Enzymes and Catalysis

• Enzymes are proteins functioning as catalysts.

Enzymes and Catalysis

• Enzymes' catalytic ability is a key function in living cells.
• Without catalysts, cellular reactions would be too slow to sustain life.
• Most enzymes are globular proteins, except for some RNA molecules.
• Enzymes are efficient catalysts.
• Some enzymes can increase reaction rates by a factor of 10^20.
• Enzymes possess enormous catalytic power, high specificity in the reactions they catalyze, and their activity can be regulated.
• Enzyme-catalyzed reactions carry out significant organic reactions like ester hydrolysis, alcohol oxidation, and amide formation.
• Enzymes allow reactions to proceed under mild pH and temperature conditions.
• Enzyme-catalyzed reactions can occur in seconds.
• Carbonic anhydrase speeds up carbon dioxide removal by combining it with water to form carbonic acid, processing 36 million molecules per minute.

Catalytic Efficiency

• Catalysts increase the rate of chemical reactions without being consumed.
• Catalysts participate in reactions but aren't permanently changed and can be used repeatedly.
• Enzymes, like other catalysts, lower the activation energy for reactions, enabling them to reach equilibrium quicker.

Specificity of Enzymes

• Enzymes exhibit specificity in the type of reaction and the substance involved.
• Strong acids catalyze the hydrolysis of any amide or ester, and the dehydration of any alcohol.
• Urease catalyzes the hydrolysis of urea.
• Enzymes with absolute specificity act on only one substance.
• Relative specificity enzymes act on structurally related substances like lipases on lipids, proteases on proteins, and phosphatases on phosphate esters.
• Stereochemical specificity enzymes act on just one of two possible enantiomers, such as D-amino acid oxidase on D-amino acids.

Regulation of Enzyme Activity

• A relatively small subset of potential reactions in a cell occurs due to the enzymes present.
• Cells manage reaction rates and product amounts by regulating enzyme activity.

Enzyme Nomenclature

• Early enzymes were named with an "-in" ending to denote their protein nature (e.g., pepsin, trypsin, chymotrypsin).
• A systematic approach to naming enzymes is the Enzyme Commission (EC) system with the International Union of Biochemistry and Molecular Biology; based on the reaction they catalyze.
• Enzymes are classified into six major groups by the reaction they catalyze.
• Each enzyme designation includes the substrate, functional group, and reaction type catalyzed.
• EC names end in -ase.
• Common names for enzymes add "ase" to the substrate name or the type of reaction.

Enzymes and Substrates

• Enzyme examples: decarboxylase, phosphatase, peptidase, esterase.
• Reactions catalyzed: formation of ester linkages, removal of carboxyl groups, hydrolysis of peptide and phosphate ester linkages.

Enzyme Cofactors

• Conjugated proteins need specific nonprotein molecules or metal ions (prosthetic groups) for function.
• If the nonprotein part is firmly bound, it's a true prosthetic group.
• If the nonprotein component is loosely bound, it is called a cofactor.
• An organic substance cofactor is known as a coenzyme.
• A cofactor can be an inorganic ion like Mg2+, Zn2+, or Fe2+.
• The protein part of an enzyme is called an apoenzyme.

Cofactor Examples

• Apoenzyme + cofactor (coenzyme or inorganic ion) yields an active enzyme.
• Vitamins are often precursors to coenzymes.
• Biotin becomes biocytin, which functions in carboxyl group removal or transfer.
• Folacin is modified to tetrahydrofolic acid to function as a one-carbon group transfer agent.
• Niacin forms nicotinamide adenine dinucleotide (NAD+) for hydrogen transfer.

Enzymatic Action

• Enzymes differ in structure and specificity, but a general mechanism explains their catalytic behavior which has been widely accepted.
• Enzymes and substrates interact at a small region called the active site.
• Substrate binding to the active site through intermolecular forces forms an enzyme-substrate (ES) complex.
• The conversion of substrate (S) to product (P) occurs once the complex is formed.
• The chemical transformation of the substrate occurs at the active site, helped by functional groups of the enzyme.
• Post-conversion, the product is released, freeing the enzyme to react with another substrate.

