BIO152

Topic 1

Objectives

• Describe and discuss cell theory

• Identify and analyse limiting factors that determine the maximum and minimum size of cells

• Compare and contrast features that distinguish prokaryotes from eukaryotes

• Name contents, including major organelles found in eukaryotic cell, describe structure + function

• Distinguish plant cells from animal cells based on cellular features

Introduction to Cells + Cell Theory

Cells = basic units of life

Organisms = either single-celled or multicellular

Cell Theory - History

Robert Hooke

• (1665) Built a microscope: enlarged objects 30x

• Came up with the name ‘cells’ (Latin ‘cellula’ (meaning little room))

Antoni van Leeuwenhoek

• Produced small lenses: enlarged objects 300x

• Observed living cells: in pond water and material scraped from teeth)

Various scientists (1830s)

• Robert BrownL rounded structure (in plant cells) named nucleus (Latin for kernel)

• Matthias Schleiden: plant tissues composed of cells

• Theodor Schwann: animal tissues composed of cells

Cell Theory

Explains relationship between life and properties of cell

• All organisms consist of one/more cells

• The cell = basic unit of all organisms

• All cells = from pre-existing, living cells

Functions

• Duplicate DNA

• Use DNA and RNA to make proteins

• Handle energy transfer

• Regilate exchange of materials

Cells of the World

Cells exist in different shape and sizes

Ge thierarcnerally small

• Most bacteria and archaea: 1-5nm

• Most higher plants and animals: 10-100nm

Unit of Measurement

1m = 1,000,000nm

• One um is one million times smaller than one metre

Cell Size

Cells are limited in size - despite various shapes

Size restricted by:

• Diffusion rates of molecules

  • Molecules move faster in smaller cells

• Maintaining concentrations of substances in cell

  • Higher concentration → increase collision rate → more reactions happening

• Surface area/volume ratio

  • Higher SA/V ratio = more efficient exchange of materials

Calculating Total Surface Area

Formula for surface area: 6s²

• Assuming cell is cube-shaped

• s = length of the side

  • If s = 2nm → 6 x (2)² = 24nm²

• If there are 1000 cells, TSA = 24nm² x 1000 cells = 24,000nm²

Organisms and Cells

Organisms can remain unicellular (prokaryotes) or multicellular (eukaryotes)

• Bacteria and achaea: unicellular - cells small and efficient

• Plants and animals: large, multicellular and complex in general - cells remain small and efficient.

Prokaryotes vs Eukaryotes

*Nucleoid ≠ nucleus

Bacteria, Archaea (Prokaryotes)

Bacteria

Genetic material folded and packed tightly in nucleoid.

• Escherichia coli (E.coli) has genetic material (1,300nm) inside its cell (1-2nm)

• Divided by binary fission

Archaea

Many species live in extreme environments, e.g. acidic hot springs (pH 2, temperature >100 deg C

Divided by binary fission

Animal Cells, Plant Cells (Eukaryotes)

Animal Cells

• Membrane-bound organelles require membrane fusion events to exchange materials.

• Divided by mitosis/meiosis + cytokinesis

Plant Cells

• Have walls containing cellulose and large vacuoles

• Divided by mitosis/meiosis + cytokinesis

Cell Ultrastructure

Cell (Plasma) Membrane

Cytosol + Cytoplasm

Cytosol

• Semi-fluid

• Site of chemical reactions, e.g. glycolysis

Cytoplasm

• Majority of internal volume of cell

• Comprises cytosol, organelles, cytoskeletal fibres

Ribosomes

• Not membrane-bound

• Made of RNAs and proteins

• Consists of two subunits

  • Large SU

  • Small SU

• Free (individual) or grouped (as part of rough ER)

• Function = protein synthesis

Animal Cell vs Plant Cell (Eukaryote)

Topic 2

Objectives

• Know basic structure of amino acid + classify different amino acids according to R groups

• Describe linkage of amino acids with different bonds + classification of proteins according to structure (primary, secondary, tertiary, quaternary structures)

• Describe structure of cell membrane and major components

• Describe types of lipids, carbohydrates, proteins in membranes

• Analyse how membrane composition affects cell function with regards to cell and signal recognition, energy generation and transport processes

Proteins 1

Hierarchy of Structures

Biological molecules/structures classified into various levels:

1. Level 1 - soluble organic molecules

2. Level 2 - biological macromolecules

3. Level 3 - Supramolecular structures

4. Level 4 - organelles

5. Level 5 - cell

Amino acids

• Building blocks of protein

• Made of

  • α carbon

  • Hydrogen atom

  • Carboxyl group (carboxylic acid): (-COO^-)

  • Amino group (Amine): -NH3^+

  • R group

• Exist in two stereoisomeric forms:

  • Mirror-image forms called L- and D-amino acids

  • Only L-aa found in proteins

• 20 Different kinds

• Properties depend on R groups = classification

R Groups

Type of R group determines different characteristics (or combination) for amino acid

• Nonpolar (hydrophobic): R group mostly contain hydrocarbon group e.g., CH3. May also have N or S atoms but effect of these atoms are diminished due to hydrocarbon portion.

• Polar (hydrophillic): R group contain either hydroxyl group (OH), sulfhydryl group (SH) or amide group (CONH2).

