Metabolism (Cellular respiration + enzymes)
What are enzymes?
Globular proteins that increase the rate of a biochemical reaction by lowering the activation energy threshold without being consumed in the process.
Purpose of enzymes
They allow for reactants in a reaction to be converted into products faster, provide the force needed for collisions to occur, and they lower they activation energy needed in a reaction
Active sites
The position on the enzyme occupied by the substrate, it is on the surface of the enzyme and once the substrate binds it catalyses the reaction. The active site is often composed of open loops of polar amino acids on the exterior or the enzyme molecule (R-side chain).
What is metabolism?
the complex network of reactions that occur in living organisms. They form metabolic pathways in which one type of molecule is transformed into another via a series of small steps (chains or cycles), catalyzed by an enzyme.
Intracellular enzymes
Enzymes within a cell, examples of intracellular reactions are glycolisis (in the cytoplasm) and the Krebs cycle (matrix of mitochondria).
extracellular enzymes
enzymes outside of a cell. Examples of extracellular reactions include chemical digestion within the digestive system.
Multienzyme complexes
groups of enzymes that work together
Enzyme specificity
Each enzyme catalyzes one specific reaction, or a specific group of reactions. Enzymes are substrate specific, meaning they will only work with one particular reactant (e.g enzymes for breaking down proteins will not break down starch). This results in large numbers of different enzymes in organisms, and also allows for them to control metabolism and the rate of reaction.
Anabolism
the synthesis of complex molecules from simpler molecules (including macromolecules from monomers via condensation). ATP is needed. Examples of anabolic reactions are protein synthesis, DNA replication, and photosynthesis
Catabolism
The breakdown of complex molecules into simpler molecules (e.g hydrolysis). Energy is released and is sometimes captured as ATP. Catabolic reactions include digestion and cell respiration.
Amino acids in the active site
The chemical properties of the active site and the substrate match each other, these conditions created by a few amino acids that change the substrates enough to convert them into products. If they are not next to each other in the polypeptides that make up the enzyme, they are brought together by the folding of the polypeptide. If the structure is altered, catalysis is unlikely to happen
Lock and Key hypothesis
An outdated model. The substrate and the active site match eachother structurally and chemically, meaning the substrate will not be able to react if not chemically attracted to the active site, or if it cannot fit into its specific 3D structure. (Enzymes only work for one substrate)
Induced fit model
The proper and updated model. This model describes enzymes as dynamic protein molecules that will change shape to better accomodate the substrate. Some enzymes can catalyze multiple reactions. As the substrate approaches, it induces a conformational change in the active site, this stresses the substrate, reducing the activation energy of the reaction. This attachment creates the enzyme-substrate complex.
What is a collision?
the coming together of a substrate and an active site
Enzymes and molecular motion
Most enzyme reactions occur when the substrates are dissolved in water. These molecules are in random motion, each moving seperately. If more substrates or enzymes molecules are added, the collision rate will increase (unless all active sites/substrates are being used when more are added). If not stuck to the surface of something and immobilized, enzymes can move (however they tend to be larger and move slower). If the substrate is too large and doesn't move much, the enzyme has to move in relation to the substrate.
Denaturation
Most of the bonds that stabilize the formation of proteins (amino acids) are relatively weak and can be disrupted or broken. This will result in the change of the conformation of the protein, which is called denaturation. Once the structure is changed, the protein can no longer perform its functions, and the changes are often permanent. Enzyme activity is therefor affected by extreme temperature and pH.
Effects of temperature on enzymes
Low temperatures result in insufficient thermal energy for the activation of enzyme catalyzed reactions to occur. However increased temperature will increase the motion + speed of both the enzyme and substrate, increasing their activity. Each enzyme has an optimal temperature that enzyme activity will peak at. Extreme heat can cause denaturation through vibrations within the molecule that break intermolecular bonds or reactions, changing the structure so that the substrate can no longer bind.
affects of pH on enzymes
At the optimal pH, the maximum rate of reaction is achieved. Above and bekiw that pH, the rate decreases. This is because bonds are made and broken which change the shape of the active site. Extremes of pH can cayse denaturation.
