic. Make toxins that deter herbivores and defends
against pathogens.
 Fungus – Animal Mutualism: Some fungi share their digestive services with animals. These fungi help break down
plant material in the guts of cows and other grazing mammals. Many ants and termites use fungi to convert plant
material into substance that they can digest. They also can detoxify plants defensive compound that would other kill
or harm the ants.
 Food Industry – Brewer’s or Baker’s yeast. Ferments sugars to alcohol and carbon dioxide. Bread making. Beer
making (Saccharomyces cerevisiae).
 Healthcare – Some fungi used to produce antibiotics to treat bacterial infections.

PATHOGENIC FUNGI
 Some fungi that attack food crops are toxic to humans. Animals are much less susceptible to parasitic fungi than are
plants. The general term for a fungal infection in animals is mycosis
 Mycoses are among the most difficult diseases to diagnose and treat
– Signs of mycoses are often missed or misinterpreted
– Fungi are often resistant to antifungal agents
 Systemic mycoses: Deep within body. Histoplasmosis (Histoplasma capsulatum). Opportunistic Fungi Candidiasis
(Candida albicans).
 Subcutaneous mycoses: Beneath the skin.
 Cutaneous mycoses: Affect hair, skin, and nails
 Superficial mycoses: Localized, e.g., hair shafts
 Opportunistic mycoses: Caused by normal microbiota or environmental fungi.
 Fungal mycotoxins can cause toxicosis. Two types of toxicosis – Mycotoxicosisc caused by eating mycotoxins.
Mycetismus - Mushroom poisoning from eating a fungus.
 Mycotoxicoses: Mycotoxins produced by fungi during normal metabolic activities. Poisonous. Often consumed in
grains or vegetables. Aflatoxins are the most well-known mycotoxins. Fatal to many vertebrates. Carcinogenic at low
levels when consumed continually.
 Fungal Intoxications: Mycetismus - Mushroom Poisoning. Most mushrooms are not toxic Some produce extremely
dangerous poisons - Neurological dysfunction, Hallucinations, Organ damage or death. Poisoning typically occurs
when untrained individuals pick and eat wild mushrooms. Deadliest mushroom toxin produced by the “death cap”
mushroom.

Innate Immunity
INNATE IMMUNITY
 Innate immunity is present before any exposure to pathogens and is effective from the time of birth.
 It involves nonspecific responses to pathogens
 Innate immunity consists of external barriers plus internal cellular and chemical defences.
DEFENCES
 SKIN: The unbroken skin is a formidable barrier to microorganisms. Keratin of skin is resistant to weak acids and
bases and to bacterial toxins and enzymes. They help suppress the growth of more virulent organisms (opportunistic
infections). These normal or resident bacteria (commensals), may become dangerous if they get inside the body.
 MUCOSAE: Mucous membranes line all body cavities that open to the exterior - digestive, respiratory, urinary and
reproductive tracts. These membranes produce protective chemicals:
- Mucus which make surfaces moist and forms a sticky trap for dust, microorganisms and other
environmental pollutants that become caught.
- The cilia sweep the mucus, plus whatever is trapped in it, into the pharynx (where it is usually
swallowed). Particularly important in the respiratory tract, where the epithelial cells are also ciliated =
ciliary escalator. These surfaces must be intact to be effective.
- Mucosae may also produce secretions that inhibit microbial growth:
Acidic skin secretions (pH 3-5) from oil and sweat glands
The stomach secretes hydrochloric acid (pH 2)
Vaginal secretions are also acidic (pH 4)
Sebum from oil glands also contains chemicals that are toxic to bacteria.
Lysozyme in tears saliva and mucous secretions is an enzyme that destroys the cell wall of bacteria.
 CELLULAR INNATE DEFENCES: The second line of defense.
- Phagocytes: They are one of the first immune cells to confront foreign material within the body. The
major phagocytes in the body are called macrophages. They develop from WBCs called monocytes.
These cells ‘roam’ through the body looking for foreign material. When they encounter non-human
material they attempt to literally ‘eat’ it. Some tissues have permanent resident macrophages, eg,
Kupffer cells in the liver. Phagocytic cells recognise groups of pathogens by TLRs, Toll-like receptors
which recognize molecular patterns not found in animal cells. Eg. LPS, flagellin, dsRNA. Other types
of white blood cells are also phagocytic:
 Neutrophils: circulate in the blood are attracted by signals from infected tissue and then
engulf and destroy pathogens. Neutrophils also produce antibacterial chemicals called
defensins which are quite lethal to foreign cells. Neutrophils capture bacteria by secreting
neutrophil extracellular traps (“NETS”; composed of DNA and protein). Bacterial capture by
 Eosinophils: are weakly phagocytic but discharge destructive enzymes and are important in
defending the body against parasitic worms.
 Dendritic cells: stimulate development of adaptive immunity
 NATURAL KILLER CELLS: A second variety of non-specific attack cell. They are large cytotoxic granular lymphocytes.
They recognise and kill cells infected with viruses or cancerous cells. NK cells are not phagocytes but they release
chemicals – perforins and also granzymes that attack infected cells’ membranes, destroying the cells. NK cells also
release chemicals which amplify the inflammatory response.
 COMPLEMENT: Complement refers to a series of 30 plasma proteins normally present in the blood in an inactive
state. Activation of these proteins enhances the inflammatory response. Activation of the complement system
results in production of the membrane attack complex. The membrane attack complex proteins are inserted into
the target cell membrane. This puts a hole in the cell membrane, allowing water and ions to enter and brings about
cell lysis and death. Antimicrobial proteins! The consequences of complement activation are: Local inflammation,
Phagocytosis and Bacterial lysis.
 INTERFERONS – VIRAL INFECTIONS: Viruses are extremely small particles and they do not present any surface
recognition molecules. They cannot be phagocytized like bacteria. Viruses replicate by taking over the protein
synthesis machinery of body cells making lots more viruses to infect yet other cells. However, body cells once
infected, can produce small proteins (interferons). The interferons stimulate healthy cells to block the viruses’
ability to replicate. Helps healthy cells to defend themselves. They are non-specific, so they are active against a
variety of viruses. They also attract macrophages and natural killer cells.
  INFLAMMATION: The inflammatory response is triggered whenever tissues are injured by physical trauma, intense
heat, irritating chemicals, or infection.
- Benefits of inflammation: Prevent spread of damaging agents, disposal of cell debris and pathogens
and helps heal the damaged tissue.
- The inflammatory response, such as pain and swelling, is brought about by molecules released upon
injury or infection.
- Mast cells, a type of connective tissue, release histamine, which triggers blood vessels to dilate and
become more permeable.
- Activated macrophages and neutrophils release cytokines, signaling molecules that enhance the
immune response.
- Inflammation can be either local or systemic (throughout the body).
- Septic shock is a life-threatening condition caused by an overwhelming inflammatory response.
- Fever is a systemic inflammatory response triggered by pyrogens released by macrophages and by
toxins from pathogens. This inhibits growth of some bacteria, speeds up chemical reactions and
repair. However excess heat can denature enzymes.

