Topic 5-9: Lipids and Membranes PDF

Topic 5 – Lipids and membrane dynamics Lipids: The smallest of the macromolecules Not polymers made of monomers Hydrophobic – do not mix well with water Made primarily of hydrocarbons 3 classes of lipids – fats, phospholipids, steroids Fats: - Glycerol: alcohol - Fatty acids:...

Topic 5 – Lipids and membrane dynamics Lipids: The smallest of the macromolecules Not polymers made of monomers Hydrophobic – do not mix well with water Made primarily of hydrocarbons 3 classes of lipids – fats, phospholipids, steroids Fats: - Glycerol: alcohol - Fatty acids: hydrocarbon chain (hydrophobic chains) - Glycerol bound to fatty acids by a dehydration reaction (creates an ester linkage) - Triacyl glyceride (triglyceride): one glycerol bound to 3 fatty acids (fatty acids can be all the same or can be diFerent) Saturated fatty acids: - No double bonds between carbon atoms (saturated with hydrogen atoms, most animal fats, solid at room temperature) Unsaturated fatty acids: - One or more double bonds between carbon atoms (cis double bonds cause a kink in the hydrocarbon chain, fats of plants and fish, liquid at room temperature) Hydrogenated oils: Unsaturated fats that gave been converted into saturated fats - Hydrogen atoms have been added to the hydrocarbon chains Ex. peanut butter, margarine Allows lipids that would normally be liquid to be solid at room temperature Prevents lipids from separating out in liquid form. Fats in our diet: Saturated fats may contribute to cardiovascular diseases (atherosclerosis: plaques develop within blood vessels) Hydrogenation can produce trans double bonds: - Contributes to coronary heart disease - Baked goods, processed foods - Canada has banned artificial trans fats in foods Essential fatty acids: - Cannot be made by our bodies, therefore we need to get them in our diet Ex. omega-3 fatty acids are required for normal growth Functions of fats: - Energy storage - A gram of fat stores more than twice as much energy as a gram of a polysaccharide much as starch - Plants store energy as starch (very bulky, plant seeds store fats) - Animals store energy as fats (more compact, less bulky, stored in adipose cells, cushions vital organs, acts as insultation for the body) Phospholipids: Major constituents of cell membranes Glycerol bound to 2 fatty acids and one phosphate group - Phosphate has a negative electrical charge (has a charge – POLAR {negative charge}) - Small charged or polar molecules can be linked to the phosphate group Hydrocarbon tails are hydrophobic Phosphate group and its attachment form a hydrophilic head Phospholipids self-assemble into double layered structures called bilayers. Steroids: Carbon skeleton consisting of 4 fused rings Distinguished by the chemical groups attached to the rings Ex. cholesterol (precursor of other steroids such as sex hormones. Synthesized in the liver and obtained in the diet. High levels in the blood may contribute to atherosclerosis) Cellular membranes: Lipids, proteins, carbohydrates Phospholipids are amphipathic (hydrophilic and hydrophobic regions) Most membrane proteins are amphipathic (embedded in the phospholipid bilayer, hydrophilic region is in contact with water in the cytosol and the extracellular fluid, hydrophobic regions are in contact with the fatty acid tails) Fluid mosaic model: Protein molecules bobbing in a fluid bilayer of phospholipids Groups of proteins can be associated with specialized patches where they carry out common functions. Lipid rafts: specific lipids found in these patches. Van der Waals interactions: weak hydrophobic Interactions that hold the membrane together. Lipids shift laterally very quickly (107 times per second) Lipids can flip-flop across the membrane Proteins move more slowly (can be highly directed) Proteins can be held immobile by the cytoskeleton or extracellular matrix Fluidity aFects permeability and ability of membrane proteins to move. (too solid or fluid and proteins may be inactivated) Membranes react to temperature: - Cold phospholipids pack tightly, membrane solidifies - Temperature at which a membrane solidifies depends on the type of pf fatty acids - Steroids are inserted between phospholipids to aFect fluidity (at high temperatures (37 degrees) makes the membrane less fluid by restraining lipid movement, at low temp it hinders the close packing of phospholipids so that the membrane does not solidify) - Fluidity buFer – resists change in membrane fluidity. Evolutionary adaptations maintain the appropriate membrane fluidity for the specific environmental conditions  - Ex. fish living unextreme cold have high proportion of unsaturated hydrocarbons - Ex. prokaryotes in thermal hot springs include unusual lipids that may prevent excessive fluidity. - Ex. plants that tolerate extreme cold increase the percentage of unsaturated phospholipids in autumn. DiFusion: a substance diFuses down its concentration gradient (from an area of high concentration to an area of low concentration) DiFusion is a spontaneous process (no input of energy) Ex. dissolved oxygen diFuses into the cell across the plasma membrane (the cell uses the oxygen which constantly lowers its concentration inside the cell) Osmosis: (diFusion of water) DiFusion of free water across a selectivity permeable membrane. Free water molecules are the ones not associated with hydrophilic substances. Side with higher solute concentration has lower free water concentration. Water diFuses from the region of higher free water concentration (low solute) to that of lower free water concentration (high solute). Tonicity: the ability of a surrounding solution to cause a cell to gain or lose water. - Depends on the concentration of solutes that cannot cross the membrane (non- penetrating solutes) in relation to the inside of the cell Three types of solutions: - Hypotonic, isotonic, hypertonic Hypotonic solution: (best for plants) Lower concentration of solutes outside the cell than inside the cell (water enters the cell) Animal cells: cell will swell and lyse Plant cells: rigid cell wall expands and exerts a back pressure Turgor pressure: pressure that opposes further water uptake Turgid: cell that is very firm (this is the healthiest state for most plant cells) Isotonic: (best for animals)  Equal solute concentration inside and outside the cell (water diFuses in an out of the cell at the same rate) Animal cells: volume of an animal cell is stable, the cell is in its healthiest state Plant cells: there is no build-up of turgor pressure (cells become limp (flaccid) and the plants wilts Hypertonic solution: (not good for anyone) Higher concentration of solutes outside the cell than inside the cell (water exits the cell) Animal cell: water is lost, the cell shrivels and dies. (an increase in salinity of a lake can kill the animals) Plant cells: the plasma membrane pulls away from the cell wall at multiple places (plasmolysis) (the plant wilts and often dies) Plasmolysis also occurs in bacteria and fungi Managing tonicity: Cells without cell walls can’t tolerate excessive uptake or loss of water Cells live in isotonic surroundings - Seawater is isotonic to many marine invertebrates - Extracellular fluid around our cells is isotonic to our cells Osmoregulation: the control of solute concentrations and water balance - Paramecium plasma membranes are less permeable to water than other membranes - Contractile vacuoles forces water out of the cell as fast as it enters - Bacteria and archaea living in hypersaline