Calculating rates of reaction in enyzmes
There are two approaches:
1. Allow the reaction to happen for a fixed time and measure the amount of substrate used up or product formed.
2. Start with a known amount of substrate and allow the reaction to continue until all the substrate has been converted into products. Measure the time taken for the reaction to go to completion.
Rate of reaction is usually measured by amount of product (cubic cm) divided by time.
Enzymes and activation energy
Substrates have to pass through a transition state before they are converted into products. Energy is required to reach this transition state. it is called activation energy and is used to break bonds in substrate molecules. Energy is also released as new bonds are made and product is formed. On an enzyme, the reaction is typically exothermic (output energy is more than input energy). Endothermic is the opposite.
Non-competitive inhibition
Inhibitors fit into the allosteric site (other site on enzyme), causing a conformation change in the active site, keeping the substrate from attaching. This is usually to regulate or slow down the process. As concentration of inhibitor increases, the rate of reaction decreases as there are fewer functional active sites available for reaction.
Competitive inhibition
The inhibitor fits into the active site and prevents the substrate from entering. The higher the concentration of inhibitor, the slower the rate of reaction. The maximum rate of reaction will still be achieved, however, if more substrate is added as the amount of enzymes available has not changed.
Statins
medicines that work by competitive inhibition. They are used to treat high blood cholesterol, which can cause heart disease. They bind to the active site of the enzyme HMG-CoA reductase (catalyzes a reaction in the synthesis of cholesterol in liver cells), this lowers the rate and the body produces less cholesterol.
ACE inhibitors
the RAA system causes vasoconstriction (tightening of blood vessels) when blood pressure drops. In people with hypertension or heart failure, the action of angiotensin II can make their problem worse. ACE inhibitors are medications that inhibit angiotensin converting enzymes (ACE), they are non-competitive and reversible. They help to prevent increased blood pressure.
End-product Inhibition
the product of the last reaction of a metabolic pathway acts as non-competitive inhibitor of the enzyme involved in the reaction at the very beginning of the pathway. When enough product is available, its synthesis is turned off. When that end-product gets used by the cell, the first enzyme is reactivated (allosteric enzyme). When concentrations are too high, this end product binds to the first enzyme to inhibit it and will bind less often when concentrations are low.
Isoleucine
Example of feedback inhibition. Isooleucine is an essential amino acid (comes from food, not made by body). Bacteria synthesize it from threonine in a series of five enzyme catalyzed steps. As the concentration of isoleucine increases, some of it binds to the allosteric site of threonin deaminase (first enzyme). As the concentration of isoleucine falls, the allosteric sites of threonine deaminase are emptied and allow it to carry out its functions.
Penicilin
Peniclin is an antibiotic discovered by alexander fleming. Dorthy Hodgkin used X-ray crystallography to determine the molecular shape of penicillin. All penicilins have a beta-lactam ring however they can differ based on the side chains. In bacteria, transpeptide is an enzyme that helps catalyze the formation of cell walls. The beta-lactam ring in penicillin binds irreversibly to transpeptidase and acts as an inhibitor, leaving the bacteria with weak cell walls that eventually burst.
Penicillin resistant bacteria
many bacteria have become enzyme resistant to penicilin due to a mutation that causes an enzyme to be produced called penicillinase. This breaks bonds within penicillin and as a result it can no longer bind to transpeptidase. Over time, these bacteria became dominant because they survived and reproduced while the non-resistant bacteria died off. Changing side chains of penicillin prevents resistance. Another form of resistance occurs due to mutations in the transpeptidase gene. The mutations can cause a structural change to the active site, and as a result, penicillin will no longer be able to bind to the active site.