Adaptive Immunity
THE SPECIFIC IMMUNE RESPONSE
 Adaptive immunity is the defence mechanisms that target specific foreign macromolecules (antigens) often
associated with pathogens. These mechanisms are not “in place” but require exposure to pathogen (or foreign
molecule). Initially takes several days.
 Involves two components: “antibody-mediated” immunity (humoral immunity) and “cell-mediated” immunity.
 Only occurs in vertebrates and involves lymphocytes (white blood cells).
 B-lymphocytes (B cells) that mature in bone marrow  antibody-mediated immunity.
 T-lymphocytes (T cells) that mature in the thymus  cell-mediated immunity.

ANTIGENS
 Antigens are specific molecules, or parts of molecules, that the body recognises as foreign.
 The small accessible part of an antigen that binds to an antigen receptor is called an epitope
 Examples of antigens are: Foreign proteins, nucleic acids, sugars, lipids, pollen grains, microorganisms (bacteria,
fungi, and virus particles), etc. Blood group antigens are polysaccharides.
 ANTIGEN RECEPTORS: Exposure to the antigen activates B and T cells with antigen receptors specific for parts of that
antigen. B cells and T cells have receptor proteins that can bind to foreign molecules. Each individual lymphocyte is
specialised to recognise a specific type of molecule.

ANTIBODIES
 Binding of a B cell antigen receptor to an antigen is an early step in B
cell activation. This gives rise to cells that secrete a soluble form of
the protein called an antibody or immunoglobulin (Ig).
 Secreted antibodies are similar to B cell receptors but lack
transmembrane regions that anchor receptors in the plasma
membrane.
  Antibodies have:
- 2 identical light chains and 2 identical heavy chains joined together by disulfide bonds.
- Both have a constant region and a variable region.
- The variable region binds to antigen.
- The constant region differs depending on the class of the antibody and ensures that each antibody
generates an appropriate immune response for a given antigen.
 Antibodies are very specific protein molecules that mediate the humoral immune response. Antibodies are produced
and secreted by B lymphocytes in response to binding of antigens. Only one sort of antibody is produced by a
lymphocyte. Antibodies form the gamma-globulins in blood plasma. Antibodies are specific to a particular antigen –
The diversity of antibody molecules comes from random genetic rearrangement.

ANTIGEN RECOGNITION BY B CELLS

 Bound antibody acts as receptor. Same specifity and antigen binding
capacity as secreted antibody. The variable region binds to antigen.
 First step in B cell activation leading to formation of cells that can
secrete antibody.