environments have mechanisms that balance their internal and external solute concentrations to ensure water doesn’t move out of the cell. Peripheral membrane proteins: - Proteins determine the function of the membrane - Peripheral proteins: appendages loosely bound to the surface of the membrane (often exposed to parts of their integral proteins) Integral membrane proteins: Integral proteins: penetrate the hydrophobic interior of the lipid’s bilayer  - Transmembrane proteins: span the entire membrane - Hydrophobic regions consist of non-polar amino acids usually coiled in helices - Hydrophilic regions are exposed to the aqueous solution on either side of the membrane - Hydrophilic channels allow passages through the membrane. Cell-cell recognition: cells ability to distinguish one type of neighboring cell from another. - Ex. embryonic development - Ex. rejection of foreign cells by the immune system Cells bind to molecules on the extracellular surface Membrane carbohydrates: branched chains of less than 15 sugar units - Glycolipids: carbohydrate bonded covalently to lipids - Glycoproteins: carbohydrates bonded covalently to proteins - Ex. A, B, AB, O blood types are variations in the carbohydrates in glycoproteins on the surface of red blood cells Synthesis and sidedness of membranes: - Membranes have inside and outside faces - Two layers of lipids may diFer in lipid composition - Proteins have directional orientation in the membrane - Asymmetrical arrangement of proteins, lipids, and carbohydrates in the plasma membrane is determined as a membrane is being built in the ER and Golgi apparatus. Permeability of the lipid bilayer: Small non-polar molecule dissolve in the lipid bilayer and cross easily - CO2, O2 - Hydrophobic molecules Ions and polar molecules cannot pass through the membrane directly - Hydrophilic cannot interact with the hydrophobic interior of the membrane - Small polar molecules (ex. sugars, water) pass very slowly through the membrane - Ions are even less likely to pass through Membrane proteins play key roles in regulating transport Transport proteins:  - Span the membrane and help molecules avoid contact with lipid bilayers - Channel proteins: hydrophilic channel that molecules and atomic ions use as a tunnel (ex. aquaporin allows up to 3 billion water molecules (10 at a time) per second through the membrane) Carrier proteins: change shape as they shuttle molecular passengers across the membrane. Protein specificity: - Transport proteins are specific for the substance they translocate (move): Only a certain substance or small group of related substances can cross with each protein Ex. carrier protein in the plasma membrane of red blood cells transports glucose across the membrane 50,000 times faster than it passes through on its own. (it rejects fructose which is an isomer of glucose) Passive transport: diFusion of a substance across a membrane with no energy investment Molecules diFuse due to their constant motion DiFusion: movement of particles so that they spread out into the available space. Dynamic equilibrium: after diFusion there are equal concentrations of the molecule on both sides of the membrane. Facilitated diFusion: passive transport through transmembrane transport proteins (movement down a concentration gradient) Channel proteins: provide corridors for specific molecules or ions to pass through - Ion channels: transport ions - May open and close in response to stimuli Ex. nerve cells ion channels open in response to electrical stimuli Ex. open or close when a specific substance binds to the channel Carrier proteins: a subtle change in shape translocate the solute-binding site across the membrane. - Triggered by the binding and release of the transported molecule Active transport (against concentration gradient): uses energy to move solutes against their concentration gradients Carrier proteins Cell maintains internal concentrations of small solutes diFerent from the environmental concentration ATP hydrolysis provides the energy for most active transport - Transfers its terminal phosphate group directly to the transport protein - Induces the protein to change shape Ex. sodium-potassium pump Animal cells have internal concentration of K+ that are higher than Na+ that are lower than the extra cellular fluid (pumps Na+ out and K+ into the cell) Ion pumps maintain membrane potential - Voltage: electrical potential energy, a separation pf opposite charges - Cytoplasmic side of the membrane is negative in charge relative to the extracellular side (unequal distribution of anions and cations on the 2 sides) - Membrane potential: voltage across a membrane (ranges from -50 to -200mV) Passive transport of cations into the cell and anions out of the cell is favoured Electrochemical gradient - Active transport is needed when electrical forces oppose the concentration gradient - Membrane proteins may contribute to membrane potential (ex. sodium-potassium pump) net transfer of positive charge from cytoplasm to the extracellular fluid Electrogenic pump: transport proteins that generates voltage across a membrane - Sodium-potassium pump in animals - Proton pump in plants, fungi, and bacteria (actively transports protons (hydrogen ions) out of the cell Generating voltage across membranes stores energy that can be trapped for cellular work (ATP synthesis during cellular respiration) Cotransport: - Coupled transport by a membrane protein - A solute can do work as it moves down its concentration gradient across the membrane - Cotransport: couples the downhill diFusion of a solute to the transport of a second substance against its concentration gradient  - Ex. plant cells use H+ generated by proton pumps to drive the active transport of amino acids, sugars, and other nutrients into the cell (plants use H+/ sucrose cotransporter to load sucrose into cells in the veins of leaves) ____________________________________________________________________________________ Topic 6 - proteins and enzyme function: Proteins: nearly every function of a living being depends on proteins Proteins are more than 50% of the dry mass of most cells Humans have tens of thousands of diFerent proteins Each has a specific structure and function. Storage protein: storage of amino acids Hormonal proteins: coordination of an organism’s activities Receptor proteins: response of cell to chemical stimuli Defensive proteins: protection against diseases Transport proteins: transport of substances Structural proteins: support Contractile and motor proteins: movement Enzymes: most are proteins (some small RNAs act as enzymes) - Enzymes regulate metabolism by acting as catalysts - Catalyst – chemical agent that selectively speeds up chemical reactions without being consumed by the reaction Enzymatic proteins – function is selective acceleration of chemical reactions. Ex. digestive enzymes catalyse the hydrolysis of bonds in food molecules. Protein structure: 20 amino acids - Polypeptide: polymer of amino acids - Protein: biologically functional molecule made up of one or more polypeptides, each folded and coiled into a specific 3D structure. Amino acids: Sidechains  - Polar – hydrophilic - Non-polar – hydrophobic - Electrically charged (negative – acidic due to the presence of a carboxyl group, positive – basic due to the presence of an amino acid) The physical and chemical properties of the side chain determine the characteristics of an amino acid (aFects the functional role in a polypeptide). Peptide bond: the carboxyl group of