Biochemistry Past Paper PDF 2024

28 DUHA AL-NADER HUSNI MEDHAT Dr. NAFEZ ABU-TARBOUSH 1  Enzymes -enzymes are mostly proteins that serve as biological catalysts that speed up chemical reactions(catalysis) within a biological system, while metals are natural catalysts...

28 DUHA AL-NADER HUSNI MEDHAT Dr. NAFEZ ABU-TARBOUSH 1  Enzymes -enzymes are mostly proteins that serve as biological catalysts that speed up chemical reactions(catalysis) within a biological system, while metals are natural catalysts that primarily function outside biological systems -They’re present in low concentrations because they can be reused -An enzyme increases rate of reaction by lowering activation energy and isn’t consumed during the reaction which means that enzymes catalyze reactions without being permanently changed & are considered reactants that have their own mechanism. The enzyme binds to the substrate forming an enzyme-substrate complex resulting in a conformational change in the enzyme, the shape change is temporary, and once the reaction is completed and products are released the enzyme retains its original form and can be reused again. -Enzyme activity is regulated through activation and inhibition -They’ve optimal PH and so the negatively and positively charged amino acids in the enzyme’s active site which is crucial for binding the substrate to break bonds & form new ones in the products -Enzymes are products of DNA expression, DNA mRNA protein General properties of proteins The function of nearly all proteins depends on their ability to bind other molecules (ligands) Two properties of a protein characterize it’s interaction with ligands: Affinity: the strength of binding between a protein and other molecule Specificity: the ability of a protein to bind one molecule in preference to other molecules (how specific/selectively it binds to its target) 2 -when affinity decreases, specificity might decrease or when affinity increases specificity might decrease but they usually go together. -If the binding between the enzyme and its substrate is very strong it implies that the enzyme only reacts with this substrate (high specificity). -If the binding between the enzyme and the substrate is weak it means it can bind to other substrates thereby specificity decreases and affinity decreases at the same time. Are enzymes important? In the human body, almost every metabolic process involves the use of enzymes Enzymes are capable of speeding up chemical reactions and they’re reusable which means huge amount of products can be generated using a low concentration of enzymes. Green tea which is (minimally oxidized) contains antioxidant and differs from black tea (fully oxidized) in drying methods black-sun green- shadow. In cells of tea there are polyphenols that’s why it’s used as an antioxidant. Phenol has a cyclic structure that can accommodate 3 radicals thereby functioning as an antioxidant. crushed leaves are exposed to the oxygen in air, a polyphenoloxidase (requires molecular oxygen) breaks up polyphenols into tannins which impart the darker color and characteristic flavors & scents. 3 -Amoxicillin is the most used antibiotic *When a certain texture of candies is desired sweet viscous materials can be added such as corn syrup instead of honey which is considered expensive. *Deserts with liquid core, sucrase enzymes breaks down sucrose into monosaccharides which are more soluble in this core. *Enzymes (proteases, peptidases) are added to meat to break peptide bonds of proteins thereby fractioning these proteins making them more soluble & accordingly meat becomes softer and more expensive. *Enzymes are added to solutions of contact lenses, break down any deposits on surfaces of contact lenses preventing eye infections. *Enzymes (proteases, lipases) are added to washing powders to remove tough stains. *Main 4 points of enzymes definition: 1-Mostly proteins 2-Catalyze biochemical reactions (biological catalysts) 3-present in very low amounts 4-they can be reused & by the end of reaction they are undergone no change The biological catalysts, enzymes What are enzymes? (specialized proteins, small amounts, acceleration, no change). Ribozymes are the exception (ribozymes are RNA molecules that function as enzymes, so they are not 4 proteins) -Enzymes speed chemical reactions by reducing time required leading to faster reaction. -Reactions can occur with or without the presence of enzymes, and what controls the occurrence of a reaction (whether to go or not to go) is the energy of reactants & products not the enzymes. Enzymes are the most efficient catalysts known Usually in the range of 10^6 10^14 Non-enzymatic catalysts (10^2 to 10^4) The actions of enzymes are fine-tuned by regulatory processes Examples: catalase (10^8) & carbonic anhydrase (10^7) Catalase breaks hydrogen peroxide into water and oxygen gas. -Platinum surface speeds the reaction, but it isn’t a biological catalyst since it isn’t contained within a biological system. Placing H2O2 on platinum surface increases speed of rxn by 10,000 folds. -Catalase increases speed of reaction by 100,000,000 folds. 