• Neutral R group has no charge, may contain highliy electronegative atom, e.g. O, N or S.

• Negatively charged: R groups contain negatively charged group (e.g. -COO^-)

• Positively charged: R groups contain positively charged group (e.g. -NH3^+)

Proteins

• Important macromolecules in all organisms/cells

• Greek: proteios = first place

• Made of amino acids linked together by peptide bonds

  • Each protein has unique amino acid sequence

  • 2 a.a. = dimer, 3 a.a. = trimer, 4 a.a. = tetramer

  • Structure important for functionality

• Amino group = start (N-terminus)

• Carboxyl group = end (C-terminus)

Peptide Bonds

• Linkage of two amino acids via covalent bonding

• Condensation reaction

  • Water removed

  • O from carboxyl group

  • 2 H from amino group

Hydrogen Bonds

• Donors (e.g. Amine/Hydroxyl group of a.a.) have H atom linked to more electronegative atom of acceptor (e.g. Carbonyl, sulfhydrall groups of a.a.)

• Weak bond

• Abundant in proteins

Disulfide Bonds

• Form between sulfur atoms of two cystein amino acid residue via covalent bonding

  • Via oxidation: removal of H atoms

    

  • Disruption of disulfide bonds denatures proteins

    • Via reduction: addition of 2 H atoms to regen sulfide groups (SH)

Ionic Bonds

• Opposites attract

• R groups can be + or - charged

• Electrostatic interactions between R groups

  • + bind to - (attract)

  • Similar charge (repel)

• Too high/low pH value affects ionic bonds

  • Results in proteins denaturing

Van Der Waals Interactions

• Transient attraction of two nonpolar molecules

• Typically weak, energy of 0.1-0.2 kcal/mol

  • Only effective if molecules are close

Hydrophobic Interactions

• Tendency of hydrophobic molecules excluded from water interaction

Primary Structure

Linear sequence of amino acid in polypeptide

• Amino acid linked by peptide bonds

Sequence determines higher orders of structures

Secondary Structure

Backbone of polypeptide

• Linear sequence fold upon itself

Structure stabilised by H bonds

• Intermolecular (between CO and NH groups of amino acids of two different polypeptides)

• Intramolecular (between CO and NH groups of amino acids of same polypeptide)

Structural patterns:

• a-helix

• b-sheet

Alpha Helix

• Spiral in shape; coils around itself

• Stabilised by H bonds

  • CO group bind to NH group of peptide bond, 4 a.a. downstream

• Spiral turn has 3.6 a.a.

  • 4th a.a. close to 3rd and 5th a.a.

    • Makes structure stretchable

• Some a.a. disrupt alpha helix (helix breaker)

  • Proline (lacks H needed for H bonding)

    

Beta sheet

• Sheet-like conformation

• Forms when two parts of polypeptide chain lie side by side

• Stabilised by hydrogen bonds

Motifs

• Combinations of alpha helices and beta sheets

  • Connected via looped regions of varying length (random coils)

  • E.g. β-α-β, hairpin loop and helixturn-helix motifs

Tertiary Structure

Higher order of folding

• Polypeptide chain folded back on itself → condensed structure

• Depends on interaction between side chains

Stabilised by

• Disulfide bonds, H bonds, ionic bonds, van der Waals interaction and hydrophobic interaction

Two main classes

• Fibrous proteins vs Globular proteins

Proteins 2

Objectives

• Describe classification of proteins according to function

• Describe how pH affects charge of proteins

• Describe how proteins can be separated based on given examples

• Describe denaturation effects on protein structure

Proteins - Summary + Categories

• Proteins made up of amino acids: Carboxyl, amino and R groups

  • Properties dependent on the R group

  • Four levels: Primary, secondary, tertiary and quaternary structure

• Properties of a protein determined by

  • Types of amino acid

  • Number of amino acids

  • Amino acid sequence

• Protein properties are important for its role and purpose

Can be categorised into:

Category	Function	Example
Enzymatic	Selective acceleration of chemical reactions	Digestive enzymes catalyse hydrolysis of bonds in food molecules
Defensive	Protection against disease	Antibodies inactivate and help destroy viruses and bacteria
Storage	Storage of amino acids	Casein, protein of milk, is major source of amino acids for baby mammals. Plants have storage proteins in seeds. Ovalbumin is protein of egg white, used as amino acid source for developing embryo.
Transport	Transport substances	Haemoglobin, iron-containing protein of vertebrate blood, transports oxygen from lungs to other parts of body. Other proteins transport molecules across membranes.
Hormonal	Coordination of an organism’s activities	Insulin, a hormone secreted by pancreas, causes other tissues to take up glucose, thus regulating blood sugar concentration.
Receptor	Response of cell to chemical stimuli	Receptors built into membrane of nerve cell detect signalling molecules released by other nerve cells.
Contractile and motor	Movement	Motor proteins are responsible for undulations of cilia and flagella. Actin and myosin proteins are responsible for contraction of muscles.
Structural	Support	Keratin is protein of hair, horns, feathers, etc. Insects and spiders use silk fibres to make cocoons and webs. Collagen and elastin proteins provide fibrous framework in animal connective tissues.

pH on Proteins

pH changes protonation state of charged residues

Acid	Neutral	Alkaline/Basic
Low pH	Uncharged	High pH
Increase H+		Decrease H+
Nett +ve charge	Isoelectric or pl (~has net electric charge of 0)	Nett -ve charge

Isoelectric Point (pl)

pH at which proteins are at 0 nett charge

• Positive and negative charges balanced

  • Also dependent on R groups

• Affects solubility

  • Proteins interact with each other rather than water

  • Might precipitate (fall out of solution = insoluble)

At pH away from pl, protein carries a charge (+)

• Because water is polar → forms a hydration shell around charged regions of protein.