one amino acid and the amino group of another are joined by a dehydration reaction Polypeptide backbone: repeating sequences of atoms (R groups extend from the backbone) One end of the polypeptide has a free amino group (N-terminus) and the other end has a free carboxyl group (C-terminus) Protein structure and function: A polypeptide chain may fold spontaneously Folding is driven and reinforced by the formation of various bonds between parts of the chain. Globular proteins: roughly spherical in shape Fibrous proteins: shaped like long fibres Form determines function: A protein’s specific structure determines how it works The function of a protein depends on its ability to recognize and bind to other molecules (lock and key) - Ex. endorphins bind receptors in the brain to make us happy Primary structure, secondary structure, tertiary structure, quaternary structure Sickle-cell disease: - A small simple change in amino acid sequence can greatly impact the formation of the protein. Denaturation: - Protein structure depends on the physical and chemical environment - Incorrect pH, salinity, temperature, etc. alters the weak bonds within a protein. Denaturation: the protein unravels and loses its native shape - The protein is inactive - Chemicals that disrupt hydrogen bonds, ionic bonds, and disulfide bridges - Hydrophobic solvents for the protein to fold inside out. - Excessive heat agitates the polypeptide to overpower the weak interactions. Protein folding: The folding process is very complex - Can be intermediate structures before the final stable shape - Looking at a mature structure does not reveal the stages of folding Many diseases are associated with misfolded proteins - Cystic fibrosis, Alzheimer’s, Parkinson’s, mad cow disease Some proteins are hard to determine the structure of - They do not have a distinct 3D structure until they interact with a target protein or other molecule - Intrinsically discorded proteins – they are flexible and indeterminate because they may require binding with diFerent targets at diFerent times. - May account for 20-30% of mammalian proteins Cellular metabolism: Metabolism: the totality of an organism’s chemical reactions Metabolic pathway: a molecule is altered in a series of steps resulting in a certain product - Each step of a metabolic pathway is catalyzed by a specific enzyme Roles of a metabolic pathways:  Release consumption of energy Conversion of small molecules into diFerent small molecules Digestion of large molecules into diFerent small molecules Export of chemical products to be used in other cells Catabolic pathways: releases energy by breaking down complex molecules to simpler compounds (ex. cellular respiration) Anabolic pathways: consumes energy to build complicated molecules from simpler ones. (Also called biosynthetic pathways. Ex. the synthesis of amino acids from simpler molecules) Bioenergetics: the study of how energy flows through living organisms. Energy: the capacity to cause change - Kinetic energy – energy associated with the relative motion of objects - Thermal energy – kinetics energy associated with the random movement of atoms or molecules - Heat – thermal energy transferred from one object to another - Potential energy – energy that matter possesses because of its location or structure. - Chemical energy – potential energy available for release in chemical reaction (complex molecules waiting to be broken down are high in chemical energy) Energy transformations: - Thermodynamics – the study of energy transformations that occur in a collection of matter - System – the matter under study - Surroundings – everything outside the system - Isolated system – unable to exchange either energy or matter with its surroundings - Open system – energy can be transferred between the system and its surroundings Laws of thermodynamics: First law: the energy of the universe is constant - Energy can be transferred and transformed, but it cannot be created or destroyed - The principle of conservation of energy Second law: every energy transfer or transformation increases the entropy of the universe - Most usable forms of energy are at least partly converted to thermal energy and released as heat. Entropy: Disorder: how dispersed the energy is in a system and how many diFerent energy levels are present. The loss of usable energy as heat makes the universe more disordered. Entropy: a measure of disorder or randomness. The more randomly arranged a collection of matter, the greater its entropy. Order can increase locally but there is an unstoppable trend towards randomization of the universe. Increasing entropy takes the form of increasing amounts of heat and less ordered forms of matter. If a process leads to an increase in entropy, that process can proceed without requiring an input of energy. (spontaneous process: energetically favourable) A process that leads to a decrease in energy is non-spontaneous - Happens only if energy is supplied - Entropy is decreased Ex. water flows downhill spontaneously but to move uphill it needs an input of energy (a machine can pump water against gravity). Biological order and disorder: Living systems increase the entropy of their surroundings - Cells create ordered structures from less organized starting materials - Cells take in organized forms of energy and replace them with less ordered forms. Complex organisms arose from simpler ancestors Increase of cellular organization overtime doesn’t violate the second law of thermodynamics - Entropy of a small system (organisms) may decrease as long as the entropy of the entire universe increases - Organisms are islands of low entropy in an increasingly random universe. Change in free energy: Gibbs free energy of a system = G Free energy: the portion of a systems energy that can perform work when temperature and pressure are uniform throughout a system. Change in free energy = ^H = change in the systems enthalpy (in biological systems this is equivalent to total energy) ^S = change in the systems entropy T = absolute temperature in Kelvin (K = *C + 273) We measure ^G using chemical means. - Negative ^G are spontaneous, positive or zero ^G require an input of energy - Less free energy means the system in its final state is less likely to change and is therefore more stable. - Free energy is a measure of a systems instability, its tendency to change to a more stable state. Exergonic reactions: release of energy - ^G is negative - Occur spontaneously (energetically favourable) - C6H12O6 + 6O2 > 6CO2 + 6H2O ^G = -23870 kJ/mol “Energy stored in bonds” means the potential energy that can be released when new bonds are formed if the products are of lower free energy than the reactants. Endergonic reactions: absorbs free energy from its surroundings - Stores free energy in molecules - G increases, ^G is positive - Reactions are nonspontaneous and the magnitude of ^G is the quantity of energy required to drive the reaction - Ex. photosynthesis Equilibrium: Most chemical reactions are reversible and proceed until the forward and backward reactions occur at the same rate. As a reaction proceeds towards equilibrium the free energy in the mixture of reactants and products decreases. A process is spontaneous and can perform work only when it is moving toward equilibrium If a reaction is endergonic in one direction it will be exergonic in the other direction Cellular respiration and photosynthesis are opposite of each other. (Cellular respiration releases 2870kJ/mol of energy, photosynthesis uses 