biochem - enzymes.pdf

NIKKI GIGI LEGASPI RMT, MD Learning Objectives for Today: 1. Define what are enzymes and outline their biologic roles in the human body 2. Enumerate the different types of enzymes and their functions 3. Describe substrate binding in enzymes 4. Enumerate and describe the mechanisms of enzyme...

NIKKI GIGI LEGASPI RMT, MD Learning Objectives for Today: 1. Define what are enzymes and outline their biologic roles in the human body 2. Enumerate the different types of enzymes and their functions 3. Describe substrate binding in enzymes 4. Enumerate and describe the mechanisms of enzyme catalysis 5. Define isozymes and state its biomedical importance What is a Chemical Reaction? ▪A process by which substances interact to form a new substance ▪It would always involve a chemical change– breaking or forming of chemical bonds Physical Change vs Chemical Change  Physical Change Chemical Change Chemical make-up of Formation of a new chemical substance remains the same substance (Chemical Reaction) Evidence: Changes in size, Evidence: Gas production, shape and state of matter change in temperature, color (solid, liquid and gas) change, formation of precipitate Can be reversed Hard to reverse CHO(s) CHO(l) CHO(s) CO2 + H2O Chemical Reaction ▪Always involve reactants (substrates) and products ▪Reactants/Substrates – the atoms and molecules that interact with each other ▪Products – the atoms and molecules produced by the reaction Chemical Reaction = Reactants (Substrates) & Products Hydrogen Peroxide Water and Oxygen In a chemical reaction: 1. Reactants contact each other, bonds between them are broken, and atoms rearrange and form new bonds to make the products 2. Only the atoms present in the reactants can end up in the products. No new atoms are created and no atoms are destroyed ▪Law of Conservation of Mass: Total mass of reactants must be equal to the mass of the products ▪Mass can neither be created nor destroyed ▪Stoichiometry – chemistry of balancing chemical reactions ▪mass of reactants = mass of products Practice Samples for Balancing Equations All Chemical Reactions in the Body Depend on Enzymes Importance of Enzymes ▪All biological reactions within human cells depend on enzymes ▪Dr. Richard Wolfenden stated that creating DNA and RNA would take 78 million years in water without enzyme “Without enzyme catalysts, slowest known biological reaction takes 1 trillion years. Enzymes can make this happen in 10 milliseconds” -Dr. Richard Wolfenden  “Without catalysts, there would be no life at all, from microbes to humans. It makes you wonder how natural selection operated in such a way as to produce a protein that got off the ground as a primitive catalyst for such an extraordinarily slow reaction.” -Dr. Richard Wolfenden What are Enzymes? ▪Are biochemical catalysts ▪Catalysts – they accelerate/speed up chemical reactions without an interchange ▪Almost all are protein and globular Parts of Enzyme ▪Active Site - the part of the enzyme that acts as a Catalyst ▪E-S Complex – when the substrate binds at the active site of enzyme ▪Allosteric Site – regulatory site (positive or negative modulator) Parts of Enzyme Cofactors vs Coenzymes ▪Not all enzymes are able to catalyze reactions on their own ▪Some need a little help: Cofactor/Coenzyme What are Cofactors? ▪Inorganic/organic molecules that bind in a transient, dissociable manner either to enzyme or substrate ▪Two (2) types of Cofactors: 1. Metal ions – inorganic molecules (Fe, Zn, Mg, Cu) 2. Coenzymes – organic molecules, loosely bound Metal Ions ▪Participate directly into the catalysis ▪May stabilize the enzyme or substrate, or help convert the substrates to products ▪Example: DNA Polymerase (Mg++) Coenzymes ▪Organic carrier cofactors ▪Serve as recyclable shuttles or group transfer