BCH 351 Exam 2 Study Guide: Enzymes, Carbohydrates, and Nucleic Acids

Enzyme Kinetics

General Properties of Enzymes

Enzymes are biological catalysts that accelerate chemical reactions in living organisms. They are highly specific for their substrates and operate under mild physiological conditions.

• Specificity: Enzymes typically act on a single substrate or a group of closely related substrates.

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

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

Single Substrate Enzyme Kinetics

Single substrate enzymes follow characteristic kinetic behavior, often described by the Michaelis-Menten model.

• Substrate Binding: The enzyme binds to its substrate to form an enzyme-substrate (ES) complex.

• Product Formation: The ES complex is converted to product, releasing the enzyme for another catalytic cycle.

Transition State Stabilization

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

• Transition State: The configuration of atoms at the maximum energy point along the reaction coordinate.

• Stabilization: Enzymes provide an environment that favors the transition state, often through specific interactions such as hydrogen bonding or electrostatic effects.

Michaelis-Menten Equation

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

• Equation:

$v = \frac{V_{max}[S]}{K_m + [S]}$

• Plot: The plot of reaction velocity (v) versus substrate concentration ([S]) is hyperbolic.

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

Pre-Steady-State and Steady-State Kinetics

• Pre-Steady-State: The initial phase of the reaction before the ES complex concentration becomes constant.

• Steady-State: The phase where the formation and breakdown of the ES complex are balanced, and the reaction rate is constant.

Michaelis-Menten and Lineweaver-Burk Plots

• Michaelis-Menten Plot: Plots v versus [S]; used to estimate Vmax and Km.

• Lineweaver-Burk Plot: Double reciprocal plot ($\frac{1}{v}$ vs. $\frac{1}{[S]}$) linearizes the data for easier parameter determination.

$\frac{1}{v} = \frac{K_m}{V_{max}} \cdot \frac{1}{[S]} + \frac{1}{V_{max}}$

Allosteric Activators and Inhibitors

• Allosteric Activator: A molecule that increases enzyme activity by binding to a site other than the active site.

• Allosteric Inhibitor: A molecule that decreases enzyme activity by binding to an allosteric site.

• Effect on Plots: Allosteric enzymes often show sigmoidal (S-shaped) kinetics rather than hyperbolic.

Enzyme Mechanisms

Exergonic vs. Endergonic Reactions

Enzyme-catalyzed reactions can be classified based on their free energy changes.

• Exergonic: Reactions that release energy ($\Delta G < 0$).

• Endergonic: Reactions that require energy input ($\Delta G > 0$).

• Reaction Coordinate Diagrams: Visualize the energy changes during a reaction, including activation energy and transition state.

Proximity Effect

• Definition: Enzymes increase reaction rates by bringing substrates into close proximity and correct orientation for reaction.

Substrate Binding Models

• Lock-and-Key Model: The enzyme active site is complementary in shape to the substrate.

• Induced-Fit Model: The enzyme changes shape upon substrate binding to better fit the substrate.

Transition State Stabilization

• Chemical Aspects: Enzymes may use hydrogen bonds, ionic interactions, or covalent bonds to stabilize the transition state.

Serine Protease Catalysis

• Catalytic Triad: A set of three amino acids (Serine, Histidine, Aspartate) that work together to catalyze peptide bond hydrolysis.

• Substrate Binding: Specificity pockets in the enzyme determine which peptide bonds are cleaved.

• Chymotrypsin Mechanism: Involves nucleophilic attack by serine, formation of a tetrahedral intermediate, and stabilization by the oxyanion hole.

Enzyme Inhibition

Types of Inhibition

• Competitive Inhibition: Inhibitor binds to the active site, preventing substrate binding. Increases apparent Km, Vmax unchanged.

• Uncompetitive Inhibition: Inhibitor binds only to the ES complex. Decreases both Km and Vmax.

• Noncompetitive Inhibition: Inhibitor binds to both the enzyme and ES complex. Vmax decreases, Km unchanged.

Type	Binding Site	Km	Vmax
Competitive	Active site	Increases	Unchanged
Uncompetitive	ES complex	Decreases	Decreases
Noncompetitive	Enzyme or ES	Unchanged	Decreases

Lineweaver-Burk Plots for Inhibition

• Competitive: Lines intersect at y-axis.

• Uncompetitive: Lines are parallel.

• Noncompetitive: Lines intersect left of y-axis.

Mechanism-Based Inhibitors

• Definition: Inhibitors that bind irreversibly to the enzyme, often by mimicking the transition state or forming a covalent bond.

• Distinguishing Feature: They inactivate the enzyme through a chemical reaction, not just by occupying the active site.

