Enzyme Kinetics

General Properties of Enzymes

Enzymes are biological catalysts that accelerate chemical reactions without being consumed. They exhibit high specificity for their substrates and operate under mild physiological conditions.

• Specificity: Enzymes recognize and bind specific substrates, often through precise molecular interactions.

• Efficiency: Enzymes can increase reaction rates by factors of 106 or more.

• Regulation: Enzyme activity can be modulated by various mechanisms, including allosteric regulation and covalent modification.

• Example: Hexokinase catalyzes the phosphorylation of glucose in glycolysis.

Single Substrate Enzyme Features

Single substrate enzymes catalyze reactions involving one substrate molecule. Their kinetics are often described by the Michaelis-Menten model.

• Substrate Binding: The substrate binds to the enzyme's active site, forming an enzyme-substrate complex.

• Product Formation: The complex undergoes a chemical transformation, releasing the product.

Transition State Stabilization

Enzymes lower the activation energy by stabilizing the transition state, the high-energy intermediate between reactants and products.

• Transition State: The configuration with the highest energy along the reaction pathway.

• Stabilization: Enzymes provide an environment that favors the transition state, often through hydrogen bonding, electrostatic interactions, or induced fit.

Michaelis-Menten Equation and Kinetic Parameters

The Michaelis-Menten equation describes the rate of enzymatic reactions as a function of substrate concentration.

• Equation:

• Plot: The Michaelis-Menten plot is hyperbolic, showing velocity versus substrate concentration.

• Key Parameters: Km (Michaelis constant), Vm (maximal velocity), kcat (turnover number), and kcat/Km (catalytic efficiency).

• Lineweaver-Burk Plot: A double reciprocal plot linearizes the Michaelis-Menten equation.

Pre-Steady-State and Steady-State Kinetics

Pre-steady-state kinetics examines the initial formation of enzyme-substrate complexes, while steady-state kinetics analyzes the reaction once intermediate concentrations are constant.

• Pre-Steady-State: Early phase, before equilibrium is reached.

• Steady-State: The rate of formation and breakdown of the enzyme-substrate complex is balanced.

Allosteric Activators and Inhibitors

Allosteric enzymes are regulated by molecules that bind at sites other than the active site, affecting activity.

• Allosteric Activator: Increases enzyme activity, often shifting the Michaelis-Menten plot to lower Km or higher Vm.

• Allosteric Inhibitor: Decreases activity, shifting the plot to higher Km or lower Vm.

Enzyme Mechanisms

Exergonic vs. Endergonic Reactions

Enzyme-catalyzed reactions can be exergonic (energy-releasing) or endergonic (energy-consuming), as shown in reaction coordinate diagrams.

• Exergonic: ; spontaneous.

• Endergonic: ; non-spontaneous.

Proximity Effect

Enzymes increase reaction rates by bringing substrates into close proximity and proper orientation.

• Effect: Reduces entropy and increases effective concentration of reactants.

Lock-and-Key vs. Induced-Fit Models

These models describe how substrates bind to enzymes.

• Lock-and-Key: Substrate fits exactly into the active site.

• Induced-Fit: Enzyme changes shape upon substrate binding for optimal interaction.

Serine Protease Catalysis and Specificity

Serine proteases use a catalytic triad (Ser, His, Asp) to hydrolyze peptide bonds.

• Catalytic Triad: Serine acts as nucleophile, histidine as base, aspartate stabilizes histidine.

• Substrate Binding: Specificity determined by the shape and charge of the binding pocket.

• Example: Chymotrypsin cleaves peptide bonds adjacent to aromatic residues.

Enzyme Inhibition

Types of Inhibition

Enzyme inhibitors reduce activity by different mechanisms.

• Competitive: Inhibitor binds active site; increases Km, no change in Vm.

• Uncompetitive: Inhibitor binds only to enzyme-substrate complex; decreases both Km and Vm.

• Noncompetitive: Inhibitor binds elsewhere; decreases Vm, no change in Km.

Mechanism-Based Inhibitors

These inhibitors mimic the substrate and irreversibly inactivate the enzyme by forming a covalent bond.

• Distinction: Unlike reversible inhibitors, mechanism-based inhibitors permanently modify the enzyme.

Cofactors

Essential Ions vs. Coenzymes

Cofactors are non-protein molecules required for enzyme activity.

• Essential Ions: Metal ions (e.g., Mg2+, Zn2+) that assist in catalysis.

• Coenzymes: Organic molecules; can be prosthetic groups (tightly bound) or cosubstrates (loosely bound).

Types of Coenzymes

• Prosthetic Groups: Permanently attached to enzyme (e.g., FAD).