agents that transport many substrates from point of generation to point of utilization ▪Indirectly participate in catalysis ▪Example: ▪Vitamin derivatives ▪Examples: Nicotinamide Adenine Dinucleotide (NAD), Thiamine pyrophosphate (TPP), Flavin adenine dinucleotide (FAD), Coenzyme A (CoA) Holoenzyme vs Apoenzyme  APOenzyme = ENZYME ONLY HOLOenzyme = ENZYME + COFACTOR Enzymatic Process ▪The catalytic properties of enzymes are surprising ▪Example: Catalase is such a high activity enzyme that it transforms harmful H2O2 into water and oxygen: 2H2O2 2H2O + O2 ▪One molecule of this catalase repeats this reaction 107 times in 1 second (turnover) ▪Most enzyme accelerate reactions at similar high rates, but there are some exceptions ▪1 molecule of lysozyme enzyme, which dissolves glycosidic bonds in the bacterial cell wall, can bind to only 1 molecule at a time Two Models for Substrate Binding 1. Lock and Key Hypothesis 2. Induced Fit Hypothesis The Lock-and-Key Hypothesis ▪Is a model postulated by Emil Fischer ▪Lock = enzyme ▪Key = substrate ▪Only the correctly sized key (substrate) fits into the key hole (active site) of the lockhole (enzyme) The Induced Fit Hypothesis ▪A more recent model, backed up by evidence and more widely accepted ▪Proposed by Daniel Koshland ▪The shape of active sites are not exactly complementary, but change shape in the presence of a specific substrate to become complementary How do Enzymes Speed Up Chemical Reactions? ▪Chemical reactions involve breaking and reforming of chemical bonds ▪The breaking of bonds needs energy to get started even if the overall process is energetically favorable ▪The energy needed to get started is called the Activation Energy Activation Energy = EA ▪Activation energy: minimum energy requirement that must be met for a chemical reaction to occur Enzymatic Mechanisms to Facilitate Catalysis 1. Catalysis by Proximity 2. Catalysis by Strain 3. Covalent Catalysis 4. Acid-Base Catalysis  Catalysis by Proximity ▪For molecules to interact, they must be in bond-forming distances to each other ▪Active Site – creates a region of high local substrate concentration in which the molecules are oriented in a position ideal for them to chemically interact  Catalysis by Strain ▪Enzymes typically bind substrates in a conformation that is somewhat unfavorable ▪This strained conformation mimics the structure of the Transition State Intermediate Which substance is a more effective enzyme effector/inhibitor: A substrate analog? A transition state analog? Covalent Catalysis ▪Involves formation of a covalent bond between the enzyme and one or more substrates ▪Modified enzyme becomes a reactant (transient) ▪Usually cysteine, serine & histidine  Acid-Base Catalysis ▪The prosthetic groups in an enzyme acts as an acid or a base and removes/donates protons of the substrate ▪Acid catalysis: sensitive to H+ or protons ▪Base catalysis: OH- ions  How to Lower the Activation Energy? ▪The Activation Energy is lowered by achieving a Transition State Stabilization ▪How to achieve the transition state: 1. Approximate and orient substrates correctly (Proximity) 2. Putting stress on the bonds within the molecule (Strain) 3. The enzyme may provide another lower-energy way for the reaction to take place (Covalent) 4. Inducing charge distribution on substrates (Acid-Base) Enzymes are Effective & Highly Specific Catalysts Characteristics of Enzymes: 1.Neither consumed nor permanently altered in the reaction 2.High Specificity and Selectivity 3. Stereospecific  “Three-Point Attachment Model” of Enzymes Enzymes usually bind to at least three (3) sites of the substrate ◦ First attachment: highest affinity and most specific (attaches to C- terminus and N-terminus) ◦ Second attachment: lower affinity (usually through phospholipid head groups) ◦ Third attachment: lowest affinity (Hydrophobic interactions) ▪Each enzyme catalyzes only one type of reaction, and the reaction exhibits selectivity to the substrate and its products ◦For example, an enzyme polypeptide of Trypsin cleaves from where the amino acid of lysine or arginine are present ◦Chymotrypsin does the same job with phenylalanine, tyrosine and tryptophan Naming of Enzymes 1. First principle: Enzymes usually end with –ase (Exceptions: Trypsin, Chymotrypsin, Pepsin) Examples: Succinate oxidase, cytochrome oxidase, hexokinase 2. Second principle: Enzymes are mainly classified and named according to the reaction they catalyze Examples: Redox reactions: Oxidoreductases, dehydrogenases, oxidases Protein degradation: Protease; Lipid degradation: Lipase 3. Third Principle: First noun in the enzyme name is usually the substrate Example: Glucose-6-Phosphate dehydrogenase, Pyruvate dehydrogenase Classification of Enzymes Enzyme Classification (OTH LIL) 1.Oxidoreductases 2.Transferases 3.Hydrolases 4.Lyases 5.Isomerases 6.Ligases Oxidoreductases Are class of enzymes that catalyze oxidation-reduction reactions Catalyze the transfer of electrons from one molecule (Oxidant) to another molecule (Reductant) A- + B A + B- Examples: ◦Oxidases: when molecular oxygen acts as acceptor of hydrogen or electrons ◦Dehydrogenases: transfer hydrogen to a NAD+/NADP+ or a Flavin enzyme ◦Peroxidases: found in peroxisomes (reduction of hydrogen peroxide) ◦Hydroxylases: add OH groups to its substrates ◦Oxygenases: incorporate molecular oxygen into organic substrates ◦Reductases: catalyse reductions  Transferases Enzymes transferring a group e.g. methyl group, glycosyl or phosphoryl group from one compound (donor) to another compound (acceptor) A-B + C A + B-C Examples of Transferases Kinases Phophorylases Hydrolases ▪Catalyze hydrolytic cleavage of C-C, C-O, C-N and other covalent bonds ▪Breakage of bonds through addition of water Lyases ▪Enzyme that catalyzes the breaking of chemical bond through means other than hydrolysis or oxidation, and in turn forms a double bond ▪Break C-C, C-O, C-N bonds Isomerases ▪Catalyze reactions that transfer functional groups within a molecule so that isomeric forms are produced ▪Examples: ▪ Racemases – interconvert L and D stereoisomers ▪ Mutases – transfer groups between atoms ▪ Isomerases – transfer groups to form isomers  Ligases ▪Catalyze the joining together (ligation) of two molecules in reactions coupled to ATP hydrolysis ▪Examples: ▪ Ligases ▪ Synthase ▪ Synthetase Isozymes Isozymes ▪Isozymes – are physically distinct versions of a given enzyme, each of which catalyzes the same reaction ▪Enzymes that differ in structure (amino acid sequences) but catalyzes the same reaction  LDH (Lactate Dehydrogenase) Enzyme ▪Enzyme responsible for the conversion of pyruvate (end product of glycolysis) into lactic acid ▪This conversion is necessary when a cell has little oxygen There are 5 human isoforms of LDH in humans LDH-1 (HHHH): Heart, RBC LDH-2 (HHHM): Reticuloendothelial System LDH-3 (HHMM): Lungs LDH-4 (HMMM): Kidney, Placenta LDH-5 (MMMM): Liver Predominant form in the serum: LDH-2 What happens in… 1. Acute Myocardial Infarction? 2. Liver disease? Learning Objectives for Today: 1. Define what are enzymes and outline their biologic roles in the human body 2. Enumerate the different types of enzymes and their functions 3. Describe substrate binding in enzymes 4. Enumerate and describe the mechanisms of enzyme catalysis 5. Define isozymes and state its biomedical importance References: 1. https://www.slideshare.net/karthi131087/final-44690151 2. http://facweb.northseattle.edu/lizthomas/Lecture%208.pdf 3. Harper’s Biochemistry, 30th Edition 4. https://socratic.org/questions/how-does-an-enzyme-lower-the-activation-energy