Enzymes & Metabolism Enzyme characteristics • Made of protein Enzymes and • Catalysts: – reactions occur 1,000,000 times faster with Metabolic Pathways enzymes – Not part of reaction • Not changed or affected by reaction • Used over and over Mechanism of Enzyme Action Why do cells need enzymes? • Ability of enzymes to lower activation energy • Not enough time to leave things to chance due to structure. • Most probable reactions are not the required • Each type of enzyme has has a highly-ordered, • Important molecules do not always occur in large characteristic 3-dimensional shape quantities (conformation). – Ridges, grooves, and pockets lined with specific amino acids. But since life processes are dependent upon enzymes, – Pockets active in catalyzing a reaction are called organisms cannot survive at high temperature the active sites of the enzyme. – Heat denatures proteins Reactants must collide for reaction Mechanism of Enzyme Action (continued) to occur • Substrates have specific shapes to fit into the active sites (lock- and-key model): – Substrate fits into active sites in enzyme. – Perfect fit may be induced: • Enzyme undergoes structural change. – Enzyme-substrate complex formed, then dissociates. – Products formed and enzyme is unaltered. Heyer 1 Enzymes & Metabolism Without enzymes, collisions are An enzyme brings reactants random together by binding to them Induced Fit Model Induced Fit Model 1. Enzyme binds 2. Substrates orient substrate, forms for productive 3. Productive collision 4. Reaction occurs, complex collision produces transition products are state released Naming of Enzymes Enzymes as Catalysts • Enzyme name ends with suffix “-ase.” • Name = substrate – action – “-ase” – E.g., glucose phosphorylase is an enzyme that adds a phosphate to glucose. – If the “action” is left out of the name, assume the action is hydrolysis. E.g., a protease catalyzes the hydrolysis of proteins into oligopeptides or amino acids. • Different organs may make different enzymes (isoenzymes) that have the same activity. – Differences in structure do not affect the active sites. Heyer 2 Enzymes & Metabolism Regulation of Enzyme Activity Reversible Reactions • Rate of enzyme-catalyzed reactions measured by the rate that • Some enzymatic reactions are reversible. substrates (reactants) are converted to products. – Both forward and backward reactions are catalyzed by • Factors regulating enzyme activity: same enzyme. 1. Mass action: concentration of substrates and products. carbonic anhydrase 2. Concentration of enzyme. • H 0 + C0 H C0 • Gene expression. 2 2 2 3 • Post-translational modification. • Proteolysis. • Law of mass action: 3. Alteration of enzyme structure. – Principal that reversible reactions will be driven from the • Activation. side of the equation where concentration is higher to side • Temperature. where concentration is lower. • pH. • But most biological reactions have complex organic 4. Cofactors and Coenzymes. substrates which require separate enzymes to catalyze 5. Inhibitors. the reverse reaction. Enzyme Activation Substrate Concentration • Enzymes may be produced in an inactive form. • Final post-translational modification occurs only • At a specific [enzyme], when enzyme is needed. rate of product formation increases as the [substrate] – In pancreas, digestive enzymes are produced as increases. inactive zymogens, which are activated in lumen of • Plateau of maximum intestine. velocity occurs when enzyme is saturated. • Part of the polypeptide is hydrolyzed off. • Additional [substrate] • Protects against self-digestion. does not not increase – In liver cells, enzyme is inactive when produced and reaction rate. is activated by addition of phosphate group. • Phosphorylation/dephosphorylation: Activation/inactivation of an enzyme. Enzyme Shape Effect of Temperature • Function of the enzyme is dependent on its shape • Rate of reaction increases as temperature increases. • Destruction of shape destroys function • Reaction rate plateaus, slightly above body temperature (37o C). – Heat • Reaction rate decreases as temperature increases. – Acid (H+) • Enzymes denature at high temperatures. Optimal temperature for Optimal temperature for typical human enzyme enzyme of thermophilic (heat-tolerant) bacteria Rate of reaction 0 20 40 80 100 Temperature (Cº) (a) Optimal temperature for two enzymes Functional shape Denatured Figure 8.18 Heyer 3 Enzymes & Metabolism Enzyme helpers: Cofactors Effect of pH • Each enzyme exhibits peak activity at narrow pH range (pH optimum). • pH optimum reflects the pH of the body fluid in which the enzyme is found. • If pH changed, so is no longer within the enzyme range; reaction will decrease. • Alter shape or charge polarity of enzyme: makes active site available to substrate • Metal ions from dietary minerals. Substrate Active site Enzyme Inhibitors Cofactors and Coenzymes Enzyme • Needed for the activity of specific • Competitive Inhibitors: enzymes. Competitive ß inhibitor • Cofactor: Shape at least partially fits – Attachment of cofactor causes a in the enzymes active site conformational change of active site. È blocks substrate from binding. – Participate in temporary bonds between enzyme and substrate. • Coenzymes: • Allosteric “different shape” Inhibitors: – Organic molecules derived from ß Bind to a different location H20 soluble vitamins. – Transport H+ and small molecules of the enzyme protein from one enzyme to another. — the allosteric site È alters the shape of the protein Allosteric inhibitor È active site no longer fits substrates Figure 8.19 Most enzymes have allosteric regulation Enzymes are Binding Proteins • Proteins change shape when regulatory molecules bind to sites. • I.e., they function by binding to a substrate (ligand) – Allosteric activators (cofactors) and/or inhibitors • Other important biological regulation by means of binding proteins: Allosteric activater stabilizes active from • Receptors for hormones & neurotransmitters Allosteric enyzme Active site with four subunits (one of four) • Transporters across cell membranes • Carriers of low-solubility or fragile substances • Immune recognition Ligand Regulatory site (one of four) Activator Binding site Active form Stabilized active form Oscillation Allosteric inhibitor Binding stabilizes inactive form protein Allosteric activators and inhibitors. In the cell, activators and inhibitors dissociate when at low concentrations. All binding proteins have similar characteristics: The enzyme can then oscillate again. 1. Specificity 2. Saturation Non-functional Inactive form Inhibitor Stabilized inactive form 3. Competitive Inhibition active site Figure 8.20 Heyer 4 Enzymes & Metabolism Coupled Reactions •Catalysis of one reaction allows the catalysis of a second reaction by a Product Coupled Reactions: Bioenergetics different active site on the same enzyme. B Product • Energy transfer from one molecule to another Substrate C couples chemical reactions Reaction 1 A • Exergonic reaction: reaction releases energy • Endergonic reaction: reaction requires energy Enzyme • Coupled bioenergetic reactions: the energy released by the exergonic reaction is used to power the Substrate endergonic reaction. Y Product Reaction 2 Z •Both reactions must occur for either to occur. Bioenergetics Free Energy & Entropy • More free energy (higher G) • Flow of energy in living systems obeys: • Less stable • At maximum stability • 1st law of thermodynamics: • Greater work capacity – The system is at equilibrium – Energy can be transformed, but it cannot be created or destroyed. A spontaneous change • The released free energy • 2nd law of thermodynamics: can be harnessed to do work – Energy transformations increase entropy (degree of . disorganization of a system). – Only free energy (energy in organized state) can be used • Less free energy (lower G) to do work. • More stable • Less work capacity (a) Gravitational motion. (b) Diffusion. Molecules (c) Chemical reaction. • Systems tend to go from states of higher free energy to states of Objects move spontaneously in a drop of dye diffuse In a cell, a sugar lower free energy. from a higher altitude to a until they are randomly molecule is broken lower one. dispersed. down into simpler Figure 8.5 molecules. Exergonic and Endergonic reactions in metabolism Coupled Reactions: Bioenergetics • An exergonic reaction • An endergonic reaction • Cells must maintain highly – Proceeds with a net release – Is one that absorbs free organized, low-entropy state at of free energy and is energy from its the expense of free energy. spontaneous surroundings and is – Cells cannot use heat for energy. nonspontaneous • Energy released in exergonic Reactants Products reactions used to drive endergonic reactions. Amount of energy Amount of – Require energy released in released energy exergonic reactions (ATP) to be (∆G <0) released Energy Energy (∆G>0) directly transferred to chemical- Products Reactants Free energy Free energy bond energy in the products of endergonic reactions. Progress of the reaction Progress of the reaction (a) Exergonic reaction: energy released (b) Endergonic reaction: energy required Figure 8.6 Heyer 5 Enzymes & Metabolism Endergonic and Exergonic The Role of ATP: Reactions Energy transfer from one molecule to • Endergonic: – Chemical reactions that another couples chemical reactions require an input of energy to make reaction “go.” • Products must contain A-P-P~P + glucose --> more free energy than reactants. glucose-6~phosphate + A-P-P • Exergonic: – Convert molecules with • Exergonic