2870 kJ/mol of energy.) Equilibrium and metabolism: Systems at equilibrium are at minimum of G and can do no work A cell that reaches a metabolic equilibrium is dead. Defining feature of life: metabolism is never at equilibrium The flow of materials in and out of the cell and the continuous use of products keeps the metabolic pathways from reaching equilibrium. A catabolic pathway releases free energy in a series of reactions. Cellular work: - Chemical work: the pushing of endergonic reactions that would not occur spontaneously - Transport work: the pumping of substances across membranes against the direction of spontaneous movement. - Mechanical work: such as the beating of cilia, contraction of muscle dells, movement of chromosomes during cellular reproduction. - Energy coupling: the use of an exergonic process to drive and endergonic reaction. - ATP: the primary source of energy that powers cellular work. ATP: - Adenosine triphosphate - Sugar ribose with the nitrogenous base adenine and a chain of three phosphate groups. - Bonds between the phosphate groups are broken by hydrolysis - Terminal phosphate group is released - ATP becomes adenosine diphosphate (ADP) Phosphorylation: the transfer of a phosphate group from ATP to another molecule Phosphorylated intermediate: recipient molecule with the phosphate group covalently bonded to it (more reactive than the original molecule) Transport and mechanical work: ATP hydrolysis leads to the change in a proteins shape and often its ability to bind another molecule. Regeneration of ATP: A muscle cell recycles its entire pool of ATP in less than a minute. - 10 million molecules of ATP consumed and regenerated per second per cell Speed of reactions: - A spontaneous chemical reaction can be slow that it is almost imperceptible - Changing one molecule into another requires contorting the starting molecule into a more unstable state before the reaction can proceed. - When the new bonds of the product form, energy is released as heat and the molecules return to stable shapes with lower energy than the contorted state. Enzyme: macromolecule that acts as a catalyst. - Catalyst: chemical agent that speeds up a reaction without being consumed by the reaction. Activation energy (EA): Energy required to contort the reactant molecules so that the bonds can break (free energy of activation) Can be supplied by thermal energy that accelerates the reactant molecules. When the molecules have absorbed enough energy, they are in an unstable condition known as the transition rate. Enzymes: - Catalysis: catalyst selectively speeds up a reaction without itself being consumed. - Enzymes catalyze a reaction by lowering the EA barrier Enables the reactant molecule to absorb enough energy to reach the transition state at moderate temperatures - Enzymes are very specific for the reactions they catalyze. Substrate specificity: - Substate: the reactant an enzyme act on  - Enzyme-substrate complex: the enzyme bound to its substrates - Enzymes convert the substrate to the product - Most enzymes names end in “ase” Ex. sucrase will only act on sucrose and will not bind to other disaccharides The specificity of an enzyme results from its shape which is a consequence of amino acid sequence. Enzyme- substrate complex: Active site: regio of the enzyme molecule that binds to the substrate - Complementary fit between the shape and charge of the active site and the substrate. Shape of is enzyme is dynamic Induced fit: shape change in the active site so that the enzyme binds the substrate more tightly. - Interactions between chemical groups - Forces chemical groups of the active site into positions that enhance their ability to catalyze the chemical reaction. *Catalytic cycle Enzymatic function: - Provides a microenvironment conductive to a particular type of reaction Ex. active site as a pocket of low pH in a neutral cell facilitates H+ transfer to the substrate. - Amino acids in the active site directly participate in the chemical reaction Can involve brief covalent bonding between the substrate and the site of an amino acid of the enzyme. Saturation: The more substrate molecules that are available, the more frequently they access the active sites of the enzyme molecules. Upper limit for speed of reactions - Substrate concentration will be high enough that all enzyme molecules have their active site engaged. - As soon as a product leaves an active site, another substrate molecule enters. - The enzyme is saturated - Rate of the reaction is determined by the speed at which the active site converts substrate to product - The only way to increase the rate of product formation is to add more enzymes. Enzymatic conditions: Optimal conditions: the conditions under which an enzyme works best. This favours the most active shape of the enzyme. Temperature: - Rate of an enzymatic reaction increases with increasing temp. - Substrates collide with active sites more frequently. - Too hot and the proteins will denature. pH: - Acidic or basic environments aFects enzymes - H+ and OH- interact with chemical groups on amino acids. Cofactors: non-protein helpers required for catalytic activity - Ex. perform electron transfers - Can be bound tightly to the enzyme (permanent) or may bind loosely and reversibly. Inorganic such as metal atoms - Zinc, iron, copper Organic such as vitamins (coenzymes) Enzyme inhibitors: Some bind covalently which is usually irreversible. Some bind weakly which is usually reversible. Competitive inhibitors: reduce productivity of enzymes by blocking substrates from entering the active sites. - Inhibition is overcome by increasing the concentration of substrate. Non- competitive inhibitors: impede enzymatic function by binding to another part of the enzyme. - Causes the enzyme to change shape so that the active site becomes less eFective. Allosteric regulation of enzymes: Allosteric regulation: a proteins function at one site is aFected by the binding of a regulatory molecule to a separate site. Most enzymes controlled allosterically are constructed from 2 of more subunits, each with its own active site. The complex oscillates between a catalytically active shape and a catalytically inactive shape. An activating or inhibiting regulatory molecule binds a regulatory molecule binds a regulatory site (allosteric site) often located where the subunits join. Activator: stabilizes the shape that has functional active sites. Inhibitor: stabilizes the inactive form. Cooperativity: another type of allosteric activation Substrate molecules binding one active site in a multi-subunit enzyme triggers a shape change to all subunits to increase catalytic activity at the other active sites. Amplifies the response of enzymes to substrates. Feedback inhibition: The metabolic pathway is halted by the inhibitory binding of its product to an enzyme that acts early in the pathway. Allosteric inhibition of the enzyme by the final product of the reactions. Prevents the cell from easting resources when there is already enough of a product. Localization of enzymes: Teams of enzymes for a specific metabolic pathway can be arranged into a multienzyme complex to facilitate the sequence of reactions. Some have fixed locations in the cell and act as structural components of membranes. Others are in solution within membrane-enclosed eukaryotic organelles. ____________________________________________________________________________________ Topic 7: carbohydrates and catabolic pathways Monosaccharides: simple sugars, molecular