• This water-protein interaction increases protein’s solubiliity in solution.

At pl point, there is no overall nett charge in protein

• Hence, less attraction between protein and water, and proteins more likely to interact with each other = protein precipitating out of solution.

Separation of Proteins

Properties depend on R groups of amino acids

• Nett charge of proteins

• Size

• Solubility - pl

Methods used in cell biology:

Charge: Electrophoresis

Separates mixture of proteins in gel matrix with voltage

Cathode (-) and anode (+) ends of gell

• Opposite charges attract

Protein mix will separate based on

• Charge

• Rate proportional to size of charge

Chromatography

• Separation of molecules suspended in a phase

  • Due to molecules having different properties (e.g. charges, affinity to certain molecules, polarity)

Charge: Ion Exchange Chromatography

• Beads trap/slows dwn flowing proteins of opposite charge

• Proteins of same charge flow through (elute) first

• Collected in fracions to purify proteins

• To get the proteins retained in column, change buffer (different pH and salt concentration)

Size: Gel filtration

• Beads slow down small proteins

  • Migrate slower (“Included”) in resin and elute last

• Larger proteins travel faster (“excluded”) through resin and elute first

• Collected in fractions to purify proteins

Solubility: Isoelectric point precipitation

• Separated based on solubility difference in water

  • Charge of protein attracts water forming hudrating shell and solubilises

• At pH = protein pl, no nett charge

  • proteins interact with each other rather than water

  • Might precipitate

Solubility: Ammonium Sulphate Precipitation

(NH4)2SO4 stripcs away hydration shell

• Decreases solubility allowing proteins to precipitate

Different proteins precipitate at different concentrations of (NH4)2SO4

Protein Denaturation

• Structures must be maintained for biological function

• Agents may cause loss of structures (2 degree, 3 degree and/or 4 degree) by disrupting bonds

• Loss off structure + function = denaturation

Factors causing denaturation

• Organic solvents - disrupt H bonds (2,3,4)

• Detergents - disrupt hydrophobic bonds (3, 4)

• pH extremes - disrupt H bonds (2,3,4) and ionic (electrostatic) bonds (3,4)

• Heat - increase kinetic energy and disrupt all non-covalent bonds (2,3,4)

Enzymes

Objectives

• Distinguish between reaction types of six major classes of enzymes

• Identify properties of enzymes

• Explain how enzymes function and evaluate the influence of various factors on reaction rates

Enzyme Nomenclature (Naming) + Classification

• Suffix “ase”

• Naming based on what it reacts with and how

  • E.g. pyruvate decarboxylase

    • Remove carboxyl group from pyruvate

    • Forming acetaldehyde and CO2

Enzyme Properties

Catalyst: speeds up reactions

• Equilibrium is attained more rapidly with enzymes

Stable: enzymes retain active structural conformation during reactions

Specific: only catalyse a single reaction (or a group of closely related reactions)

• Specific choice of substrates

• Tight regulation in cell (prevent chaos 

Equilibrium Constant (Keq)

• Measure directionality of reversible reaction (A ←→ B)

• Reaction will strive towards Keq

• Same regardless whether enzyme is present

• E.g.

  • Glucose-6-phosphate (G6P) ←→ fructose-6-phosphate (F6P)

Activation Energy (Ea)

• Activation energy - reactants must gain enough energy to collide effectively

• Transition state - high energy, unstable arrangeent reactants must reach

• Higher free energy - transition state has more free energy than starting reactants

• Catalysts - lower activation energy so transition state is easier to reach

• Temperature increases - gives particles more magnetic energy to overcome energy barrier

Enzymes as Catalysts

• Lowers activation energy requirement

• Forms transient, reversible complexes with substrate, facilitates interaction and stabilises intermediate transition state

  • Substrate binding occurs at active site of enzyme

  • Induced fit model: conformation change in active site as substrate binds

• Changes the rate at which Keq is achieved

Catalytic enzymes are dependent on:

• Enzyme concentration [E]

• Substrate concentration [S]

• Temperature

• pH

Enzyme concentration [E]

• As [E] increases, reaction velocity increases

  • Positive linear relationship

  • When [S] is in excess

Substrate concentration [S']

• As [S] increases, reaction velocity increases until a certain point

  • Low [S]: reaction velocity increases linearly with [S]

  • Km: [S] where reaction velocity is half of max velocity

  • High [S]: reaction velocity increases independent of [S] until Vmax

  • Vmax: Maximum velocity (rate of reaction)

    • Where all available enzymes are working at maximum

Importance of Km, Vmax

• Used to calculate rate at which substrate → product

  • Vmax: measure of potential maximum rate of reaction

  • Km (Michaelis constant): [S] where reaction velocity is half of max velocity

    • Tells binding affinity between anzyme and substrate

      • Km has inverse relationship with binding affinity of an enzyme for its substrate