1 mole is converted per second (reaction without catalase) one hundred million molecules are converted per second (reaction with catalase) *The difference in activity between catalase and platinum emphasizes that enzymes are the best-known catalysts. 5  *When a piece of meat is dipped into hydrogen peroxide solution, catalase in meat will react with H2O2 which produce the oxygen & bubbles appears. How to express an enzymatic reaction? In enzymatic reactions, reactants are known as substrates We can simply express an enzymatic reaction using this formula Biochemical reactions occur inside cells, while in enzymes reactions occur in active site. where E is the free enzyme; S is the free substrate, ES is the enzyme-substrate complex; P is the product of the reaction; and EP is the enzyme-product complex before the product is released, enzyme is undergone no change at end of rxn (exits as it entered) Active sites of enzymes -Some proteins function as enzymes while others don’t, depending on the presence of active site which makes them function as enzymes. A specific three-dimensional shape which includes a region where the biochemical reaction takes place Contains a specialized amino acid sequence that facilitates the reaction Enzymes provide a binding site for other materials which can change the shape of active site (since proteins can change in shape) making it better (activators) or worse(inhibitors) 6  Within the active site are two sub-sites, the binding site (hydrophobic interaction with substrate) and the catalytic site (responsible for reaction), The binding & catalytic site may be the same in case of having small active site Binding site: binds substrate through ionic, H-bonding or other electrostatic forces, or hydrophobic interactions (weak bonds) Catalytic site: contains the catalytic groups Features of active sites Active sites; structures that look like canals, clefts or crevices (not present on the surface of enzyme) in order to interact with the substrate and initiate chemical reaction Compatible with the substrate Water is usually excluded after binding unless it participates in the reaction Substrates are bound to enzymes by multiple weak attractions in other words non-covalent bonds (electrostatic, hydrogen, van der Waals, & hydrophobic) Binding occurs at least at three points (chirality). To discriminate between isomers such as chiral or achiral, D or L rotation enzyme binds to the substrate at 3 or more binding sites. If binding occurs at 2 binding sites enzymes won’t specify whether it’s D or L orientation of the substrate thereby binding occurs at 3 binding sites or more. 7 Forms by groups from different parts of the amino acid sequence usually forming a domain made of multiple secondary structures -lined with amino acids that aren’t in a specific order (ex; side chain from AA #13 and side chain from AA #271) , side chains of these A.A constitute the active site and project toward the space allowing the active site to react with substrate. -if they were projected toward the interior, all active sites of all enzymes would be the same (R chains pointing interiorly, identical backbone toward the space), and this is lead us to loose the main feature of enzymes. Takes up a relatively small part of the total volume, comprising approx. 1-2% of enzyme, rest 98% stabilizes the conformation or 3D structure of active site by huge number of A.As that preserve and strengthen conformation resulting in an enzyme which is highly specific to a substrate. The “extra” amino acids also help create the three-dimensional active site & in many enzymes, may create regulatory sites. -Release the product to become reusable again allowing them to catalyze many reactions without being consumed. -Covalent bonds are not involved in initial binding of substrate to the active site. Only toxins, poisons &drugs bind covalently to enzymes, anything physiological can’t bind covalently and reaction is carried on normally. However, it’s possible for a covalent bond to be formed (as an intermediate, temporarily) and broken down during the catalyzed reaction. 8 How do enzymes work (model wise) Binding leads to formation of transition-state Usually, substrate binds by non-covalent interactions to the active site The catalyzed reaction takes place at the active site, usually in several steps Two models, lock-and-key vs. induced-fit model Glucose and hexokinase, phosphorylation *Improving the binding site for ATP & excluding water (might interfere with the reaction) Lock & key model This theory has failed or has been suppressed by other models for the following reasons 1-More than one key can interact with the same lock, for example, glucokinase doesn’t only react with glucose it can react with galactose and fructose but less efficiently. 