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Lecture 12: Enzyme Catalysis Topics: Catalytic Strategies Steps in a Reaction Summary: The chemical reactions catalyzed by most enzymes can be classified into one of 6 general types of reactions. A few catalytic strategies are used by most enzymes regardless of the particular chemistry they perform. Enzyme-catalyzed reactions proceed in an organized series of steps each of which can be considered separately. Key Concepts: Covalent catalysis Nucleophile Acid-Base Catalysis Metal-ion Catalysis Catalysis by Approximation Electrostatic Effects, Desolvation, Induced Fit Displacement reactions Lecture 13:Mechanism of Chymotrypsin Topics: Chemical Mechanism of Chymotrypsin Summary: Chymotrypsin is a protease and its activity is regulated by controlled cleavage of its backbone. Its chemical mechanism proceeds in two stages: 1. Nucleophilic attack on substrate by Ser 195 to form acyl-enzyme complex followed by 2. Deacylation though nucleophilic attack by water on the acyl intermediate. Key Concepts: Meaning of burst phase and labelling of Serine 195 Catalytic triad: Roles of His 57, Asp 102, and Ser 195 in mechanism Occurrences of acid-base catalysis and covalent catalysis in mechanism Lecture 14: Regulation of Proteins 1: Allosteric Control of ATCase Topics: Overview of Regulatory Mechanisms Description of ATCase Allosteric Properties of ATCase Summary: Aspartate transcarbamoylase (ATCase) is an allosteric enzyme which carries out the first step in the synthesis of pyrimidine nucleotides. Allosteric enzymes use changes in conformation to switch between different states which have different levels of activity. Binding of allosteric effectors can control the switch between states, and thereby increase or decrease the enzyme activity to exert control over biological processes. Key Concepts: Types of Regulation Feedback inhibition Allosteric transition in ATCase ATP and CTP as allosteric effectors of ATCase Lecture 15: Regulation of Proteins 2: Allosteric Control of Hemoglobin Topics: Hemoglobin and Myoglobin Allosteric Transition in Hemoglobin Physiological Role of Hemoglobin Summary: Hemoglobin is an oxygen transport protein that carries oxygen from the lungs, where oxygen levels are high, to tissues where it is needed. Hemoglobin exhibits cooperative binding of oxygen which is the result of a conformational switch between a low-affinity state and a high-affinity state. The affinity of hemoglobin for oxygen can be regulated by allosteric effectors to improve its physiological performance. Key Concepts: Function of hemoglobin Conformational changes due to oxygen binding Cooperativity Role of BPG Role of Bohr Effect Lecture 16: Regulation of Proteins 3: Isozymes and Covalent Modification Topics: Isozymes Covalent Modification Protein Kinase A Summary: Isozymes are enzymes which have the same activity but different kinetics or regulatory properties- differential expression of isozymes allows control over enzyme activity. Many proteins are regulated by covalent modification. The most common such modification is phosphorylation. Protein kinase A carries out phosphorylation of a wide variety of targets in response to cyclic AMP. Key Concepts: Isozymes of LDH Regulation of histones by acetylation Kinases and Phosphatases Role of Protein Kinase A Lecture 17: Regulation of Proteins 4: Proteolytic Activation Topics: Examples Activation of Digestive Enzymes Blood Clotting Summary: Zymogens are inactive protein precursors which must be converted to their active forms by specific proteolytic cleavage events. A variety of digestive enzymes are synthesized as zymogens in the pancreas. They are activated by proteolysis, and further control of their activities is achieved through the action of specific inhibitor proteins. A cascade of zymogen activations resulting in the controlled creation of fibrin aggregates is the molecular basis of blood clotting. Key Concepts: Zymogens Control of activation Roles of inhibitor proteins (Serpins) Emphysema Activation Cascades Mechanism of blood clotting Hemophilia Lecture 18: Introduction to Membranes Topics: Lipid Structure Properties of Lipid Bilayers Summary: Biological membranes are composed of lipids and proteins and form the boundary of the cell and its compartments. Phospholipids and glycolipids are formed of fatty acids esterified to a platform (backbone) molecule and contain other groups such as alcohols or sugars. Lipids spontaneously assemble into bilayers which are largely impermeable to charged and polar molecules and which form closed compartments. Key Concepts: Fatty acids Phospholipids Glycolipids Cholesterol Micelles Bilayers Vesicles Permeability of bilayers Lecture 19: Membrane Proteins Topics: Architecture of Membrane Proteins Fluid Mosaic Model Protein Targeting Summary: Different membranes have different protein contents which confer their different capabilities. Membrane proteins associate with lipid bilayers in a variety of ways. In some cases the amino acid sequence can suggest structural features in such proteins. Proteins have internal sequence codes for the organelle to which they should be targeted. Key Concepts: Peripheral and Integral membrane proteins Architecture of membrane proteins Hydropathy plots Fluid mosaic model Targeting sequences Lecture 20: Membrane Transport Topics: Energetics of Transmembrane Transport Active Transport Passive Transport Summary: Transmembrane concentration differences are not at equilibrium and therefore are a source of energy which can be used for other purposes. Two categories of active transporters are pumps, which use the energy of ATP hydrolysis to transport molecules across membranes, and secondary transporters, which use the energy stored in a transmembrane concentration gradient. Passive transporters facilitate diffusion of molecules through membranes and allow equilibrium to be more rapidly established. Ion channels are examples of passive transporters and play important roles in nerve cell function. Key Concepts: Thermodynamics of transport Active Transport Primary transporters (pumps) Secondary transporters (antiporters, symporters) Passive transport Ion channels Ligand-gated and Voltage-gated ion channels