        • High Km = low affinity

        • Low Km = high affinity

*Km and Vmax help calculate reaction rate at different substrate concentrations

*High Km needs more substrate to reach half-max speed → weak binding

*Low Km means enzyme reaches half-max speed at low substrate → strong binding

Temperature

• Low temperature = enzymes inactive

• Temperature up, reaction velocity up

  • Kinetic energy increases collision rate

• Extreme temperature denatures proteins

  • Non-covalent bonds in proteins affected

  • Diminished enzyme activity

pH

Most enzymes active within pH range of ~3-4 units

Extreme pH denatures enzymes

• Amino and carboxyl groups affected leading to disruption in electrostatic bonds

• Hydrogen and ionic bonds affected

• Diminished enzyme activity

Enzyme Regulation

• Substrate-level regulation

  • Regulation that affects substrate/products and enzyme interactions - reaction rate changes depending on how much substrate/product present

    • More substrate → enzyme works faster

    • More product → reaction may slow down

  • Not efficient - regulation type is basic and often not enough to control enzyme activity in cells

• Feedback inhibition

• Allosteric regulation

  • Allosteric inhibition

  • Allosteric activation

• Covalent modification

  • Addition or removal of chemical group

    • E.g. phosphorylation/dephosphorylation

  • Proteolytic cleavage

    • E.g. Enterokinase removes a hexapeptide segment from trypsinogen

Membranes and Transport

Membrane Recap

• Defines boundaries of cell and any internal compartments

• Site for biochemical functions

• Transport solutes in/ut of cell (selective permeability)

• Detect and transmit electrical/chemical signals

• Provide mechanisms for cell-to-cell contact, adhesion, communication

Membrane Structure

Fluid mosaic model

• Proposed in 1970s

• Fluid: lipids, proteins can move

  • Side-ways (lateral)

  • Around (rotation)

  • Flip-flip (transverse)

• Mosaic: presence of different kinds of proteins (mainly) and lipids such as cholesterol

Components:

• Phospholipid bilayer

  • Hydrophilic heads

  • Hydrophobic tails

  • Proteins - lipid ratios differ among membranes (protein:lipid)

    • E.g. outer mitochondria (~1.2:1), inner mitochondria (~3.5:1)

  • Carbohydrates

*Membranes that do lots of work, have more proteins

*Membranes that are more structural have more lipids

Membrane Lipids

1. Phospholipids

2. Glycolipids

3. Sterols

1. Phospholipids

• Most abundant

• Consists of polar head (attached to phosphate), backbone, non-polar fatty acid tails

• E.g. Phosphoglycerides, phospholingolipids

Phospholipid structure

• Polar head

  • Phosphate group

  • Variable R group (e.g. serine, ethanolamine, inositol)

  • Charged, therefore hydrophillic (polar)

• Backbone

  • Glycerol/sphingosine

• Non-polar tails

  • Hydrophobic

  • Fatty acids

    • Long unbranched hydrocarbon chain with carbonyl group

    • Saturated or unsaturated

Fatty Acids

2. Glycolipids

• Lipid with carbohydrate group

• E.g. glycoglycerolipids/glycospingolipids

3. Sterols

• Have four-ringed hydrocarbon

• E.g. Cholesterol, phytosterols (plant)

Membrane Carbohydrates

Oligosaccharides that can be attached to proteins (glycpproteins) or lipids (glycolipids)

• Help signalling

• Act as molecular markers (antigen) for recognition of substances

  • E.g. glycosphingolipid: A antigen and B antigen

    • Cell surface markers for different group of red blood cells

Membrane Proteins

Mosaic part of model

Integral proteins

• Hydrophobic regions embedded within lipid bilayer

• Classified by mode of attachment

  • Integral monotopic proteins

  • Transmembrane protein that acts as proton pump/channel

Peripheral proteins

• Extrinsic - lacks hydrophobic sequences

• E.g. Enzymes

Lipid-anchored

• Hydrophilic proteins - on membrane surfaces

• E.g. glycosylphosphatidylinositol (GPI) - anchored membrane proteins

Membrane Protein Function

• Transport

  • Ion homeostasis

• Enzymatic

  • Localises metabolic pathways such as ETC and ATP production

• Singal transduction

  • Binds to signalling molecule and relays messages

• Cell-to-cell recognition

  • Functions as markers for cellular identification

• Cell junctions

  • Glue cells together for barrier

• Structure attachment

  • Attachment to cytoskeleton

Membrane Permeability

• Selective permeability

  • Hydrophobic barrier to most molecules and ions

  • Permeable to small molecules (O2, Co2, but H2O is slower)

  • Hardly permeable to larger molcules (>100 MW molecules, glucose, - MW 180)

  • Impermeable to ions (H+, Na+, K+, Cl-)

• Most substances needed by cell are hydrophilic, may require transport protein to enter cell

Transport Across Membranes

• Simple diffusion (passive)

• Osmosis (passive)

• Facilitated diffusion (passive)

• Active transport (active)

Simple diffusion

• High → Low

  • Bidirectional movement depends on concentration gradient

• Does not require energy

• Does no require membrane protein

• Slow; may not meet cellular needs

• E.g. Gases (02, CO2)