Energy released by carbon compunds in ATP
all living cells need a continuous supply of energy to carry out various processes such as active transport and protein synthesis. Cell respiration is the controlled release of energy. Each covalent bond ina glucose, amino acid or fatty acid represents stored chemical energy, when they are broken, energy is released and used to make ATP (adenosine triphosphate).
What is the ATP molecule?
Adenosine triphosphate consists of a 5-carbon sugar (ribose) which is bonded to a nitrogenous base and 3 phosphate groups. ATP is a nucleotide. The bond connecting the phosphates are considered high energy bonds because the phosphates are negatively charged and repel one another. These bonds can be broken by a hydrolysis reaction, releasing energy in the process.
ATP becoming ADP
When ATP splits up to become ADP (adenosine diphosphate) and phosphate, most of the energy that is released is converted into heat energy (all energy ends up as heat eventually). This energy can be used to raise the temperature of the organism, but is eventually lost to the environment.
Creating ATP from ADP
The formation of ATP by adding a phosphate and hydrogen ion to ADP is an endergonic reaction (energy is absorbed and stored in bonds). This is created by the oxidation of nutrients (carbohydrates, proteins, etc). Adding a phosphate group is called phosphorylation. When energy is needed within the cell, ATP is converted to ADP, breaking the bond between one of the phosphates and releasing energy (exergonic reaction)
What is aerobic respiration?
Aerobic respiration needs the presence of oxygen to proceed. It occurs in cells that have mitochondria (happens in mitochondria). A large yield of ATP is given (36 per glucose). Water is a waste product recycled in the cell, and CO2 is excreted through gas exchange.
What is anaerobic respiration?
Occurs in the absence of oxygen. Carbon dioxide and ethanol are produced in yeast (plants + yeast: glucose --> ethanol + carbon dioxide + 2 ATP), and lactic acid is produced in in humans and other animals (animals: glucose --> lactate + 2 ATP). The overall yield of ATP is very low (2 ATP per glucose from glycolysis). Its useful for short, rapid bursts of ATP production, when oxygen supplies are low, and in low-oxygen environments.
Matrix (Mitochondria diagram)
contains enzymes and solutes for link reaction and krebs cycle. The compartment enclosed within the inner membrane. It is the gel like substance inside the membrane.
Outer mitochondrial membrane
Acts as a barrier to larger molecules (the membrane that borders the mitochondria).
Inner mitochondrial membrane
The site for electron tranpsort chain and oxidative phosphorylation. (the membrane encasing the matrix)
Cristae (mitochondria diagram)
To maximise surface area for reaction. Cristae are the folds of the inner membrane.
Small inter-membrane space
more efficient generation of H+ concentration gradient. This is the region between the outer and inner membranes of the mitochondria.
70S Ribosomes (mitchondria diagram)
Used for protein production, they are the dark dots inside the matrix.
Naked loops of DNA (mitochonria diagram)
Mitochondrial dna or mDNA. Passed unchanged from mother to child. These are the lighter dots within the matrix, are not often visible on an electron micrograph
What are oxidation and Reduction?
Many biochemical reactions are either classified as oxidation or reduction. Oxidation is the loss if electrons and hydrogen, but the gaining of an oxygen. Reduction is the opposite. OILRIG.
What is NAD?
NAD (nicotinamide adenine dinucleotide) is the most common hydrogen carrier. NAD+ + 2H+ + 2e- produces NADH + H+ via reduction and vice versa. (the simplified form of NAD is just NAD+)
What is FAD?
FAD (flavin adenine dinucleotide) is a less frequently used hydrogen carrier. FAD produces FADH2 via reduction and vice versa.
Glycolysis step 1
Hexokinase reaction: phosphorylation of hexoses (mainly glucose). ATP is dephosphorylated (removal of a phosphate). The phosphate group replaces the hydroxyl group of carbon 6 (glucose is phosphorylated - addition of a phosphate). This forms glucose 6-phosphate (G6P).
