Chapter 2 Enzyme
Contents • Properties of enzymes • Structural features of enzymes • Mechanism of enzyme-catalyzed reactions • Kinetics of enzyme-catalyzed reactions • Inhibition of enzymes • Regulation of enzymes • Clinical applications of enzymes • Nomenclature
§ 1.1 General Concepts A + B → C + D • spontaneous reaction only if DG is negative. • at equilibrium if DG is zero. • spontaneously impossible if DG is positive.
Catalyzed reactions • Reactants need to pass over the energy barrier, G+. • Catalysts reduce the activation energy and assist the reactants to pass over the activation energy.
Need for special catalysts Chemical reactions in living systems are quite different from that in the industrial situations because of • Fragile structures of the living systems • Low kinetic energy of the reactants • Low concentration of the reactants • Toxicity of catalysts • Complexity of the biological systems
Enzymes • Enzymes are catalysts that have special characteristics to facilitate the biochemical reactions in the biological systems. • Enzyme-catalyzed reactions take place usually under relatively mild conditions.
§ 1.2 Characteristics Enzyme-catalyzed reactions have the following characteristics in comparison with the general catalyzed reactions: • common features: 2 “do” and 2 “don’t” • unique features: 3 “high”
Common features • Do not consume themselves: no changes in quantity and quality before and after the reactions. • Do not change the equilibrium points: only enhance the reaction rates. • Apply to the thermodynamically allowable reactions • Reduce the activation energy
Unique features • Enzyme-catalyzed reactions have very high catalytic efficiency. • Enzymes have a high degree of specificity for their substrates. • Enzymatic activities are highlyregulated in response to the external changes.
§ 1.3.b High specificity Unlike conventional catalysts, enzymes demonstrate the ability to distinguish different substrates. There are three types of substrate specificities. • Absolute specificity • Relative specificity • Stereospecificity
Absolute specificity Enzymes can recognize only one type of substrate and implement their catalytic functions.
Relative specificity Enzymes catalyze one class of substrates or one kind of chemical bond in the same type.
Stereospecificity Lactate dehydrogenase can recognize only the L-form but the D-form lactate. The enzyme can act on only one form of isomers of the substrates.
§ 1.3.c High regulation • Enzyme-catalyzed reactions can be regulated in response to the external stimuli, satisfying the needs of biological processes. • Regulations can be accomplished through varying the enzyme quantity, adjusting the enzymatic activity, or changing the substrate concentration.
§ 2.1 Active Center • Almost all the enzymes are proteins having well defined structures. • Some functional groups are close enough in space to form a portion called the active center. • Active centers look like a cleft or a crevice. • Active centers are hydrophobic.
Lysozyme Residues (colored ) in the active site come from different parts of the polypeptide chain .
Two essential groups The active center has two essential groups in general. • Binding group: to associate with the reactants to form an enzyme-substrate complex • Catalytic group: to catalyze the reactions and convert substrates into products
§ 2.2 Molecular Components • Simple enzymes: consists of only one peptide chain • Conjugated enzymes: holoenzyme = apoenzyme + cofactor (protein) (non-protein) • Cofactors: metal ions; small organic molecules
Metal ions • Metal-activated enzyme:ions necessary butloosely bound. Often found in metal-activated enzyme. • Metalloenzymes: Ions tightly bound. • Particularly in the active center, transfer electrons, bridge the enzyme and substrates, stabilize enzyme conformation, neutralize the anions.
Organic compounds • Small size and chemically stable compounds • Transferring electrons, protons and other groups • Vitamin-like or vitamin-containing molecule
Coenzymes • Loosely bind to apoenzyme. Be able to be separated with dialysis. • Accepting H+ or group and leaving to transfer it to others, or vise versa. Prosthetic groups • Tightly bind through either covalent or many non-covalent interactions. • Remained bound to the apoenzyme during the course of reaction.
To understand the molecular details of the catalyzed reaction. • Proximity and orientation arrangement • Multielement catalysis • Surface effect
Lock-and-key model Both E and S are rigid and fixed, so they must be complementary to each other perfectly in order to have a right match.
Induced-fit model The binding induces conformational changes of both E and S, forcing them to get a perfect match.
Hexokinase catalyzing glycolysis • Hexokinase, the first enzyme in the glycolysis pathway, converted glucose to glucose-6-phosphate with consuming one ATP molecule. • Two structural domains are connected by a hinge. • Upon binding of a glucose molecule, domains close, shielding the active site for water.
Initial velocity • The reaction rate is defined as the product formation per unit time. • The slope of product concentration ([P]) against the time in a graphic representation is called initial velocity. • It is of rectangular hyperbolic shape.
Intermediate state Forming an enzyme-substrate complex, a transition state, is a key step in the catalytic reaction. initial intermediate final
Rate constants • K1 = rate constant for ES formation • K2 = rate constant for ES dissociation • K3 = rate constant for the product released from the active site
§ 4.2 Michaelis-Menten Equation • The mathematical expression of the product formation with respect to the experimental parameters • Michaelis-Menten equation describes the relationship between the reaction rate and substrate concentration [S].
Assumptions • [S] >> [E], changes of [S] is negligible. • K2 is negligible compared with K1. • Steady-state: the rate of E-S complex formation is equal to the rate of its disassociation (backward E + S and forward to E + P)
Describing a hyperbolic curve. Km is a characteristic constant of E [S] << Km 时，v ∝ [S] [S] >> Km 时，v ≈ Vmax
Significance of Km • the substrate concentration at which enzyme-catalyzed reaction proceeds at one-half of its maximumvelocity • Km is independent of [E]. It is determined by the structure of E, the substrate and environmental conditions (pH, T, ionic strength, …)
Km is a characteristic constant of E. • The value of Km quantifies the affinity of the enzyme and the substrate under the condition of K3 << K2. The larger the Km，the smaller the affinity.