Osmosis

• Diffusion of water across selective permeable membrane

  • Dependent on relative osmolarity (total solute concentration)

• Different species have different osmolarities

Types of solutions:

• Hypertonic

  • Higher solute concentration outside cell

• Isotonic

  • Same solute concentration to cell

• Hypotonic

  • Higher solute concentration inside cell

Facilitated

• Requires membrane proteins

  • Channel protein or carrier protein

  • Selective permeability (controlled passage)

• High → low

• Faster than simple

• Does not require energy

• Temperature-sensitive

• Can be saturated at high solute

• E.g. transport of glucose via carrier protein

Active

• Requires membrane carrier protein (pump)

• Requires energy (ATP)

• Molecules able to move against concentration

  • Uptake scarce essential nutrients

  • Remove waste and secretory products

  • Maintain [ions]

• E.g. Na+/K pump (Na+ out and K+ in)

Transport Processes

• Transport may differ in number of solutes transported and direction of movement

  • Uniport

  • Symport

  • Antiport

Topic 3

Objectives

• Define and discuss examples of autotrophs and heterotrophs

• Consider Solar and chemical sources of energy

• Define Metabolism

• Compare catabolism and anabolism

• Introduce role of ATP as an energy currency

• Compare Anaerobic vs aerobic respiration

• Understand ATP’s role as an “energy currency”

• Describe catabolic and anabolic metabolism

• General overview of anaerobic vs aerobic catabolism

• Introduce intermediate electron acceptors

• Define and describe the steps of glycolysis

• Understand the fate of pyruvate

Introduction to Energy Metabolism

Energy

Cell growth requirements:

1. Energy

2. Chemical building blocks

Building blocks = C O N P S

Building blocks are used to make macromolecules

Macromolecules:

Carbon sources

Organisms classified by how they obtain carbon

1. Autotrophs - C from atmospheric CO2

2. Heterotrophs - C from other organisms

Autrotrophs

• ‘Self feeding’

• Obtain energy from CO2

• Energy dependent process

• How autotrophs obtain energy classifies them as either photosynthetic or chemosynthetic

Photosynthetic organisms

• Use solar energy

• 
  

  

• Green plants

• Photosynthetic bacteria and algae

Chemosynthetic organisms

• Use chemical energy

  • From oxidisation of inorganic molecules

    • E.g. nitrifying bacteria

      

Heterotrophs

• ‘Feeding off others’

• Carbon is derived from assimilated compounds

  • E.g. glucose, a 5-carbon sugar

• Energy also obtained from these compounds

  • 
    

    

• Animals

• Most bacteria

• Non-green plant cells

Carbon sources

• Not all cells in an organism are the same

  • E.g in plants:

    • Roots = heterotrophic

    • Leaves - autotrophic

• Cells can be flexible

  • E.g. leaves:

    • During day, sunlight = auto

    • At night, no sunlight = hetero

Cellular metabolism

Metabolism = all of enzyme-catalysed reactions in a cell

Two phases:

1. Catabolism

2. Anabolism

Catabolism

• Degradative reactions of metabolism

• Larger compounds are broken down into smaller ones generating energy

• E.g.

  

Anabolism

• Synthetic (building) reactions of metabolism

• Smaller compounds combined into larger ones

• Requires energy

• E.g.

  

Cellular processes require energy for:

1. Anabolism - generation of more complex biomolecules

2. Transport - active transport of molecules within/between cells

3. Mechanical work - e.g. muscle contraction

All these processes require an input of energy such as ATP

ATP

• Energy carrier, transmitter, currency

• NOT energy store

• Nucleoside triphosphate

Nucleoside triphosphates

• Only differ in their nitrogen bases

  • 
    

    

• Used in

  • Polysaccharide synthesis (UTP)

  • Protein synthesis (GTP)

  • Lipid synthesis (CTP)

  • RNA synthesis (CTP, GTP, UTP)

Ribonucleoside triphosphates

Deoxyribonucleoside triphosphate

• If change sugar from ribose to deozyribose

  • 
    

    

• Used in DNA synthesis

  • dATP

  • dCTP

  • dGTP

  • dTTP

DNA = deoxyribonucleic acid

Ribo vs deoxyribonucleoside triphosphates

ATP hydrolysis

• ATP energy released via hydrolysis

• 
  

  

ATP generation

• ATP generated via catabolism in cell

• Two processes:

  • Fermentation

  • Aerobic respiration

1. Fermentation

• Under anaerobic conditions ( no O2)

• 
  

  

  
  

• Lactic acid fermentation

  • E.g. in muscle, in absence of O2

• 
  

  

  
  

• Ethanol fermentation

  • E.g. in yeast, in absence of O2

• 
  

  

2. Aerobic respiration

• Fuel molecules (e.g. glucose) completely oxidised to CO2

• ATP produced by both substrate-level phosphorylation and oxidative phosphorylation

• Hydrogen ions (H+) and their associated electrons (e-) are transferred from fuel molecules to O2

• O2 is reduced to H2O while fuel molecules are oxidised to CO2

  • This is an oxidation-reduction reaction

• Under aerobic conditions (O2)

  • 
    

    

    
    

    Summary table:

Glycolysis

Cellular Energy

Cells require energy (ATP) to grow

ATP - Adenosine triphosphate

Cells generate ATP via catabolic metabolism

ATP Generation

Catabolism

• Fermentation

  • Lactic acid fermentation

  • Ethanol fermentation

• Aerobic respiration

NAD^+ = nicotinamide adenine dinucleotide

FAD = flavin adenine dinucleotide

Lactic Acid Fermentation (Anaerobic)

(slides + Khan Academy)

E.g. in muscle (O2 absent)

• NADH transfers electrons directly to pyruvate → pyruvate as byproduct

• Lactate = deprotonated form of lactic acid (deprotonated = removal of H ion or proton from molecule by base in acid-base reaction)

• Lactic acid produced in muscle cells transported via bloodstream to liver then converted back to pyruvate and processed in remaining reactions of cellular respiration

• 4 ATP is produced in lactic acid fermentation but 2 ATP is consumed in initial priming steps, leaving 2 ATP

Ethanol Fermentation (Anaerobic)

(slides + Khan Academy)

E.g. in yeast (O2 absent)

• NADH donates electrons to a derivative of pyruvate, produces ethanol

• Pyruvate to ethanol:

  • Carboxyl group removed from pyruvate and released as CO2, producing two-carbon molecule (acetaldehyde)

  • NADH Passes electrons to acetaldehyde, regenerating NAD^+ and forming ethanol

Aerobic Respiration I

1. Glycolysis

• Glucose (6C) converted to 2 pyruvate (2× 3C). Produces ATP and converted NAD^+ to NADH

2. Pyruvate oxidation

• Each pyruvate goes into mitochondrial matrix, converted to two-carbon molecule bound to coenzyme A (AKA acetyl CoA)

• CO2 released

• NADH generated

3. Citric acid cycle

• Acetyl coA from last step, combines with 4-carbon molecule

• ATP, NADH, FADH2 produced

• CO2 released

4. Oxidative phosphorylation

• NADH and FADH2 from other steps deposit electrons in electron transport chain, turning back into NAD+ and FAD.

• As electrons move down chain, energy released and used to pump protons out of matrix (forms gradient)

• Protons flow back into matrix through ATP synthase (enzyme), produces ATP

• At end of ETC, O2 accepts electrons and takes up protons to form water

Glycolysis Steps

1. Glycolysis phosphorylated to glucose-6-phosphate

   • Consumption of 1 ATP

   • ATP → ADP

   • One-way reaction catalysed by hexokinase

   • Once phosphorylated, glucose → polar + “trapped” in cell with phospholipid membrane

2. Glucose-6-phosphate isomerised to fructose-6-phosphate

   • Substrate and product have same molecular weight

   • Reversible reaction catalysed by phosphoglucoisomerase (an isomerase)

3. Fructose-6-phosphate phosphorylated to fructose-1,6-bisphosphate

   • Consumption of 1 ATP

   • One-way reaction catalysed by phosphofructokinase

   • ATP → ADP

4. Fructose-1,6-biphosphate cleaved to produce 2× 3-carbon molecules

   • dihydroxyacetone phosphate (DHAP) and glyceraldehyde-3-phosphate (G3P)

   • Reversible reaction catalysed by aldolase

5. Only G3P can proceed through glycolytic pathway

   • DHAP isomerised to G3P

     • DHAP → G3P

   • Reversible reacction catalysed by triose phosphate isomerase

   • All reactions here will occur twice for the one glucose — started with glycolysis 6C glucose, continue with 2× 3C metabolites

6. Glyceraldehyde-3-phosphate oxidsed to 1,3-biphosphoglycerate

   • Requires 1x NAD^+ and 1x P

   • Coupled to reducion of NAD+ to NADH

   • Reversible reaction catalysed by G3PDH

   • Note: Pi (inorganic phosphate) and NAD^+ substrates

7. 1,2-biphosphoglycerate dephosphorylated to 3-phosphoglycerate

   • One phosphate group transferred to ADP, forming first ATP

   • Substrate level phosphorylation

   • Reversible reaction catalysed by phosphoglycerate kinase

8. 3-phosphoglycerate converted to 2-phosphoglycerate

   • Remaining phosphate group transferred to different carbon

   • Reversible reaction catalysed by phosphoglyceromutase

9. 2-phosphoglycerate converted to phosphoenolpyruvate (PEP)

   • Reversible reaction catalysed by enolase (a DH)

10. Phosphoenolpyruvate is dephosphorylated to pyruvate

    • Phosphate group transferred to ADP, forms ATP (second generation of ATP)

[STEPS 6-10 OCCUR TWICE FOR EACH GLUCOSE]

AFTER glycolysis:

• Aerobic respiration @ mitochondrion

• Ethanol fermentation @ cytoplasm

• Lactic acid fermentation @ cytoplasm

Glycolysis Steps (REWRITTEN)

1. Hexokinase
   • Glucose becomes glucose-6-phosphate.
   • A phosphate is added.
   • Glucose is trapped inside the cell.
   • Uses 1 ATP.
   • Irreversible.

2. Phosphoglucoisomerase
   • Glucose-6-phosphate becomes fructose-6-phosphate.
   • Molecule is rearranged to prepare for splitting.
   • Reversible.