Glycolysis step 2
Phosphohexose isomerase: Isomerization of g6p to fructose 6 phosphate. Isomerases convert molecules from one isomer to another. This enzyme catalyzes the rebersible isomerization g6p to f6p.
Glycolysis step 3
Phosphofructokinase-1 reaction: transfer of phosphoryl group from ATP to corner 1 of F6P to produce fructose 1,6 bisphosphate. ATP is dephosphorylated. The phosphate group replaces the hydroxyl group on carbon 1 (F6P becomes phosphorylated). Fructose 1,6 bisphosphate is now formed.
Glycolysis step 4
Aldolase reaction: cleavage of fructose q,6 bisphosphate into glyceraldehyde 3 phosphate (an aldose) and dihydroxy acetone phosphate. The aldolase enzyme catalyses the cleavage of F1,6 Bisphosphate into DHAP and G3P.
Glycolysis step 5
Triose phosphate mutase reaction: conversion of DHAP to glyceraldehyde 3 phosphate. This isomerase reaction converts the DHAP to G3P, so there are now two G3Ps in total (the next 5 steps happen twice at the same time because there are two g3p molecules).
Glycolysis step 6
Glyceraldehyde-3-phosphate dehydrogenase reaction (GAPDH): conversion of g3p to bisphosphoglycerate. (BPG). NAD removes the hydrogen off of inorganic phosphate, becoming reduced to NADH (this will be used to make more ATP later). The phosphate group attaches itself to g3p, converting it to BPG.
Glycolysis step 7
Phosphoglycerate kinase reaction: transfer of phosphoryl group from BPG to ADP, generating ATP. BPG becomes dephosphorylized forming 3-phosphoglycerate (3PG). The phosphate group will attach itself to an ADP molecule forming ATP (substrate level phosphorylation).
Glycolysis step 8
phosphoglycerate muatase reaction: Conversion of 3-phosphoglycerate to 2-phosphoglycerate (2-PG). The phosphate group switches places with the hydroxyl group on carbon 2. This formes 2PG.
Glycolysis step 9
Enolase reaction: Dehydration of 2-PG to phosphoenolpyruvate (PEP). A dehydration reaction occurs where the hydrogen on carbon 2 and the hydroxyl group on carbon 3 form water, creating a double bond between carbon 2 and 3. PEP is then formed.
Glycolysis step 10
Pyruvate kinase reaction: Transfer of phosphoryl group from PEP to ADP generating ATP and pyruvate. PEP becomes dephosphorylated, forming 2 pyruvate molecules. ADP becomes phosphorylated and forms 2 ATP molecules.
Link reaction
This links glycolysis (which brings glucose outside of the cell into the cytoplasm) with the Krebs Cycle (which happens inside the mitochondria). Pyruvate enters the mitochondrion matrix by active transport. Enzymes remove one carbon dioxide and hydrogen from the pyruvate. (the link reaction is called oxidative decarboxylation). Hydrogen is accepted by NAD to form NADH. the product is an acetyl group that reacts with coenzyme A. Acetyl CoA enters the Krebs cycle.
Cell respiration using fatty acids
Fatty acids can also be a source of energy in respiration. Fatty acids have a long chain of carbon atoms. CoA can oxidize this chain (break it down). It makes Acetly CoA with two carbons and carries them to the Krebs cycle. Glycolysis is not needed in this case, but the reaction is slower. If there are an odd number of carbons, the remaining carbon atom is released as carbon dioxide. Generally, the fatty acid produces 20% more ATP than the glucose molecule.
What is the krebs cycle?
The krebs cycle reduces electron carriers in preparation for oxidative phosphorylation (carbon is released as CO2 as a by-product). The reduced forms of NAD and FAD (NADH + H and FADH2) carry H+ ions and electrons to the electron transport chain, which is situated in the folds on the inner mitochondrial membrane (the cristae).