• Cosubstrates: Transiently associated (e.g., NAD+).

ATP in Group Transfer Reactions

ATP acts as a cofactor by transferring phosphate groups in metabolic reactions.

• Example: Phosphorylation of glucose by hexokinase.

NAD(P)+/NAD(P)H and FAD/FADH2 in Redox Reactions

These cofactors participate in oxidation-reduction reactions by accepting or donating electrons.

• NAD(P)+: Accepts electrons, becomes NAD(P)H.

• FAD: Accepts electrons, becomes FADH2.

Monosaccharides

Functions of Carbohydrates

Carbohydrates serve as energy sources, structural components, and signaling molecules.

• Energy: Glucose is a primary energy source.

• Structure: Cellulose in plants, chitin in fungi.

• Signaling: Glycoproteins in cell recognition.

Classification of Saccharides

• Monosaccharides: Single sugar units (e.g., glucose).

• Disaccharides: Two monosaccharides linked (e.g., sucrose).

• Oligosaccharides: 3-10 monosaccharides.

• Polysaccharides: Many monosaccharides (e.g., starch).

Aldoses vs. Ketoses

Monosaccharides are classified by their carbonyl group.

• Aldoses: Have an aldehyde group (e.g., glucose).

• Ketoses: Have a ketone group (e.g., fructose).

Fischer Projections and Chirality

Fischer projections are 2D representations of monosaccharides, showing stereochemistry.

• Chirality: Monosaccharides have multiple chiral centers.

• Example: D- and L-glucose differ at the chiral center furthest from the carbonyl.

Stereochemical Terms

• Stereoisomer: Same formula, different spatial arrangement.

• Enantiomer: Mirror images.

• Diastereomer: Not mirror images.

• Epimer: Differ at one chiral center.

Cyclization of Monosaccharides

Monosaccharides cyclize to form hemiacetals (aldoses) or hemiketals (ketoses).

• Anomeric Carbon: New chiral center formed during cyclization.

• Haworth Projection: Shows cyclic structure.

• Pyranose: Six-membered ring.

• Furanose: Five-membered ring.

Disaccharides

Glycosidic Bonds and Connectivity

Disaccharides are formed by glycosidic bonds between monosaccharides.

• Reducing Sugar: Has a free anomeric carbon.

• Non-Reducing Sugar: Both anomeric carbons are involved in the bond.

• Example: Sucrose (non-reducing), lactose (reducing).

Polysaccharides and Glycoproteins

Functional Roles and Structure of Polysaccharides

Polysaccharides serve as energy storage and structural materials.

• Starch: Storage in plants; composed of amylose and amylopectin.

• Glycogen: Storage in animals; highly branched.

Glycoproteins

Proteins with covalently attached carbohydrate chains.

• O-linked: Carbohydrate attached to serine/threonine.

• N-linked: Carbohydrate attached to asparagine.

• Function: Cell recognition, signaling.

ABO Blood Classification

Blood types are defined by specific glycoprotein antigens on red blood cells.

• Donor/Recipient: Compatibility depends on antigen presence.

Nucleic Acids

Components of a Nucleotide

Nucleotides are composed of a nucleobase, a pentose sugar, and a phosphate group.

• Nucleobase: Purine (A, G) or pyrimidine (C, T, U).

• Nucleoside: Nucleobase + sugar.

• Nucleotide: Nucleoside + phosphate.

Structures of Nucleotides

• Purines: Adenine (A), Guanine (G).

• Pyrimidines: Cytosine (C), Thymine (T), Uracil (U).

• Ribose: Sugar in RNA.

• Deoxyribose: Sugar in DNA.

• Phosphoanhydride: Between phosphate groups.

• Phosphoester: Between phosphate and sugar.

DNA vs. RNA Nucleotides

• DNA: Contains deoxyribose and T.

• RNA: Contains ribose and U.

DNA and RNA: Structures and Functions

Linking Nucleotides

Nucleotides are linked by phosphodiester bonds, forming polynucleotide chains.

• Directionality: 5' to 3' end.

• Watson-Crick Base Pairs: A-T (or A-U in RNA), G-C.

• Hydrogen Bonds: A-T: 2 bonds; G-C: 3 bonds.

Major Classes of RNA and Functions

• mRNA: Messenger RNA; carries genetic information.

• tRNA: Transfer RNA; brings amino acids to ribosome.

• rRNA: Ribosomal RNA; structural and catalytic role in ribosome.

Secondary Structures of RNA

• Hairpin: Stem-loop structure.

• Bulge: Unpaired nucleotides in a loop.

• Pseudoknot: Complex folding pattern.

Additional info: Where original notes were brief, academic context and examples were added for completeness.