3. Phosphofructokinase (PFK)
   • Fructose-6-phosphate becomes fructose-1,6-bisphosphate.
   • Second phosphate added.
   • Commits molecule to glycolysis.
   • Uses 1 ATP.
   • Irreversible.

4. Aldolase
   • Fructose-1,6-bisphosphate splits into DHAP and G3P.
   • Produces two 3-carbon molecules.
   • Reversible.

5. Triose Phosphate Isomerase
   • DHAP is converted to G3P.
   • Only G3P continues through glycolysis.
   • From here, all steps occur twice per glucose.
   • Reversible.

6. G3P Dehydrogenase
   • G3P becomes 1,3-bisphosphoglycerate.
   • G3P is oxidised.
   • NAD+ becomes NADH.
   • Pi is added.
   • Reversible.

7. Phosphoglycerate Kinase
   • 1,3-bisphosphoglycerate becomes 3-phosphoglycerate.
   • High-energy phosphate transferred to ADP.
   • Produces ATP.
   • Substrate-level phosphorylation.
   • Reversible.

8. Phosphoglyceromutase
   • 3-phosphoglycerate becomes 2-phosphoglycerate.
   • Phosphate moved to another carbon.
   • Reversible.

9. Enolase
   • 2-phosphoglycerate becomes phosphoenolpyruvate (PEP).
   • Water removed.
   • Creates high-energy PEP.
   • Reversible.

10. Pyruvate Kinase
    • PEP becomes pyruvate.
    • Phosphate transferred to ADP.
    • Produces ATP.
    • Irreversible.

Overall Glycolysis Equation
• Glucose + 2 ADP + 2 Pi + 2 NAD+ → 2 pyruvate + 2 ATP + 2 NADH
• Input: 2 ATP
• Output: 4 ATP
• Net: 2 ATP

Fate of Pyruvate

Aerobic respiration:
• Occurs in mitochondria.
• Pyruvate becomes acetyl-CoA then CO2.
• Produces large amounts of ATP.
• Requires oxygen.

Ethanol fermentation:
• Pyruvate becomes acetaldehyde then ethanol + CO2.
• Regenerates NAD+.
• Occurs in cytoplasm.

Lactic acid fermentation:
• Pyruvate becomes lactate.
• Regenerates NAD+.
• Occurs in cytoplasm.

Energetics of Fermentation
• Only 7 percent of glucose’s energy is released.
• Only 30 percent of that is trapped as ATP.
• Overall efficiency is about 2 percent.
• Most energy remains in lactate or ethanol.
• Fermentation produces little ATP.

Why Ferment?
• Glycolysis needs NAD+.
• Without oxygen, NADH cannot be oxidised in mitochondria.
• Fermentation regenerates NAD+.
• Allows glycolysis to continue.
• Prevents energy shutdown.

TCA Cycle (Kreb’s Cycle)

• Occurs in the mitochondrial matrix

Purpose:

• Completely oxidises acetyl CoA to CO2.

• Produces NADH and FADH2 (electron carriers).

• Produces 1 GTP (equivalent to ATP).

• Regenerates oxaloacetate to continue the cycle.

Aerobic/Anaerobic Fate of Glucose

Without oxygen:

• Pyruvate becomes lactate (muscle, microbes) or ethanol + CO2 (yeast).

• NADH is re-oxidised to NAD+.

• Allows glycolysis to continue.

With oxygen:

• Pyruvate becomes acetyl CoA.

• Acetyl CoA enters the TCA cycle.

• NADH and FADH2 feed electrons into the electron transport chain to generate ATP.

Steps

1. Pyruvate to Acetyl CoA
   • Pyruvate (3C) is decarboxylated to acetyl CoA (2C).
   • Releases CO2.
   • Produces NADH.
   • This is the link between glycolysis and the TCA cycle.

2. Acetyl CoA + Oxaloacetate to Citrate
   • Acetyl CoA (2C) combines with oxaloacetate (4C) to form citrate (6C).
   • CoA is released and recycled.

3. Citrate to Isocitrate
   • Citrate is rearranged to isocitrate.
   • Prepares molecule for oxidation.

4. Isocitrate to Alpha-Ketoglutarate
   • Isocitrate (6C) is oxidised to alpha-ketoglutarate (5C).
   • Produces NADH.
   • Releases CO2.

5. Alpha-Ketoglutarate to Succinyl CoA
   • Alpha-ketoglutarate (5C) is oxidised to succinyl CoA (4C).
   • Produces NADH.
   • Releases CO2.
   • By this step, all 3 carbons from pyruvate have been released as CO2.

6. Succinyl CoA to Succinate
   • Succinyl CoA becomes succinate.
   • Produces GTP (equivalent to ATP).
   • Only substrate-level phosphorylation in the TCA cycle.

7. Succinate to Fumarate
   • Succinate is oxidised to fumarate.
   • Produces FADH2.
   • Only step producing FADH2.

8. Fumarate to Malate
   • Fumarate is converted to malate.

9. Malate to Oxaloacetate
   • Malate is oxidised to oxaloacetate.
   • Produces NADH.
   • Oxaloacetate is ready to combine with another acetyl CoA.