Krebs Cycle process
Acetyl CoA enters the krebs cycle. Acetyl group (2 carbon) joins a 4 carbon sugar to form a 6 carbon sugar. Oxidative decarboxylation of the 6C sugar to a 5C compound produces CO2. This happens again, making the 5C compound into a 4C compound and producing CO2. The process is oxidative as NAD and FAD are reduced by the addition of hydrogen. Two CO2 are produced per molecule of pyruvate, along with three NADH + H+ and one FADH2. One ATP is produced by substrate level phosphorylation per molecule of pyruvate. NADH and FADH2 provide electrons to the electron transport chain.
Oxidative phosphorylation - ETC first steps
A series of integral protein complexes act as electron carriers, forming the electron transport chain. Electron carries deposit their electrons and H+ ions. Oxidative phosphorylation happens across the inner membrane. NADH and NAD+ go back to the krebs cycle. 4 H+ are passed through the first channel using energy from electrons, e- movs along the ETC and activates the next channel which the 4 H+ pass through. E- move along, losing energy. 2 H+ pass thorugh the third channel..
ETC - generating a proton motive force
The hydrogen carriers (NADH and FADH2) are oxidized and release high energy electrons and protons. The electrons are transferred to the electron tranpsort chain, which consists of several transmembrane carrier proteins. As electrons pass through the chain, they lose energy, which is used by the chain to pump protons (H+ ions) from the matrix. The accumulation of H+ ions within the intermembrane space creates an electrochemical gradient.
ETC - reduction of oxygen
in order for the ETC to continue functioning, the de-energized electrons need to be removed. Oxygen acts as the final electron acceptor, removing the de-energized elctrons to prevent the chain from being bloxked. Oxygen also binds with free protons in the matrix to form water to maintain the hydrogen gradient. Without oxygen, hydrogen carriers cannot transfer energized electrons to the chain and ATP production is halted.
Chemiosmosis
Chemiosmosis is the diffusion of ions across a semi-permeable membrane, through a carrier protein. In this case, the ions are hydrogen protons and the carrier is ATP synthase. The flow of H+ through ATP synthase generates ATP. Yields 32 ATP.
ATP synthesis via chemiosmosis
the proton motive force will cause H+ ions to move down their electrochemical gradient and diffuse back into the matrix. This diffusion of protons is Chemiosmosis (facilitated by ATP synthase). As they move through they trigger the molecular roation of the enzyme, synthesizing ATP.
Yeast fermentation - bread
Dough is kept warm after kneading to encoruage the yeast in it to respire. Yeast can respire anaerobically or aerobically, but oxygen in dough is used up to the yeast has to respire anaerobically. The carbon dioxide produced cannot escape from the dough and forms bubbles, causing the dough to swell and rise. Ethonal produced by anaerobic cell respiration evaporates during baking.
Bioethanol
Bioenthanol is a renewable energy source produced from sugar cane or maize using yeast. Starch and cellulose in the plant are broken down into sugars by enzymes and fermenters are used to keep the yeast in optimum conditions. When the yeast carries out anaerobic respiration the sugars are converted to ethanol and carbon dioxide. This ethanol is then purified and water is removed.
Lactic acid fermentation
This anaerobic respiration occurs when a person's exercise rate exceeds the body's ability to supply oxygen. Pyruvate molecules are converted to lactic acid. However, this causes stiffness and fatigue as more lactate accumulates in muscule tissue. Once O2 reaches the mitochondria, the lactate is transported to the liver where it turns back to pyruvate. The demand for oxygen that builds up during this is called oxygen debt, which is paid back by panting after excercise.
Variables affecting the rate of cell respiration
Temperature (effects rate of collision), carbon dioxide concentration (CO2 increases, rate of respiration decreases), oxygen concentration (O increases, rate of respiration increases), Glucose (increases, rate also increases), type of cell (some cells require more ATP than others and will have a higher rate).
respirometer
A device that determines an organism's respiration rate by measuring the rate of exchange of O2 and CO2.