Cofactors

Types of Cofactors

• Essential Ions: Metal ions required for enzyme activity (e.g., Mg2+, Zn2+).

• Coenzymes: Organic molecules that assist enzymes. Two types:

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

  • Prosthetic Groups: Tightly bound to the enzyme (e.g., FAD).

ATP as a Cofactor

• Role: ATP acts as a phosphate group donor in group transfer reactions, providing energy for many cellular processes.

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

• NAD(P)+/NAD(P)H: Electron carriers in oxidation-reduction reactions.

• FAD/FADH2: Another class of electron carriers, often involved in dehydrogenase reactions.

Monosaccharides

Functions of Carbohydrates

• Energy Storage: Glucose, glycogen, and starch serve as energy reserves.

• Structural Roles: Cellulose in plants, chitin in arthropods.

• Cell Recognition: Glycoproteins and glycolipids on cell surfaces.

Classification of Carbohydrates

• Monosaccharides: Simple sugars (e.g., glucose, fructose).

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

• Oligosaccharides: 3-10 monosaccharide units.

• Polysaccharides: Many monosaccharide units (e.g., starch, cellulose).

Aldoses vs. Ketoses

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

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

Fischer Projections and Chirality

• Fischer Projection: A two-dimensional representation of three-dimensional sugar structures.

• Chirality: Monosaccharides have chiral centers, leading to stereoisomers.

Stereochemistry of Monosaccharides

• Stereoisomer: Molecules with the same formula but different spatial arrangement.

• Enantiomer: Non-superimposable mirror images.

• Diastereomer: Stereoisomers that are not mirror images.

• Epimer: Diastereomers differing at only one chiral center.

Cyclization of Monosaccharides

• Hemiacetal Formation: Aldoses cyclize to form hemiacetals.

• Hemiketal Formation: Ketoses cyclize to form hemiketals.

• Anomeric Carbon: The new chiral center formed during cyclization.

• Haworth Projection: A common way to depict cyclic sugars.

• Pyranose: Six-membered ring form.

• Furanose: Five-membered ring form.

Disaccharides

Glycosidic Bonds and Disaccharide Formation

• Glycosidic Bond: Covalent bond linking two monosaccharides via their anomeric carbon.

• Reducing vs. Non-Reducing Sugars: Reducing sugars have a free anomeric carbon; non-reducing do not.

• Examples: Sucrose (glucose + fructose), lactose (glucose + galactose).

Polysaccharides and Glycoproteins

Polysaccharide Structure and Function

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

• Glycogen: Storage polysaccharide in animals; highly branched.

• Cellulose: Structural polysaccharide in plants; linear chains of glucose.

Glycoproteins

• O-linked Glycoproteins: Carbohydrate attached to the oxygen atom of serine or threonine side chains.

• N-linked Glycoproteins: Carbohydrate attached to the nitrogen atom of asparagine side chains.

• Functional Roles: Cell-cell recognition, immune response, structural support.

ABO Blood Classification

• Basis: Determined by specific oligosaccharide structures on red blood cell surfaces.

• Blood Donors and Recipients: Compatibility depends on the presence or absence of A and B antigens.

Nucleic Acids

Components of Nucleotides

• Nucleobase: Purine (adenine, guanine) or pyrimidine (cytosine, thymine, uracil).

• Nucleoside: Nucleobase + sugar (ribose or deoxyribose).

• Nucleotide: Nucleoside + phosphate group.

Structures of Nucleotides

• Purines: Adenine (A), Guanine (G).

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

• Phosphoanhydride Bond: High-energy bond between phosphate groups (e.g., in ATP).

• Phosphoester Bond: Bond between phosphate and sugar.

• DNA vs. RNA: DNA contains deoxyribose and T; RNA contains ribose and U.

DNA and RNA: Structures and Functions

Linking Nucleotides

• Phosphodiester Bond: Connects the 3' carbon of one sugar to the 5' phosphate of the next.

• Directionality: Nucleic acids are synthesized and read from 5' to 3'.

Base Pairing

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

• Hydrogen Bonds: A-T/U pairs have 2 hydrogen bonds; G-C pairs have 3.

Classes and Structures of RNA

• Major Classes: mRNA (messenger), tRNA (transfer), rRNA (ribosomal).

• Secondary Structures: Stem-loops, hairpins, bulges, and pseudoknots.

Example: The double helix structure of DNA is stabilized by complementary base pairing and the sugar-phosphate backbone.

Additional info: Where specific structures or drawings are requested (e.g., Haworth projections, nucleotide structures), students should refer to textbook figures or lecture slides for visual representations.