Biochemistry Chapter 4 Enzymes PDF

Chapter 4 Proteins II: Enzymes Biochemistry First Edition John Tansey Lecture PowerPoints Tanea Reed/ Dr. Jose Sapien Chapter 4 Outline 4.1 Regarding enzymes 4.2 Enzymes increase reaction rate 4.3 The mechanism of an enzyme can be deduced from structural, kinetic, and spectral data 4.4 Examples o...

Chapter 4 Proteins II: Enzymes Biochemistry First Edition John Tansey Lecture PowerPoints Tanea Reed/ Dr. Jose Sapien Chapter 4 Outline 4.1 Regarding enzymes 4.2 Enzymes increase reaction rate 4.3 The mechanism of an enzyme can be deduced from structural, kinetic, and spectral data 4.4 Examples of enzyme regulation Copyright © 2019 John Wiley & Sons, Inc. Enzymes Defined ▪ Enzymes are catalysts involved in biochemical reactions. • Increase the rate of reaction • Usually globular proteins • Can be monomeric or multimeric Copyright © 2019 John Wiley & Sons, Inc. Substrate Defined ▪ Substrate is a molecule (or molecules) that acts as the reactant in a enzymatically catalyzed reaction. • More than 60% of biochemical reactions use multiple substrates. • Binds to the enzyme at the active site • When the enzyme binds to the substrate, product(s) is/are formed. Copyright © 2019 John Wiley & Sons, Inc. mage modified from "Enzymes: Figure 4," by OpenStax College, Biology, CC BY 3.0._ Source: https://www.khanacademy.org/science/ap-biology/cellularenergetics/environmental-impacts-on-enzyme-function/a/enzyme-regulation Copyright © 2019 John Wiley & Sons, Inc. Active Site Defined ▪ Active site is the location where the enzyme binds to the substrate and catalysis occurs. • Located on the surface of the enzyme • Is often in a cleft, pocket, or trench • Generally formed by residues on turns or coils Copyright © 2019 John Wiley & Sons, Inc. Topology of Active Site Figure 4.4 Topology of the active site. Copyright © 2019 John Wiley & Sons, Inc. Enzyme–Substrate Binding Models (1 of 2) ▪ Lock and key Figure 4.3A Models of substrate binding. Copyright © 2019 John Wiley & Sons, Inc. Enzyme–Substrate Binding Models (2 of 2) ▪ Induced fit Figure 4.3B Models of substrate binding. Copyright © 2019 John Wiley & Sons, Inc. Enzyme Classifications (1 of 2) ▪ Oxidoreductases catalyze reactions involving the gain or loss of electrons ▪ Transferases transfer one group to another ▪ Hydrolases cleave a bond with water Copyright © 2019 John Wiley & Sons, Inc. Enzyme Classifications (2 of 2) ▪ Lyases break double bonds using some other means than oxidation or hydrolysis ▪ Isomerases catalyze a rearrangement of the molecule ▪ Ligases join two molecules Copyright © 2019 John Wiley & Sons, Inc. Enzyme Mechanism ▪ Lowers activation energy (Ea ) to speed up (increase) reaction rate ▪ Does not change thermodynamic parameters Figure 4.5 Enzymes lower activation energy. Copyright © 2019 John Wiley & Sons, Inc. Section 4.2 Enzymes Increase Reaction Rate Copyright © 2019 John Wiley & Sons, Inc. Michaelis–Menten Constant ▪ For the reaction, ▪ (ES) = enzyme–substrate (Michaelis) complex Copyright © 2019 John Wiley & Sons, Inc. The two assumptions ▪ The first assumption was that k2 is much slower than k−1, enabling an equilibrium to be established between E, S, and the ES complex: Copyright © 2019 John Wiley & Sons, Inc. The two assumptions ▪ The second assumption (the steady state assumption) assumes that the ES complex forms rapidly from free enzyme and substrate, and it exists at a relatively unchanging concentration as the reaction proceeds until substrate is depleted Copyright © 2019 John Wiley & Sons, Inc. Michaelis–Menten Equation ▪ It is the central equation of enzyme kinetics that relates substrate concentration [S] to two fundamental properties of the enzyme: • The theoretical maximal velocity for a given concentration of enzyme (Vmax) and the Michaelis constant (KM). Copyright © 2019 John Wiley & Sons, Inc. Michaelis–Menten Equation ▪ The maximal rate achievable (Vmax) occurs when the entire enzyme is saturated with substrate. ▪ Vo = initial velocity. Copyright © 2019 John Wiley & Sons, Inc. Saturation Kinetic Curve ▪ If there is enough substrate, the enzyme will reach full saturation. What about high or low Km values?? Figure 4.7 Saturation kinetic curve. Copyright © 2019 John Wiley & Sons, Inc. Lineweaver–Burke Plot (1 of 2) ▪ Double reciprocal of Michaelis–Menten equation ▪ Easier method to interpret graphical data Copyright © 2019 John Wiley & Sons, Inc. Lineweaver–Burke Plot (2 of 2) Figure 4.8 Lineweaver–Burke plot. Copyright © 2019 John Wiley & Sons, Inc. Turnover Number Defined ▪ Turnover number (kcat) is the number of reactions the enzyme can catalyze per unit of time. ▪ Can be used to measure catalytic efficiency, which describes how often a reaction occurs for every encounter of enzyme and substrate Copyright © 2019 John Wiley & Sons, Inc. Diffusion Controlled Limit Defined ▪ Diffusion controlled limit is an occurrence when ratelimiting step becomes the diffusion of enzyme and substrate together. ▪ The enzymes catalyze a reaction nearly every time they encounter a substrate. ▪ The rate must be between 108 and 109 M−1 sec−1. Copyright © 2019 John Wiley & Sons, Inc. Inhibitors ▪ Prevent the generation of products ▪ Can be irreversible or reversible Copyright © 2019 John Wiley & Sons, Inc. Suicide Inhibitors Defined ▪ Suicide inhibitors directly poison the enzyme. ▪ Covalently modify the active site of the enzyme, irreversibly blocking its function ▪ Inactivate enzyme ▪ Examples include pesticides and nerve agents. Copyright © 2019 John Wiley & Sons, Inc. Copyright © 2019 John Wiley & Sons, Inc. Irreversible Enzyme Inhibitors Figure 4.9 Irreversible enzyme inhibitors. Copyright © 2019 John Wiley & Sons, Inc. Competitive Inhibition ▪ Recognizes molecules similar to the shape of the substrate that binds to the active site ▪ Competes directly with the substrate ▪ Can be overcome if the substrate concentration is high Copyright © 2019 John Wiley & Sons, Inc. Michaelis–Menten Equation for Competitive Inhibition Copyright © 2019 John Wiley & Sons, Inc. Competitive Enzyme Inhibitors Figure 4.10 Competitive inhibition. Copyright © 2019 John Wiley & Sons, Inc. Uncompetitive Inhibition ▪ Binds to the ES complex ▪ Decreases Vmax ▪ Decreases KM Copyright © 2019 John Wiley & Sons, Inc. Michaelis–Menten Equation for Uncompetitive Inhibition Copyright © 2019 John Wiley & Sons, Inc. Uncompetitive Inhibition Plots Figure 4.11 Uncompetitive inhibition. Copyright © 2019 John Wiley & Sons, Inc. Mixed Inhibition ▪ Bind in the presence or absence of substrate ▪ A combination of competitive and uncompetitive inhibitors ▪ Effective regardless of substrate concentration Copyright © 2019 John Wiley & Sons, Inc. Michaelis–Menten Equation for Mixed Inhibition Copyright © 2019 John Wiley & Sons, Inc. Mixed Inhibition Plots Figure 4.12 Mixed inhibition. Copyright © 2019 John Wiley & Sons, Inc. Summary of Inhibition Inhibitor Apparent Km Apparent Vmax None Km Vmax Competitive αKm Vmax Uncompetitive Km/α′ Vmax/α′ Mixed αKm/α′ Vmax/α′ Copyright © 2019 John Wiley & Sons, Inc. Copyright Copyright © 2019 John Wiley & Sons, Inc. All rights reserved. Reproduction or translation of this work beyond that permitted in Section 117 of the 1976 United States Act without the express written permission of the copyright owner is unlawful. Request for further information should be addressed to the Permissions Department, John Wiley & Sons, Inc. The purchaser may make back-up copies for his/her own use only and not for distribution or resale. The Publisher assumes no responsibility for errors, omissions, or damages, caused by the use of these programs or from the use of the information contained herein. Copyright © 2019 John Wiley & Sons, Inc.