formulas are multiples of CH2O Sugars: - Carbonyl group - Multiple hydroxyl groups - Aldose sugar (aldehyde) - Ketose sugar (ketone) Size ranges from 3-7 carbons long (most are hexoses, trioses and pentoses are common) - Isomers: placement of parts around one asymmetric carbon Linear and ring forms Abbreviated ring structure Most pentoses and hexoses form ring structures Most stable than linear structures. Disaccharides: 2 monosaccharides joined by a glycosidic linkage using a dehydration reaction - Ex. maltose = 2 molecules of glucose - Ex. sucrose = glucose + fructose - Ex. lactose = glucose + galactose Lactose intolerance: people lack the enzyme lactase Polysaccharides: hundreds or thousands of monosaccharides joined by glycosidic linkages. Function: - Storage – starch, glycogen - Building materials – cellulose, chitin Starch: Storage molecule of plants Granules inside plant plastids such as amyloplasts Polymer of glucose monomers Joined by 1-4 linkages 1-6 linkages at branching points Glycogen: (animals store up their glucose) Vertebrate storage molecule in liver and muscles cells Extensively branched polymer of glucose like starch Branched structure means more free ends for hydrolysis Human glycogen stores are gone in a day unless we eat Low carbohydrate diets can result in weakness and fatigue. Cellulose: Components of plant cell walls Most abundant organic compound on earth Glucose monomer, 2 slightly diFerent structures (a and b) 1-4 linkages, unbranched Forms cable-like microfibrils Insoluble fibre – we lack the enzyme to hydrolyze the B linkage Some prokaryotes and protists can digest cellulose Chitin: Exoskeleton of arthropods, cell wall of fungi B linkages between glucose monomers with a nitrogen containing attachment Proteins chemically link pieces of chitin together  Becomes encrusted with calcium carbonate Catabolic pathways: metabolic pathways that release stored energy by breaking down complex molecules (high energy system to low energy system) - Transfers electrons from fuels molecules to other molecules Fermentation: degradation of organic fuel without using oxygen Aerobic respiration: degradation of organic fuel without using oxygen (cellular respiration, fuel: carbohydrates, fats, proteins molecules) Cellular respiration: C6H12O6 + 6 O2 → 6 CO2 + 6 H2O + Energy (ATP + heat) Degradation of glucose Exergonic: Releases energy (ΔG = -2870 KJ/mol) Energy from catabolism is captured in the bonds of ATP Redox reactions: Oxidation: loss of electrons (electron donor, reducing agent) Reduction: gain of electrons (electron acceptor: oxidizing agent) The degree of electron sharing in covalent bonds may change Electrons lose potential energy when they move from a less electronegative atom to a more electronegative atom. Cellular respiration is a redox reaction: - Oxidation of organic fuel molecules (loss of hydrogen) - Reduction of oxygen (gaining hydrogen) - Electrons lose potential energy and energy is released - We see the transfer of hydrogen atoms (one proton, one electron) NAD+ reduction to NADH - Glucose is broken down in a series of steps that remove electrons - NAD+  Nicotinamide adenine dinucleotide Made from the vitamin niacin Coenzyme that carries electrons from one molecule to another NAD+ is reduced to NADH Stages of cellular respiration: Substrate level phosphorylation: - Substrate-level phosphorylation: phosphate group is transferred from a substrate to ADP to make ATP - Method by which ATP is produced in glycolysis and the citric acid cycle STAGE 1: glycolysis: - Glucose is split into two 3-carbon sugars which are oxidized and rearranged to make two molecules of pyruvate  - Energy investment phase – ATP is used - Energy payoF phase – ATP is produced by substrate level phosphorylation, NAD+ is reduced to NADH - Net yield: 2 ATP and 2 NADH Steps of glycolysis – energy investment phase: Steps of glycolysis – energy payoF phase: STAGE 2: Pyruvate oxidation and the citric acid cycle: - Pyruvate oxidation Pyruvate enters the mitochondria in eukaryotes 1. Carboxyl group is removed – release of CO2 2. Oxidation of carbon to form acetate 3. Coenzyme A is attached to acetate to form acetyl CoA CoA is made from vitamin B Citric acid cycle: Tricarboxylic acid cycle (TCA cycle) or Krebs cycle Acetyl CoA is oxidized to CO2 Cycle generation 1ATP  3 NADH 1 FADH2 2 CO2 Acetate binds to oxaloacetate to form citrate 2 carbon atoms break oF as molecules of CO2 Oxaloacetate is regenerated to be used again. STAGE 3; Oxidative phosphorylation: Glycolysis and the citric acid cycle produce only 4 ATP molecules per glucose molecule (substrate level phosphorylation) Most energy is still tied up in the electrons carried by NADH and FADH2 2 steps to release this energy and trap it in the bonds of ATP - Electron transport chain - Chemiosmosis Electron transport chain (ETC): Molecules in a membrane: mostly integral membrane protein complexes NADH shuttles electrons to the starting molecule of the ETC Electrons are passed from carrier to molecule releasing energy at each step.  O2 captures the electrons along with hydrogen ions at the end of the chain to form water. Cristae in the mitochondria increases surface area so that thousands of copies of ETC are present. Prosthetic groups (non-protein components) are bound to the proteins Carriers alternate between reduced and oxidized states as they accept and donate electrons. ETC establishes an H+ gradient Exergonic flow of electrons from NADH and FADH2 to pump H+ across the inner mitochondrial membrane into the intermemebrane space. Electron carriers accept and release protons (H+) along with electrons Proton-motive force: H+ gradient Chemiosmosis: ATP synthase: enzyme that makes ATP from ADP and an inorganic phosphate\ - Ion pump running in reverse - Uses the energy of an H+ ion gradient to power ATP synthesis - High H+ concentration in the intermembrane space (acidic) - Low H+ concentration in the mitochondrial matrix (basic) ATP production by cellular respiration: Glycolysis: 2 ATP via substrate level phosphorylation Citric acid cycle: 2 ATP via substrate level phosphorylation Oxidative phosphorylation: - 1 NADH = 2.5 ATP - 1 NADH2 = 1.5 ATP Total = about 32 ATP ADP > ATP stores 30.5 KJ/mole 34% of the energy in glucose is transferred to ATP Remainder is lost as heat - Maintains our body temperature - Dissipates through sweating and cooling mechanisms Hibernation: Internal body temperature is lower than normal but still much higher than the external air temperature Brown fat: tissue packed full of mitochondria - Uncoupling protein: inner mitochondrial membrane protein allows H+ to flow down their concentration gradient without generating ATP - Ongoing oxidation is stored fuels (fats), generating heat without ATP Fermentation and anaerobic respiration: Cells oxidize organic fuel and generate ATP without the use of oxygen. Anaerobic respiration; uses an electron transport chain - Some prokaryotes that live in anaerobic environments - Terminal electron acceptor is not oxygen - Ex. sulphate-reducing bacteria use SO42- as the final electron acceptor to make H2S as a by-product instead of H2O Fermentation: does NOT use an electron transport chain - Organisms use glycolysis for energy production (2 ATP) - Glycolysis produces NADH - NADH is recycled to NAD+ by transferring electrons to pyruvate Alcohol fermentation: Pyruvate is converted to ethanol CO2 is released Many bacteria and yeast Used for brewing, winemaking and baking Lactic acid fermentation: Pyruvate