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Biology Midterm 2 – Study Notes Latest Update.
Chapter 5 – Cell Membranes and Signalling 5.3 Membrane Proteins 5.3 a They Key Functions of Membrane Proteins
- Membrane proteins separate into four functional categories - Transport o Substances cannot freely diffuse through membrane o A protein provides a hydrophilic channel – allows movement of molecule o Membrane protein changes shape – shuttle molecules from one side of membrane to other - Enzymatic Activity o Enzymes = membrane proteins o Enzymes associated with respiratory & photosynthetic electron transport chains - Signal Transduction o Membranes contain receptor proteins on outer surface – bind to chemicals o Binding – receptors trigger changes on inside surface of membrane – lead to transduction of signal through the cell - Attachment/Recognition o Proteins exposed to internal & external membrane surfaces – attachment points for cytoskeleton elements & components in cell-cell recognition 5.3 b Integral Membrane Proteins Interact with the Membrane Hydrophobic Core - Integral membrane proteins – proteins embedded in phospholipid bilayer o Traverse entire lipid bilayer at least once – transmembrane proteins - Transmembrane proteins o Interact with aqueous environment on both sides of membrane & hydrophobic core o Have distinct regions that differ in polarity – domains o Domain – interacts with lipid bilayer ▪ Consists of nonpolar amino acids – form secondary structure (alpha helix) o Transmembrane proteins exposed on each side of membrane are composed of polar amino acids - Given amino acid sequence – simple to determine if it is a transmembrane protein o To look for ▪ Stretches of nonpolar amino acids ▪ Stretches – 17-20 amino acids in length
- Peptide length needed to span lipid bilayer o Transmembrane proteins span membrane more than once ▪ Ex. Protein = three membrane-spanning domains
- Pr imary sequence shows three regions of nonpolar amino acids linked by regions dominated by polar & charged amino acids
- Polar amino acids found in portions in proteins exposed to aqueous environment on each side of membrane **5.3 c Peripheral Membrane Proteins Interact with the Membrane Hydrophilic Surface
- Peripheral membrane proteins** – second group of membrane proteins o Positioned on surface of membrane o Do not interact w/ hydrophobic core of membrane o Held to membrane surfaces by noncovalent bonds – hydrogen bonds and ionic bonds o Found on cytoplasmic side of plasma membrane & form part of cytoskeleton o Made up of polar and nonpolar amino acids 5.4 Passive Membrane Transport - Hydrophobic nature of membranes restricts free movement of molecules - O2 diffuse rapidly across membranes – vital role in cellular respiration 5.4 a Passive Transport Is Based on Diffusion - Passive transport – movement of a substance across a membrane w/o chemical energy (ATP) - Diffusion – drives passive transport o Net movement of a substance from a high concentration to a low concentration o Above absolute zero (-273 degrees Celsius) molecules are in constant motion ▪ Molecules become uniformly distributed in space o Primary mechanism of solute movement within a cell o Driving force behind diffusion – increase in entropy - Initial state – molecules are more concentrated in one region/one side of membrane o Energy is more localized - Diffusion occurs – entropy increases
▪ Specific for water – does not allow diffusion of ions such as protons ▪ 3D models show presence of positive charges in centre of channel that repel transport of proteins o Gated Channel – found in all eukaryotes ▪ Switch b/w open, closed, and intermediate states ▪ Critical to the movement of ions – sodium, potassium, calcium & chlorine ▪ Gates open/close by changes in voltage across membrane or binding signal molecules ▪ Opening/closing involves changes in proteins 3D shape ▪ Animals – voltage-gated ion channels used in nerve conduction & control of muscle conduction ▪ CFTR Cl-^ channel = defective in individuals w/ cystic fibrosis