Yield

Per pyruvate:

• 3 NADH

• 1 FADH2

• 1 GTP

• 2 CO2

Per glucose:

• Glucose produces 2 pyruvate.

• Therefore:

  • 6 NADH

  • 2 FADH2

  • 2 GTP

  • 4 CO2

ETC (Phase II)

• NADH produces 3 ATP.

• FADH2 produces 2 ATP.

• Oxygen is the final electron acceptor.

• Water is produced.

• ATP Yield from TCA Cycle and ETC (per pyruvate)

  • 4 NADH = 12 ATP

  • 1 FADH2 = 2 ATP

  • 1 GTP = 1 ATP

  • Total = 15 ATP per pyruvate

• ATP Yield from TCA Cycle and ETC (per glucose)

  • 2 pyruvate = 2 x 15 = 30 ATP

Glycolysis NADH and the Glycerol Phosphate Shuttle

• Glycolysis produces 2 NADH in the cytoplasm.

• These cannot directly enter mitochondria.

• Glycerol phosphate shuttle transfers electrons into mitochondria.

• Converts cytoplasmic NADH into mitochondrial FADH2.

• Each FADH2 produces 2 ATP.

• Total from glycolysis NADH = 4 ATP.

Total ATP from Complete Oxidation of Glucose

• Glycolysis: 2 ATP

• TCA/ETC: 30 ATP

• Shuttle: 4 ATP

• Total: 36 ATP per glucose

Overall Aerobic Respiration Equation

• Glucose + 6 O2 + 36 ADP + 36 Pi → 6 CO2 + 6 H2O + 36 ATP

Energetics

Energy released from glucose oxidation: 2870 kJ.

Energy trapped in ATP: 1098 kJ.

Efficiency: 38%

Much higher than fermentation (7% released, 2% trapped).

Summary of Glucose Metabolism

• Glycolysis: glucose to pyruvate, 2 ATP, 2 NADH.

• Without oxygen: fermentation regenerates NAD+.

• With oxygen: pyruvate to acetyl CoA, enters TCA cycle.

• TCA cycle produces NADH, FADH2, GTP.

• ETC uses NADH and FADH2 to generate ATP.

• Total ATP per glucose: 36.

ETC and Oxidative Phosphorylation

Location:

• ETC and ATP synthase embedded in inner mitochondrial membrane

• Cells with high energy demand have many mitochondria with extensive cristae

Respiratory control

• ETC has multiple steps with free energy release

• Electrons do not flow unless ADP is available

• ADP availability regulates ETC and OP

• Prevents unnecessary fuel burning

Transport mechanisms

• Outer membrane permeable (<5000 MW)

• Inner membrane impermeable

• ADP and Pi must enter mitochondria

• ATP must exit mitochondria

• Transport via translocase proteins

Inhibitors

ETC inhibitors

• Cyanide, azide, hydrogen sulphide, carbon monoxide

  • Block cytochrome a + a3

    • Antimycin

• Blocks cytochrome b → c1

  • Rotenone

• Inhibits NADH oxidoreductase

Oxidative phosphorylation inhibitors

• Oligomycin

  • Inhibits ATP synthase

    • Atractyloside

• Inhibits translocase proteins

Uncoupling

• ETC and OP normally coupled

• Uncouplers allow electron flow without ATP production

• Proton gradient destroyed

• Energy released as heat

• Occurs naturally in brown adipose tissue (thermogenesis)

Synthetic uncoupler: DNP

• 2,4‑dinitrophenol

• Shuttles protons across membrane

• Eliminates proton gradient

• Energy lost as heat

• Used as a diet pill in 1930s; banned due to deaths

• Still used illegally

Electron Transport Chain

• Electrons removed from NADH

• Passed through redox intermediates to O2

• O2 is final electron acceptor

• ETC proteins contain Fe atoms (Fe3+ / Fe2+)

• Purpose: oxidise NADH and FADH2, transfer electrons to O2, pump H+

ETC Pathway

• NADH oxidoreductase

• FMN

• Coenzyme Q

• Cytochrome b

• Cytochrome c1

• Cytochrome c

• Cytochrome a + a3

• O2 → H2O

FADH2 entry

• FADH2 enters later in the chain

• Produces less ATP than NADH

ETC complexes

• Located in inner mitochondrial membrane

• NADH oxidation pumps ~10 protons

Coupling of ETC and OP

• Electron flow pumps H+ into intermembrane space

• Proton gradient drives ATP synthesis

• Chemiosmotic theory: electron flow → H+ pumping → gradient → ATP

Oxidative phosphorylation

• ATP synthase uses H+ returning to matrix

• ADP + Pi + H+ → ATP + H2O

• Proton gradient provides energy

• ETC and OP are coupled processes

Gluconeogenesis

Topic 4

Photosynthesis

Prokaryotic Metabolism and Growth

Topic 5

DNA, RNA, and Transcription

Translation and the Genetic Code

Topic 6

DNA Replication

Gene Control

DNA Mutation

Topic 7

Eukaryote Cell Growth and Death

Chromosome and Mitotic Process

Meiosis and Genetic Diversity

Single Gene Inheritance

Stem Cell Differentiation and Cancer

Cellular Transport and Blood Cells

Cellular Transport and Blood Cells

Cellular Signalling and Movement

Genes and Genome

Topic 8