I reduced to form lactate Some fungi and bacteria (used for making cheese and yogurt) Human muscle cells when oxygen us scarce  - Sugar catabolism for ATP outpaces the supply of oxygen - Excess lactate is carried to the liver which converts it back to pyruvate Fermentation/anaerobic respiration/ anerobic respiration: Similarities: - Produce ATP by harvesting chemical energy of food - Use glycolysis to oxidize glucose and other organic fuels to pyruvate - NAD+ is the oxidizing agent that accepts electrons DiFerences: - The mechanism by which NADH is converted back to NAD+ - Amount of ATP produced (fermentation: 2 ATP, cellular respiration: up to 32 ATP) Classes of organisms: Obligate aerobes: require oxygen for survival (carry out only aerobic respiration) Obligate anaerobes: cannot survive in the presence of oxygen (carry out only fermentation or anaerobic respiration) Facultative anaerobes: can live with or without oxygen - Aerobic conditions: perform aerobic respiration - Anaerobic conditions: perform fermentation Evolution of glycolysis: Glycolysis is the first stage of aerobic and anaerobic respiration and fermentation Ancient prokaryotes used glycolysis to male ATP before oxygen was even present in our atmosphere Most widespread metabolic pathway on earth, suggesting it evolved very early in the history of life Occurs in the cytosol, does not require membrane-enclosed organelles. ____________________________________________________________________________________ Topic 8 - Photosynthesis Conversion of solar energy to chemical energy Autotrophs: produce organic molecules from CO2 and other inorganic materials (producers of thr biosphere) Photoautotroph: autotrophs that use light as a source of energy Heterotrophs: obtain organic material by consuming it (consumers of the biosphere) Site of photosynthesis: Enzymes and other necessary molecules are grouped together in biological membranes Originated in bacteria that have infolded regions of thr plasma membrane - Infolded plasma membrane functions similarly to the internal membrane of the chloroplast. Chloroplasts: site of photosynthesis in eukaryotic organisms Leaves and plant cells: Mesophyll cells: main location of chloroplasts - Tissue in the interior of the leaf - 30-40 chloroplast per cell Stromata (stroma): pores in a leaf that allow the entry of CO2 and the exit of O2 Veins: deliver water and sugars to all parts of the plant Chloroplast structure: Double membrane Stroma: fluid inside the inner chloroplast membrane Thylakoids: sacs of membrane suspended in the stroma Thylakoid space: space inside the thylakoids Grana (granum): stacks of thylakoids Chlorophyll: pigment molecule that absorbs light energy Photosynthesis: 6CO2 + 6H2O + LIGHT ENERGY > C6H12O6 + 602 This reaction is the opposite of cellular respiration Photosynthesis is a redox reaction: Energy + 6 CO2 + 6 H2O > C6H12O6 + 6O2 Electrons increase in potential energy when they move from water to sugar - Endergonic reaction - Energy is provided by light STAGES OF PHOTOSYNTHESIS: The light reactions: conversion of solar energy to chemical energy - Photophosphorylation: chemiosmosis adds a phosphate group to ADP The Calvin cycle: conversion of CO2 to organic molecules - Reduces carbon to carbohydrates - Dark reactions, light-independent reactions Sunlight: Electromagnetic energy, electromagnetic radiation - Travels in rhythmic waves Wavelength: the distance between the crests of electromagnetic waves - Electromagnetic spectrum: full range of radiation Visible light: can be detected as colours by the human eye Photons: Photon: discrete particles of light, packets of light energy - Each photon has a fixed quantity of energy - The shorter the wavelength, the greater the energy of each photon The atmosphere screens out a substantial amount of radiation Visible light is used in photosynthesis Photosynthetic pigments: Pigments: substances that absorb visible light (the colour we see is the wavelength of light that is reflected or transmitted) Spectrophotometer: instrument that measures the ability of a pigment to absorb various wavelength of light  Absorption spectrum: graph plotting the pigments light absorption versus wavelength Pigments: Chlorophyll a: key light capturing pigment (participates directly in the light reactions, absorbs violet-blue light, reflects green light) Chlorophyll b: accessory pigment (works in conjunction with chlorophyll a) Carotenoids: accessory light harvesting pigments (reflected light gives us the yellow, orange and red we see in the fall Photoprotection: Carotenoids protect the cells from damage due to excessive light energy Absorbs and dissipate light energy that would otherwise damage chlorophyll or form dangerous reactive oxidative molecules We eat carotenoids in our diet - Phytochemicals - Can have antioxidant properties. Excitation of chlorophyll by light: When a molecule absorbs a photon, one of the molecules electrons is elevated to an orbital where it has more potential energy - Photons energy matches the diFerence between the ground state and the excited state. Ground state: electron is in its normal orbital Excited state: electron is in an orbital of higher energy - Excited electrons drop back down to their ground state (releases the energy as heat and sometimes light, fluorescence) Photosystems: Reaction centre (RC): proteins holding a special pair of chlorophyll a molecules Light harvesting complex (LHC): pigment molecules bound to proteins Energy is passed from pigment molecules to pigment molecules within the LHC, and finally to the RC chlorophyll a molecules  Primary electron acceptor: molecule capable of accepting electrons from the RC chlorophyll a molecules 2 photosystems cooperate in the light reactions Photosystem II (PSII) functions first - RC chlorophyll a is known as P680 Photosystem I (PSI) functions second - RC chlorophyll is P700 Linear electron flow: (energy from absorbed photons is used to oxidise water on the luminal face of photosystems II (PSII) – electrons generated by this process pass through a series of electron carriers in PSII and then to the oxidized plastoquinone (PQ) that diFuse within the membrane.) Cyclic electron flow:  Uses photosystem I without using photosystem II Electrons cycle back from ferredoxin (Fd) to the cytochrome complex and then to a P700 chlorophyll No production of NADPH and no release of oxygen Production of ATP occurs Photosynthetic bacteria such as purple sulphur bacteria have only one photosystem - Hypothesis: these bacterial groups are descendants of ancestral bacteria in which photosynthesis first evolved. Cyanobacteria and plants have both photosystems and can carry out cyclic electron flow in certain conditions - Evolutionary remnant - Plants without cyclic electron flow can grow well in low light but not intense light - Hypothesis: cyclic electron flow may be photoprotective Chemiosmosis: ATP synthase couples of the diFusion of hydrogen ions down their gradient for the phosphorylation of ADP to form ATP - H+ diFuse from the thylakoid space to the stroma The Calvin cycle: Cyclic cycle Uses the chemical energy of ATP and the electrons carried by NADPH to reduce CO2 to sugar Anabolic pathway (building carbohydrates from smaller molecules and consuming energy) Output: glyceraldehyde-3-phosphate (G3P) - 3-carbon sugar - Cycle turns 3 times to make one G3P - Each turn of the cycle fixes one molecule of CO2 The Calvin cycle phase 1: Carbon fixation: incorporation of CO2 by attaching it to ribulose