- Carrier Proteins – form passageways through lipid bilayer o Each protein binds a single solute (sugar molecule) & transports it across lipid bilayer o Transfer called uniport transport o Transport step – protein undergoes conformational changes that move the solute binding site from one side of membrane to the other – transports solute ▪ Distinguishes carrier protein function from channel protein function o Transport proteins display high degree of substrate – similar to enzyme ▪ Allows cells & cellular compartments to control what gets in and out ▪ Transport proteins present in plasma membrane depend on type of cell & growth conditions o How to experimentally determine if a molecule is transported by facilitated diffusion and not simple diffusion? ▪ Facilitated Diffusion - Rate of movement is faster based on the chemical structure of the molecule being transported - Can be saturated the same as an enzyme – by substrate - Measure of rate of transport at increasing concentration differences - rate of transport of a molecule (the substrate) reaches a plateau that represents a state when all transporters are occupied by substrate ▪ Simple Diffusion - Membrane surface is the transporter thus rate of transport never reaches a plateau **5.4 d Osmosis Is the Passive Diffusion of Water
- Osmosis** – like solutes, water moves across membranes o Passive transport of water constantly occurs in living cells o Inward/outward movement of water by osmosis develops forces that cause cells to swell/shrink
o Formally defined as diffusion of water molecules across a permeable membrane from a solution of lower to higher solute o To take place – permeable membrane must allow water molecules to pass not solute o Occurs in cells b/c they contain a solution of proteins & other molecules retained in the cytoplasm by a membrane impermeable to them but permeable to water o Can occur by simple diffusion through lipid bilayer/facilitated by aquaporins
- Movement of water by osmosis is dictated by solute concentration - Solution surrounding a cell contains dissolved substances at lower concentrations – hypotonic to cell (hypo = under/below; tonos = tension/tone) o Hypotonic solution – water enters by osmosis & cell swells ▪ Animal cells – red blood cells – can swell to the point of bursting ▪ Plant cells – presence of cell wall prevents cells from bursting - Solution that surrounds a cell contains solute at higher concentrations than in cell – hypertonic (hyper = over/above) o Hypertonic solution – water leaves by osmosis ▪ Outward movement exceeds capacity of cells to replace lost water – animal & plant cells shrink - Animals, ions, proteins & other molecules are concentrated in extracellular fluid and inside cells – concentration of water inside & outside cells is equal/isotonic (iso = same) o Comes at energetic cost of constantly pumping ions from one side to the other o Ex. ATP-dependent transport of Na+^ from inside to outside the cell is essential – otherwise water would move inward by osmosis & cells would burst 5.5 Active Membrane Transport - Facilitated diffusion compared to simple diffusion increases rate of movement of molecules across membranes o Type of transport is limited to movement down a concentration gradient 5.5 a Active Transport Requires Energy - Active Transport o Transport of molecules across a membrane against a concentration gradient that requires energy o Energy is in the form of ATP – estimated about 25% of a cells ATP requirement o Concentrates molecules sugars & amino acids inside cells & pushes ions in/out of cells - Three main functions of active transport in cells and organelles o Uptake of nutrients from fluid surrounding cells even when concentrations are lower than in cells o Removal of secretory/waste materials from cells/organelles when concentration is higher outside
o Voltage across plasma membrane results from difference in charge and from unequal distribution of ions across membrane created by passive transport o Membrane potential – measures -50 to -200 millivolts ▪ Minus sign indicates charge inside cell is negative versus outside o Both a concentration difference and an electrical charge difference on the two sides of membrane – electrochemical gradient o Electrochemical gradients – store energy that is used for other transport mechanisms ▪ Ex. Electrochemical gradient across membrane is the movement of ions associated with nerve impulse transmission 5.5 c Secondary Active Transport Moves Both Ions and Organic Molecules