ANTIGEN RECOGNITION BY T CELLS

 Each T cell receptor (TCR) consists of two different polypeptide chains (called alpha and beta). The tips of the chain
form a variable (V) region; the rest is a constant (C) region
 T cell and B cell antigen receptors are functionally different. T cells bind to antigen fragments displayed or presented
on a host cell. These antigen fragments are bound to cell surface proteins called MHC molecules.
 MHC (major histocompatibility complex) molecules are host
proteins that display the antigen fragments on the cell surface.
 In infected cells, MHC molecules bind and transport antigen
fragments to the cell surface, a process called antigen
presentation.
 Antigen presenting cells (APCs) have both class I and class II
MHC, whereas most body cells have class I only. Therefore
MHC class II allow recognition of APCs. A T cell can then bind
both the antigen fragment and the MHC molecule. This
interaction is necessary for the T cell to participate in the
adaptive immune response.

ADAPTIVE IMMUNITY
 Four Major Characteristics of Adaptive Immunity:
- Immense diversity of lymphocyes and receptors
- Self-tolerant
- Proliferation of B and T cells results from activation to produce the great number of cells specific for
the antigen encountered
- Immunological memory
  HUMMORAL IMMUNE RESPONSE: antibodies help neutralize or eliminate toxins and pathogens in the blood and
lymph.

Antibody-mediated mechanism of Antigen/Pathogen Disposal: Neutralise virus by blocking ability to bind to host,
Opsonisation involves digestion by phagocytosis by macrophages and neutrophils. Complement proteins generate a
membrane attack complex that forms pores in the membrane of the foreign cell, ions and water rush in and the cell
swells and lyses.
 CELL MEDIATED IMMUNE RESPONSE: Specialised T cells destroy affected host cells.

ANTIBODY MEDIATED VS. CELL MEDIATED

 Antibody-mediated = production of antibodies by B lymphocyte plasma cells.
 Cell-mediated = production of activated T lymphocytes (cytotoxic T cells and helper T cells) which directly attack
unwanted cells:
- TC cells secrete chemicals (e.g. perforin) that destroy target cells.
- TH cells secrete chemicals (e.g. cytokines) that amplify the activity of other immune cells (including
non-specific immune system).
 B cells recognise extracellular antigens (bacteria, viruses, bacterial toxins etc).
 T cells recognise and destroy bad self-cells, (virus-infected cells, cancer cells, etc).
 Very specific: B cell has to recognise the antigen to produce antibodies that can specifically bind to that antigen, T
cell also has to recognise the antigen to produce TC and TH cells.
 Activation of T and B cells will also produce some memory T and B cells that can be activated quickly upon a
subsequent exposure to the pathogen.
 ACTIVE IMMUNITY: Natural immune response following an infection, or artificial immune response following
vaccination.
 PASSIVE IMMUNITY: mediated by the transfer of antibodies and is normally short lived. Pass through placenta to
fetus (IgG), pass to newborn in milk (IgA) and may be used as a therapy (e.g. Rabies).

Biotechnology
BIOTECHNOLOGY
 Biotechnology is a broad term generally used to describe the use of biology in industrial processes such as
agriculture, brewing and drug development.
 Traditional applications include animal breeding, brewing beer with yeast, and cheese making with bacteria.
 The term also refers to the production of genetically modified organisms (GMOs) or the manufacture of products
from GMOs.

GENE TECHNOLOGY
 Gene technology is the term given to a range of activities concerned with understanding the expression of genes,
taking advantage of natural genetic variation, modifying genes and transferring genes to new hosts.
 GM CROPS:
- In Australia, ingredients from GMOs are typically only found in highly processed foods and cannot be
purchased as fresh food.
- Products from GM soybean, canola, corn, potato, sugar beet and cotton crops have been approved for
use in food in Australia. These crops have been modified to be insect resistant, herbicide tolerant or
both.
- GM crops that have been modified for insect resistance or herbicide tolerance allow farmers to use
less herbicide and pesticide on their farms.
- Insect resistant Bollgard II cotton has reduced pesticide use by up to 80 per cent. This means fewer
chemicals in the environment, and less harm to friendly insects.
 Genetic technologies are also used to develop new vaccines and treatments for preventing and diagnosing livestock
diseases. Research which involves the genetic modification of animals to benefit animal and human health is also
being conducted.
 GENE THERAPY: Gene therapy is the insertion of a functional gene into patients who suffer from a particular genetic
disease because they have a mutation in that gene. There are two types of gene therapy that are possible:
- Germ line gene therapy is the insertion of recombinant genes into germ cells (ova or sperm).The
transgene would be heritable.
- Somatic gene therapy is the correction of genetic defects in somatic cells (i.e. non‐germ line cells).
The transgene is limited to the individual and is not heritable to the progeny.
 DUCHENNE MUSCULAR DYSTROPHY (DMD):
- Is a recessive Xlinked disorder, mostly affecting males.
- DMD is caused by a mutation in the dystrophin gene.
- Dystrophin is an important structural component within muscle tissue.
- Aden-associated viruses (AAV) are being trialed for use in gene therapy for DMD. AAV can insert the
gene at a specific site and is non-pathogenic.
- Potential problems with gene therapy: Short‐lived nature of gene therapy, immune response,
problems with viral vectors, limited to single gene disorders and potential tumour induction.