bisphosphate (RuBP) Enzyme: Rubisco - Most abundant protein in chloroplasts - Hypothesised to be the most abundant protein on earth Forms an unstable 6-carbon molecule that immediately breaks into 3-carbon molecules (3-phosphoglycerate) The Calvin cycle phase 2: Reduction: electrons from NADPH reduces 3-phosphoglycerate to glyceraldehyde 3-phosphate (G3P) For every three turns of the cycle, 6 G3P are formed - Only one is counted as a net gain - Other five are required to complete the cycle The Calvin cycle phase 3: Regeneration of RUBP: - 5 molecules of G3P are rearranged into 3 molecules of RuBP - Input of 3 ATP Calvin cycle input and output: Input - 9 ATP - 6 NADPH - 3 CO2  Output - 1 Glyceraldehyde 3-phosphate which can be used to synthesize other organic compounds such as glucose Photosynthesis is an emergent property of chloroplast which integrate with light and dark reactions to make sugars from inorganic carbon Topic 9 – DNA replication and cell division Nucleic acid structure and DNA replication Nucleic acids: - Gene: unit of inheritance (instructions to build a polypeptide) - Nucleic acid: polymer made of monomers called nucleotides - Deoxyribonucleic acid (DNA): genetic material organisms inherit from their parents. Provides directions for its own replication. Directs RNA synthesis - Ribonucleic acid (RNA): controls protein synthesis by interacting with ribosomes Polynucleotides: polymers that make nucleic acids Nucleotide: monomers of polynucleotides Nitrogenous base Five-carbon sugar Phosphate group Nucleoside: portion of the nucleotide that does not contain a phosphate group Nucleotides: Sugar (deoxyribose has one fewer oxygen atom than ribose Phosphate group attached to the 5’ carbon Nitrogenous base attached to the 3’ carbon Nucleotide or nucleoside monophosphate Nitrogenous base Pyrimidine: one 6-membered ring - Cytosine ©, thymine (T), uracil is only in RNA Purine: two rings, one 5-membered and one 6-membered  - Guanine (G), adenine (A) Nucleotide polymers: Nucleotides are linked together by a dehydration reaction - Forms a phosphodiester linkage - Sugar phosphate backbone: repeating pattern of sugars and phosphates 5’ end has a phosphate 3’ end has a hydroxyl group - Nitrogenous bases stick out from the sugar-phosphate backbone Structure of DNA: Two polynucleotide strands wound into a double helix - Strands held together by hydrogen bonds between paired nitrogenous bases Antiparallel: sugar phosphate backbones run in opposite directions from each other Complementary base pairing - A pair with T - G pairs with C Structure of RNA: Single strand of nucleic acid Complementary base pairing between regions of the same or diFerent molecules A pair with U Variable in shape and versatile functions - May have preceded DNA as the carrier of genetic information Discovering the structure of DNA: Maurice Wilkins and Rosalind Franklin performed x-ray crystallography on DNA molecules James Watson and Francis crick deciphered the data to determine that DNA is a double helix Structure of DNA: Data showed the width of the helix and the spacing of nitrogenous bases - One turn of the helix every 3.4nm - Bases are stacked 0.34nm apart - 10 base pairs per full turn of the helix Franklin suggested the sugar-phosphate backbone was on the outside of the molecule - Negative phosphates in contact with aqueous solution - Relatively hydrophobic bases hidden inside the molecule Purine must pair with pyrimidine for the helix to have a uniform diameter DNA replication: One strand act as a template for the synthesis of a second strand Nucleotides line up along the template strand and are linked to form a new strand Semiconservative model: each daughter molecule will have one parental strand, and one newly made strand Starting replication: Origin of replication: - Proteins that initiate DNA replication recognize and bind to this specific sequence of DNA - Eukaryotic chromosomes can have hundreds or thousands of origins of replication Two strands are separated to open a replication bubble Replication fork: Y-shaped region where the parental strands of DNA are being unwound Replication proceeds in both directions Replication bubble form and fuse together Enzymes: Helicase: enzyme that untwists the double helix at the replication forks  Single-strand binding proteins (SSB): binds the unpaired DNA molecules to prevent them from re-pairing Topoisomerase: enzyme that relieves the strain of tighter twisting at the replication fork (breaks, swivels, rejoins the DNA strand) Primase: synthesizes a short RNA primer Initiating synthesis of the new DNA stand: Primer: short RNA molecule, 5-10 nucleotides long New DNA strand starts from the 3’ end of the RNA primer DNA polymerase: adds nucleotides to the 3’ end of a pre-existing chain - DNA polymerase III and DNA polymerase I play major roles in E. coli - Eukaryotes have at least 11 diFerent DNA polymerases DNA pol III adds DNA nucleotides to the RNA primer and then keeps adding nucleotides to the growing end of the new strand - 500 nucleotides per second in bacteria - 50 nucleotides per second in eukaryotes Antiparallel elongation: DNA polymerase III adds nucleotides in a 5’ > 3’ direction Leading strand: replication towards the replication fork - DNA polymerase III remains in the replication fork continuously adding nucleotides to the new strand as the fork progresses - Only one primer is required for DNA pol III to synthesize the entire strand Lagging strand: DNA pol III works away from the replication fork Synthesized discontinuously as a series of segments Okazaki fragments: segments of DNA synthesized on the lagging strand 1000- 2000 nucleotides long in E. Coli 100 – 200 nucleotides long in eukaryotes Each fragment needs its own RNA primer DNA polymerase I replaces the primer un the DNA nucleotides DNA ligase joins the Okazaki fragments together into a continuous DNA strand DNA replication complex: Proteins participating in DNA replication form one single large complex - DNA replication machine - Primase acts as a brake to slow progress of the replication fork and coordinate placement of primers and rates of replication DNA moves through the complex Eukaryotes: copies of the complex may be anchored to the nuclear matrix Replicating the ends of DNA molecules: Normal DNA replication machinery cannot be complete the 5’ ends of linear daughter DNA strands  Repeated rounds of replication produce shorter and shorter DNA molecules with uneven (staggered) ends Telomers: special nucleotide sequences at the ends of DNA molecules Humans: TTAGGG is repeated 100-1000 times Telomeres: Proteins associated with telomeric DNA prevent staggered ends from activating the cells systems from monitoring DNA damage Telomeric DNA acts as a buFer zone providing protection against gene shortening Telomeres become shorter during every round of replication 1990 Calvin Harley suggested the “telomere clock” Telomeres may be connected to the aging process of tissues or organisms Germ line cells – do not shorten the telomeres Premature aging syndromes – telomere get shorter faster Telomerase: enzyme that catalyzes the lengthening of telomeres in eukaryotic germ cells Restores original length Compensate for the shortening that occurs in DNA replication Uses an RNA molecule to act as a template to extend the leading strand so that the lagging strand can also grow and maintain a given length Usually present in germ cells, not as common in somatic cells Telomeres will be at their maximum length in new zygotes