- Secondary active transport pumps use concentration gradients of an ion established by a primary pump as their energy source o Ex. Driving force for secondary active transport in animal cells is the high outside/low inside Na+^ gradient set up by the sodium-potassium pump o Transfer of solute across membrane is couples with transfer of ion supplying the driving force o Occurs by two mechanisms – symport and antiport - Symport o Co-transported solute moves through membrane channel in same direction as driving force – cotransport o Ex. Glucose and amino acids - Antiport o Driving ion moves through membrane channel in one direction – provides energy for active transport of another molecule in opposite direction – exchange diffusion o Ions are exchanged by antiport o Ex. Is the mechanism used in red blood cells for the movement of chloride ions & bicarbonate ions through a membrane channel 5.6 Exocytosis and Endocytosis - Eukaryotic cells import & export larger molecules by endocytosis and excytosis - Export of materials by exocytosis carries secretory proteins & waste materials from cytoplasm to cell exterior - Import by endocytosis carry proteins, larger aggregates of molecules/whole cells from outside into cytoplasm - Exocytosis & endocytosis contribute to back-and-forth flow of membranes b/w endomembrane system and plasma membrane - Both require energy – both processes stop if a cell’s ability to make ATP is inhibited
5.6 a Exocytosis Releases Molecules to the Outside by Means of Secretory Vesicles
- Exocytosis o Secretory vesicles move through cytoplasm & contact plasma membrane o Vesicle membrane fuses with plasma membrane – releases vesicle’s contents to cell exterior o Eukaryotic cells secrete materials to outside through exocytosis ▪ In animals, glandular cells secrete peptide hormones/milk proteins and cells lining digestive tract secrete mucus and digestive enzymes ▪ Plant cells secrete carbohydrates by exocytosis to build a strong cell wall **5.6 b Endocytosis Brings Materials into Cells in Endocytic Vesicles
- Endocytosis** o Proteins & other substances are trapped in pit like depressions that bulge inward from plasma membrane ▪ Depression then pinches off as endocytic vesicle o Takes place in eukaryotic cells by one of two distinct but related pathways - Bulk-phase endocytosis/pinocytosis o Extracellular water is taken in along with any molecules that happen to be in the solution in the water o No binding by surface receptors takes place - Receptor-mediated endocytosis o Molecules to be taken in are bound to the outer cell surface by receptor proteins o Receptors – integral proteins of the plasma ▪ Recognize & bind certain molecules from solution surrounding cell o After binding target molecules – receptors collect into a depression called coated pit b/c of network of proteins (clathrin) that coat & reinforce the cytoplasmic side o W/ target molecules attached – pits deepen & pinch free of plasma membrane to form endocytic vesicles o When in cytoplasm – endocytic vesicle loses its clathrin coat & fuse with a lysosome o Enzymes within lysosome digest contents of vesicle – breaking down into smaller molecules useful to cell o Molecular products enter cytoplasm by crossing vesicle membrane via transport proteins - Cells – white blood cells (phagocytes) – in bloodstream/protists take in large aggregates molecules, cell parts/whole cells by a process called phagocytosis - Phagocytosis – cell eating
6.1 b Coupled Oxidation-Reduction Reactions Are Central to Energy Metabolism
- Potential energy in fuel molecules – released when molecules lose electrons = oxidation
- Electrons released from molecule that is oxidized = gained by another molecule = reduced
- Oxidation & Reduction rxns = coupled processes; cannot happen without the other o Oxidation = loss of electrons o Reduction = gain of electrons - Oxidation-Reduction rxns = redox rxns
- General Redox Rxn : Oxidation Xe-^ + Y X + Ye- Reduction The redox rxn describing respiratory breakdown of glucose is as follows: Oxidation C 6 H 120 6. + 602 6CO 2 + 6H 2 O Reduction
- Oxidation comes from the rxns (where electrons are removed from fuel molecules) involve O 2 as atom accepts electrons & gets reduced
- Involvement of O 2 = essential for common oxidation rxns o Car engine requires large amounts of air (21% O 2 ) to be delivered to each piston for combustion to take place
- Concept of redox rns = more challenging to understand by two facts o First – although oxidation rxns involve O 2 , others (including cellular respiration) do not o Second – the gain/loss of electron in redox rxn = not always complete
▪ In some rxns – electrons are transferred from one atom to another ▪ In other rxns – degree changes to which electrons are shared b/w 2 atoms
- Rxn b/w methane & oxygen = redox rxn, where degree of electrons sharing changes
- Figure 6.