2-Proteins are dynamic in nature and have huge number of conformations for each protein at least one of them is active and is called ‘meta conformation’, so a protein can have million conformations while the lock and key theory suggests that an enzyme has a static conformation. Induced-fit model As the name implies fit is induced and it wasn’t there from the beginning. -What stimulates complementarity is the binding of substrate. Initial affinity between substrate and active site causes the active site to bind to the substrate & change in shape to better enclose the substrate -Glucokinase (phosphorylates glucose) transfers phosphate group from ATP to glucose, and phosphate group is added on carbon number 6. 9 -Glucose enters active site and the enzyme adds phosphate on glucose. Energy and biochemical reactions Reactions occur to form a more stable substance (lower energy scale). In energy graphs products should be of lower energy scale compared to reactants ΔG= ΔH-TΔS Spontaneous vs. non-spontaneous, favorable vs. non-favorable, exergonic vs. endergonic, exothermic vs. endothermic, switch of signs ΔG, ΔG° -difference between energy of products and reactants and since energy of products is lower ΔG is always -ve Biochemical pathways; storage (endergonic) & release (exergonic) Kinetics (rate) vs. Thermodynamics (favorability) -Spontaneous is a biochemical term which implies that products have lower energy scale compared to reactants and the difference in energy will be released to the surrounding that’s why it’s called exergonic. -Endergonic reactions are non-spontaneous, unfavorable and ΔG is +ve *In real life, both don’t occur spontaneously unless an enzyme is incorporated as they take millions of years to occur 10  How do enzymes work (from an energy point of view) Enzymes speed up reactions, but have no relation to equilibrium or favorability What is an activation energy (ΔG°‡) concept? Is the energy required to convert any substance to the transition state Specificity varies (stereoisomers), however, there is none non-specific Spontaneous vs. rate! What is the transition state? An unstable high-energy state that substrates must pass through during the reaction *Transition-state complex binds more tightly to the enzyme compared to substrate In physics, for a reaction to occur the molecules collide with each other (random collisions) producing energy & this energy must be sufficient to change the compound from the stable condition to the unstable condition, it either becomes a product or reverts back into reactant. In most cases this energy isn’t sufficient to reach instability which accounts for the use of enzymes Glucose is stable under normal conditions but it should be converted to a substance that’s unstable to change into something else. 11 Alternative pathways - Substrates of enzymatic reactions often undergo several transformations when associated with the enzyme and each form has its own free energy value -Which one is the activation energy? The graph shows 3 Intermediates, the one of the highest peak from reactant energy level is the activation energy. - Activation energy & final ΔG calculation: ΔG remains the same as the energy of products and reactants doesn’t change & activation energy is lowered -Enzymes don’t interfere with the starting or ending points of a reaction Adenosine deaminase 12 How do enzymes work? (mechanism of action) Proximity effect: Bring substrate and catalytic sites together Orientation effect: Hold substrate at the exact distance and in the exact orientation necessary for reaction allowing hydrophobic and polar interactions in nearest attack conformation which is the best conformation of enzyme to attack the substrate Catalytic effect: Provide acidic, basic, or other types of groups required for catalysis (converting substrate) Energy effect: Lower the energy barrier by inducing strain in bonds in the substrate molecule Catalysis by proximity and orientation (Done by all enzymes) Enzyme-substrate interactions orient reactive groups and bring them into proximity with one another favoring their participation in catalysis Such arrangements have been termed nearattack conformations (NACs) NACs are precursors to reaction transition states 13 Catalysis by bond strain (specific to certain enzymes) -In this form of catalysis, the induced structural rearrangements produce strained substrate bonds (to facilitate its breakage) reducing the activation energy. Example: lysozyme The substrate, on binding, is distorted from the typical 'chair' hexose ring into the 'sofa' conformation, which is similar in shape to the transition state Catalysis involving proton donors (acids) and proton acceptors (bases) -The R groups of amino acids in the active sites are positively and negatively charged therefore can act as donors or acceptors of protons from substrate. -Histidine is an excellent proton donor/acceptor at physiological pH -Example: serine proteases 14 Covalent catalysis -A substrate can never bind initially to the active site through a covalent bond -During this mechanism a covalent intermediate forms between the enzyme or coenzyme and the substrate -Examples of this mechanism is proteolysis by serine proteases, which include digestive enzymes (trypsin, chymotrypsin, and elastase) ‫تمت كتابة هذا الشيت صدقة جارية عن روح والدة زميلنا عمرو رائد من دفعة تيجان‬ ‫دعواتكم لها بالرحمة والمغفرة‬ 15