Telomeres in cancer cells: Shortening telomeres may protect organisms from cancer Large tumors have cells that contain short telomeres Further shortening would lead to self-destruction of tumor cells Cancerous somatic cells often have high telomerase activity - Allows these cancer cells to persist - Capable of unlimited cell division - Same is seen in immortal cell lines  Could telomerase inhibition be used as a possible cancer therapy? Proofreading ad repairing DNA: One in 10(5) nucleotides are paired incorrectly Only one in 10(10) nucleotides remain incorrect DNA polymerase proofread each added to a growing strand Mismatch repair: enzymes remove and replace incorrectly paired nucleotides DNA becomes damaged all the time and maintenance of the molecule is important Mutations: permanent changes in the DNA sequence Repair mechanisms: almost 100 known in E. coli, about 170 known in humans Repair mechanisms: Nuclease: DNA-cutting enzyme Segment of the damaged strand is cut out (excised). Gap is filled in with nucleotides Undamaged strand is a template Gap is filled by a DNA polymerase and DNA ligase Ex. UV light (sun exposure) causes thymine dimers to form Cell division: the reproduction of cells - A parent cell replicates its DNA and distributes the copies to two daughter cells Asexual reproduction: division gives rise to a full new organism Multicellular eukaryotes undergo cell division - Development from a single cell zygote to a full organism - Renewal and repair of cells within the multicellular organism Genetic material: Genome: all the DNA in a cell - Prokaryotes: one single DNA molecule - Eukaryotes: many DNA molecules Chromosomes: packaged structures of DNA - Each chromosome carries hundreds or thousands of genes - Gene: a single unit of information that specifies an organism’s inherited traits Chromatin: the full complex of DNA and proteins  - Maintains the structure of the DNA - Controls the activity of the genes Number of chromosomes: Each eukaryotic species has a specific number of chromosomes in each cell Somatic cell: all body cells except gametes Humans have 46 chromosomes (2 sets of 23 chromosomes) Gametes: reproductive cells (eggs and sperm) Humans have 23 chromosomes (1 set) Chromosomes: Sister chromatids: duplicate copies of the chromosomes Cohesions: protein complexes that binds sister chromatids together Centromere: region where the sister chromatids are most closely attached to one another (proteins bind the centromeric DNA) Arm: portion of the chromatid to either side of the centromere Mitosis and cytokinesis: Mitosis – division of the genetic material in the nucleus Cytokinesis – division of cytoplasm (one cell becomes 2) Meiosis – production of daughter cells that have half the number of chromosomes as the parent cell - Production of gametes - Occurs in testes and ovaries in humans - Fertilization of egg with sperm cell returns the chromosomes number to 46 Cell cycle: the life of a cell from the time it is first formed during division of a parent cell into two new daughter cells to the time it divides itself into a new daughter cell Two main stages: 1. Interphase: growing phase 2. Mitotic phase: cell division Interphase: 90% of the cell cycle Metabolic activity is high as the cell performs its normal functions The cell - Grows - makes more cytoplasm - makes more proteins - makes more copies of organelles - replicates the DNA 3 phases of interphase: G1 phase – first gap (normal cellular functions) S phase – synthesis phase (DNA replication) G2 phase – second phase (normal cellular functions, the cell finishes preparing for cell division) Mitotic phase (M phase) Cell division produces two identical daughter cells 2 stages: 1. Mitosis – the nucleus and its contents are divided into two new daughter nuclei 2. Cytokinesis – the cytoplasm divided into two and the daughter cells are separated Cell cycle control system: G1 checkpoint – ensures the environment is correct for growth G2 checkpoint – ensures DNA is undamaged and fully replicated M checkpoint – ensures duplicated chromosomes are properly attached to the mitotic spindle Growth factors: protein released by certain cells that stimulates other cells to divide Essential nutrients: nutrients needed for a cell to make new macromolecules and organelles and energy for replication Density-dependant inhibition: crowded cells stop dividing (cells form a single layer and then stop growing) Anchorage dependence: cells must be attached to a substratum to divide Phases of mitosis: 1. G2 of interphase: Nuclear envelope encloses the nucleus Duplication of the centrosome Centrosomes – organize the spindle microtubules Centrosomes consist of two centrioles Chromosomes are duplicated DiFuse mass inside the nucleus 2. Prophase: Chromatin becomes tightly coiled Condenses into discrete chromosomes Nucleoli disappears Each duplicated chromosome appears as two sister chromatids joined at their centromeres Mitotic spindle forms Asters: microtubules that extend from the centrosomes Centrosomes move away from each other Propelled by the lengthening microtubules between them 3. Prometaphase: Nuclear envelope fragments Microtubules extend into the nuclear region Kinetochores form Specialized protein structure at the centromere of each chromatid Microtubules attach to kinetochores Microtubules interact with those from the opposite pole of the spindle 4. Metaphase: Centrosomes are at opposite poles Chromosomes are aligned at the metaphase plate Microtubules from opposite poles are in a tug of war for the sister chromatids Kinetochores of sister chromatids are attached to microtubules originating from opposite spindle poles Non- kinetochore microtubules from opposite poles overlap each other 5. Anaphase: Cohesion molecules are cleaved by the enzyme separase Sister chromatids separate becoming distinct chromosomes Chromosomes move toward opposite ends of the cell as kinetochore microtubules shorten - Motor proteins attached to kinetochores “walk” the chromosomes along the microtubule - Chromosomes are “reeled in” by motor proteins at the spindle poles The cell elongates as non-kinetochore microtubules lengthen - Region of overlap between microtubules is reduced as motor proteins walk them away from one another 6. Telophase: Two daughter cells nuclei form - Nuclear envelopes arise from the fragments of the parent cells nuclear envelope and other parts of the endomembrane system Nucleoli reappear  Chromosomes become less condensed Spindle microtubules are disassembled (depolymerize) Mitosis is complete Cytokinesis occurs at the same time Cytokinesis in animal cells: Cleavage Cleavage furrow: shallow groove in the cell surface near the old metaphase plate - Contractile ring of actin microfilaments - Actin interacts with myosin causing the ring to contract Cleavage furrow deepens until the parent cell is pinched in two Cytokinesis in plant cells: Vesicles carrying cell wall material move to the middle of the cell where they coalesce - Makes a cell plate as the cell wall material collect Cell plate enlarges - Membrane fuses with the plasma membrane at the perimeter of the cell - Two daughter cells each with its own plasma membrane Cell wall forms between the daughter cells Binary fission: Division in half Cell division done by prokaryotes DNA replication The cell elongates The plasma membrane pinches inward The parent cell is divided into two daughter cells with complete genome No mitotic spindle, no microtubules, proteins related to actin and tubulin may be involved.