- Blue dots = position of electrons in covalent bonds of reactants & products
- Reactant - methane o Electrons = shared equally b/w carbon & hydrogen atoms
- Product – carbon dioxide o Electrons = closer to oxygen than carbon b/c oxygen atoms = more electronegative
- Carbon atom – has partially lost shared electrons in rxn; methane = oxidized
- Reactant – oxygen / Product - water o Two oxygen atoms share electrons equally o Oxygen reacts w/ hydrogen from methane – producing water, where electrons = closer to oxygen atom than hydrogen atom o Means each oxygen atom has partially gained electrons; oxygen = reduced o B/c of this – rxn b/w methane & oxygen releases heat o Energy = released as electrons in C-H bonds of methane move closer to electronegative oxygen atoms that form carbon dioxide 6.1 c Cellular Respiration Is Controlled Combustion - Like gasoline & methane – glucose can undergo combustion & burn - Combustion of glucose o Releases energy as electrons are transferred to O 2 – reducing it to water & carbon in glucose = oxidized to carbon dioxide - Spontaneous rxn – substrate molecules need to reach transition site for it to proceed o Requires energy input: activation energy - To get glucose to ignite o Use a flame to provide high activation energy - Within a cell – oxidation of glucose occurs through a series of enzyme catalyzed rxns
- each w/ a small activation energy - Thermodynamically : the two processes = identical o Both exergonic = same energy in free energy (delta G) of -686kcl/mol o Difference: burn glucose = energy released as heat = not available to drive metabolic rxns - Process of cellular respiration = controlled combustion o Energy of C-H bonds – not liberated suddenly, producing heat, but is released in a stepwise fashion, w/ energy transferred to other molecules - Cellular respiration – oxidation of food molecules occurs in presence of dehydrogenases
o Citric acid cycle & oxidative phosphorylation occur in specialized membrane- bound organelle called mitochondrion
- Mitochondrion : membrane bound organelle o Referred to as powerhouse of cell b/c location of citric acid cycle & oxidative phosphorylation = largest generator of ATP in cell o Composed of two membranes: outer & inner – define two compartments ▪ Intermembrane space – found b/w outer & inner membrane, matrix (interior aqueous environment) 6.3 Glycolysis: The Splitting of Glucose o 10 enzyme-catalyzed rxns that lead to oxidation of six-carbon sugar glucose, producing two molecules of three-carbon compound pyruvate o Potential energy released in oxidation leads to synthesis of NADH & ATP **6.3 a Glycolysis Is a Universal and Ancient Metabolic Process
- Glycolysis** = first metabolic pathway studied & best understood in terms of o Enzymes involved o mechanisms of action o how pathway is regulated to meet energy needs of cell - First experiments investigating glucose = 100 years ago o First to show one could study biological rxns in an isolated, cell-free system o Experiments became foundation of modern biotechnology - Glycolysis = fundamental & most ancient of all metabolic pathways - Supported by 3 facts o Glycolysis = universal ▪ Found in all 3 domains of life; Archaea, Bacteria & Eukarya o Glycolysis = not dependent on presence of O 2 ▪ Became abundant in Earth’s atmosphere 2.5 bill years ago – 1.5 bill years after scientists though life first evolved o Glycolysis occurs in cytosol of cells using soluble enzymes ▪ Thus, does not require sophisticated ETCs & internal membrane systems to function **6.3 b Glycolysis Includes Energy-Requiring and Energy-Releasing Steps
- Key features** o Energy investment followed by payoff o No carbon is lost o ATP is generated by substrate-level phosphorylation - Energy investment followed by payoff o Consists of two distinct phases ▪ Initial five-step energy-requiring phase followed by a five-step energy releasing phase o Initially – two molecules of ATP are consumed as glucose & fructose- 6- phosphate become phosphorylated
o Investment of two ATP for each glucose molecules leads to an energy reward ▪ Reward = four ATP & two NADH molecules are produced during energy- releasing phase
- No carbon is lost o Rxns of glycolysis convert glucose into two molecules of the three-carbon compound pyruvate = no carbon is lost o Since glucose was oxidized – potential energy in two molecules of pyruvate = less than one molecule of glucose - ATP is generated by substrate-level phosphorylation o During glycolysis – ATP = generated by pross of substrate-level phosphorylation o This mode of ATP synthesis involves transfer of phosphate group from a high- energy substrate molecule to ADP – producing ATP o Substrate-level phosphorylation = mediated by specific enzyme – also mode of ATP synthesis used in citric acid cycle 6.4 Pyruvate Oxidation and the Citric Acid Cycle - Two molecules of pyruvate synthesized by glycolysis contains usable free energy 6.4 a Pyruvate Oxidation Links Glycolysis and the Citric Acid Cycle - B/c rxns of citric acid cycle are localized to mitochondrial matrix – pyruvate synthesized during glycolysis must pass through both outer & inner mitochondrial membranes o Large pores in outer membrane – allow pyruvate to diffuse through o Inner membrane – crossing this requires a pyruvate-specific membrane carrier - Once pyruvate is in matrix – it is converted into acetyl-CoA through a process – pyruvate oxidation - Pyruvate oxidation o Starts with a decarboxylation rxn where the carboxyl group (-COO-) is lost as carbon dioxide ▪ Rxn = understandable b/c carboxyl group contains no usable energy o Followed by oxidation of remaining two=carbon molecule – producing acetate ▪ Acetate – dehydrogenation rxn leads to transfer of two electrons & proton to NAD+, forming NADH o Lastly, acetyl group reacts w/ coenzyme A (CoA) – forms high- energy intermediate acetyl-CoA - Goal of rxns that make up citric acid cycle = liberating electrons in acetyl-CoA that still contain three C-H bonds 6.4 b The Citric Acid Cycle Oxidizes Acetyl Groups to Carbon Dioxide - Citric acid cycle consists of eight enzyme-catalyzed reactions o Seven = soluble enzymes located in mitochondrial matrix o One enzyme is bound to matrix side of inner mitochondrial membrane ▪ Combined – rxns result in oxidation of acetyl groups to carbon dioxide with the synthesis of ATP, NADH and FAD; reduced = FADH 2
6.5 Oxidative Phosphorylation: Electron Transport and Chemiosmosis
- Following citric acid cycle – all carbon atoms originally in glucose are completely oxidized & released as carbon dioxide
- Besides ATP formed by substrate-level phosphorylation – the potential energy originally in glucose now exists in molecules of NADH & FADH 2
- Role of ETC coupled with process of chemiosmosis extract potential energy in these molecules & synthesize additional ATP 6.5 a The Electron Transport Chain Converts the Potential Energy in NADH and FADH 2 into a Proton-Motive Force - Respiratory ETC comprises a system of components that in eukaryotes is found in inner mitochondrial membrane o Chain facilitates transfer of electrons from NADH 2 & FADH 2 to oxygen - Chain consists of 4 protein complexes: o Complex I – NADH dehydrogenase o Complex II – succinate dehydrogenase o Complex III – cytochrome complex o Complex IV – cytochrome oxidase ▪ Complex II = single peripheral membrane protein ▪ Remaining complexes are composed of multiple proteins ▪ Ex. 40 individual proteins make up complex I - Electron flow b/w complexes = facilitated by two mobile electron shuttles o Ubiquinone – hydrophobic molecule in core of membrane ▪ Shuttles electrons from complexes I & II to complex III o Second shuttle – cytochrome c = located on intermembrane space side of membrane ▪ Transfers electrons from complex III to complex IV – cytochrome oxidase 6.5 b Electrons Move Spontaneously along the Electron Transport Chain
- In ETC, not proteins themselves that transfer electrons – rather electron transported is facilitated by nonprotein molecules – called prosthetic groups
- Protein subunits of each complex I, III & IV bind a number of prosthetic groups very precisely o Allows for electron transport
- Prosthetic groups = redox-active cofactors that alternate b/w reduced & oxidized states as they accept electrons from upstream molecules & donate electrons to downstream molecules o Common prosthetic group = molecule heme, component of cytochromes, including cytochrome c ▪ Heme = component of many biologically important components – including hemoglobin, where it is critical to molecule’s ability to carry O 2 o Heme group ▪ Contains central redox-active iron atom that alternates b/w Fe2+^ & Fe3+
- During electron transport – one of the prosthetic groups of complex I FMN, is reduced by electron donation from NADH on matrix side of inner membrane o FMN donates electron to Fe/S prosthetic group – in turn donates electron to ubiquinone
- Process of reduction followed by oxidation of each carrier continues along entire chain until electrons are donated to oxygen – reducing it to water o Protons used in formation of water = abundant in aqueous environment of cell
- Prosthetic groups & other electron carriers = organized in a specific way o From high to low free energy
- NADH = high potential energy b/c it contains high-energy electrons, thus can be readily oxidized o By contrast, O 2 (terminal electron acceptor of chain) = strongly electronegative & can be reduced
- As a consequence – electron movement along chain = thermodynamically spontaneous – down a free energy gradient 6.5 c Chemiosmosis Powers ATP Synthesis by a Proton Gradient - Although goal of cellular respiration = synthesis of ATP, electron transport from NADH/FADH 2 to O 2 – does not actually produce ATP o Electrons are passed along a chain of electron carriers until they are donated to oxygen, producing water - How ATP is produced o NADH = more potential energy than O 2 - Energy release in electron transport chain o Energy released during electron transport is used to do work o Specifically work of transporting protons across inner mitochondrial membrane from matrix to intermembrane space ▪ As a consequence of this proton pumping across inner membrane, H+ concentration becomes higher in intermembrane space than in matrix - Proton translocation occurs at distinct sites along ETC o Within complexes I & IV – specific protein components use energy released from electron transport for proton pumping o As ubiquinone molecules accept electrons from complexes I & II – they pick up protons from matrix o After migrating through membrane & donating electrons to complex III – ubiquinone retains a neutral charge by releasing protons into intermembrane space
- Spinning of headpiece of ATP synthase represents the smallest molecular rotary motor known in nature - Active transport pumps o Use energy from ATP to transport ions across membranes against concentration gradients o An ATP synthase operating in reverse o Doesn’t synthesize ATP – uses free energy from hydrolysis of ATP to provide the energy necessary to pump ions across a membrane - Harnessing potential energy present in proton gradient to synthesize ATP = fundamental to all forms of life & developed early in evolution of life o Shown by fact that ATP synthase complex found in mitochondria is structurally similar to ATP synthase complexes found in thylakoid membrane of chloroplast & plasma membrane of bacteria & archaea 6.5 e Electron Transport and Chemiosmosis Can Be Uncoupled - Generation of ATP by ATP synthase = linked/coupled to electron transport by proton gradient established across inner mitochondrial membrane - Electron transport & chemiosmotic generation of ATP = separate & distinct processes that are not always completely coupled - Example o Possible to have high rates of electron transport & yet no ATP generated by chemiosmosis o Uncoupling of two processes occur when mechanisms prevent formation of a proton-motive force - Class of chemicals – ionophores o Form channels across membranes through which ions, including protons can freely pass o Consequence – in presence of ionophores ▪ Proton pumping during electron transport is followed by protons flowing back unto matrix through ionophore channels ▪ Proton gradient = prevented from becoming established - Ionophores – referred to as uncouplers o Very toxic b/c of their ability to inhibit oxidative phosphorylation o In 1930s – low concentrations of chemical uncouplers were used as diet drugs ▪ Although people lost weight – overdoses resulting in death not uncommon - When electron transport = uncoupled from chemiosmotic synthase of ATP – free energy released during electron transport is not conserved by a proton-motive force – instead lost as heat o Organisms take advantage of this as a means of regulating body temperature by altering expression of a group of transmembrane proteins o These uncoupling proteins = localized to inner mitochondrial membrane & similar to chemical uncouplers, form channels through which protons can freely flow o This mechanism of regulating body temp = important in animals o Example
▪ Hibernating mammals & newborn infants – activity of uncoupling within mitochondria of brown adipose fat = important mechanism of heat generation 6.6 The Efficiency and Regulation of Cellular Respiration
- Calculate efficiency w/ which cellular respiration extracts energy from glucose 6.6 a What Are the ATP Yield and Efficiency of Cellular Respiration?
- ATP molecules produced by oxidative phosphorylation o Recent research suggests that for each NADH that is oxidized – for each pair of electrons that travels down ETC – 10 H+^ are pumped into inner membrane space o B/w three and four H+^ are needed to flow back through ATP synthase for synthesis of one molecule of ATP ▪ Gives b/w 2.5 & 3.3 molecules of ATP synthesized for every NADH oxidized by ETC ▪ For each NADH oxidized – three ATP are synthesized ▪ B/c oxidation of FADH 2 skips proton-pumping complex I – only about two molecules of ATP are synthesized for each FADH 2 oxidized
- Detailed accounting of ATP yield for each molecule of glucose oxidized (Figure 6.18)
- Products of glycolysis include two molecules of ATP & two molecules of NADH