Lock and Key Theory vs Flexible Enzymes

• The lock-and-key theory explains the specificity of enzymes.
• Enzyme surfaces fit specific substrates in an active site to create an ES complex.
• The lock-and-key theory requires rigid enzyme conformations, at odds with research indicating enzyme flexibility.

Enzymatic Activity Numbers

• Enzymatic activity measures an enzyme's ability to speed up a reaction.
• The turnover number is the number of product molecules formed by one enzyme molecule per minute.
• Turn over number for carbonic anhydrase has a turnover number of 36 million molecules per minute.
• Most enzymes' turnover numbers are closer to 1000 molecules per minute.
• Enzyme assays are experiments that assess enzyme activity.
• Blood enzyme assays are common in clinical labs.
• These can be done by observing the rate of color change or pH change.

Factors Affecting Enzyme Activity

• The concentration of an enzyme that is typically low compared to substrate, increases reaction rate.
• Reaction rate is proportional to enzyme concentration; doubling [E] doubles the rate.

Substrate Concentrations

• Increasing substrate concentration increases the reaction rate to a maximum (Vmax).
• The maximum rate is reached when the enzyme is saturated with substrate.
• The rate of enzyme-catalyzed reactions increases with temperature.
• Enzymes denature beyond a certain temperature, due to their nature as proteins.
• Each enzyme-catalyzed reaction shows maximum activity at an optimum temperature, usually, 25-40°C.

pH Effects

• Altering pH influences the acidic and basic side chains in enzymes.
• Enzymes denature at pH extremes.
• Pickling uses acetic acid to deactivate bacterial enzymes.
• It will be effective at preserving food.
• Most enzymes operate best near pH 7, however some enzymes are more effective at low pH, e.g., pepsin in the stomach.

Enzyme Inhibition

• Enzyme inhibitors lower the rate of enzyme-catalyzed reactions.
• Many poisons and medicines act as enzyme inhibitors.
• Some cells contain substances that inhibit specific enzymes.
• This offers the cell an internal control of metabolism.
• Irreversible inhibition involves an inhibitor forming a covalent bond with an enzyme, inactivating it; an example is cyanide.
• Cyanide blocks cell respiration by interfering with the iron-containing enzyme cytochrome oxidase.
• Sodium thiosulfate is an antidote for cyanide poisoning.
• It converts cyanide into thiocyanate which does not bind to cytochrome.
• Lead and mercury ions binding to -SH groups on enzymes causes denaturation and permanent neurological damage
• Administering chelating agents to bind metal ions to excreted in the urine treats heavy metal poisoning.

Antibiotics As Inhibitors

• Antibiotics inhibit life processes essential to bacteria.
• Interfering with cell wall construction, sulfa drugs and penicillins inhibit transpeptidase.
• Reversible inhibitors bind reversibly to an enzyme; setting up equilibrium and preventing catalysis.
• Shifting the equilibrium removes it.
• There are two types: competitive and noncompetitive.

Competitive Inhibition

• Competitive inhibitors bind to the enzyme's active site and compete with normal substrates.
• Competitive inhibitors are often similar to substrates.
• In competitive inhibition, increasing substrate concentration or decreasing enzyme concentration can reverse equilibrium.

Noncompetitive Inhibition

• Noncompetitive inhibitors binds at a different site from the active site.
• This alters the enzyme's 3D shape, so the substrate no longer fits properly resulting in decreased or no binding with the substrate.
• Noncompetitive inhibitors are often dissimilar to substrates.
• Increasing substrate levels do not surmount the effect of noncompetitive inhibition.

Enzyme Regulation

• Enzymes require sensitive controls.
• Activation of zymogens, allosteric regulation, and genetic control.
• Zymogens are inactive enzyme precursors.
• Certain enzymes in active form could damage the internal structures of cells.
• These enzymes are stored as inactive precursors that are activated when required.
• Zymogen activation usually needs the cleavage of one or more peptide bonds.
• Examples pepsin, trypsin, chymotrypsin, and blood clotting enzymes.

Examples of Zymogens and Active Enzymes

• Chymotrypsinogen converts to chymotrypsin.
• Pepsinogen converts to pepsin.

Allosteric Regulation

• Compounds (modulators) alter enzymes by changing the 3D conformation.
• Modulators either increase (activators) or decrease activity (inhibitors).
• Noncompetitive inhibitors are examples of this.
• Allosteric enzymes have quaternary structures with modulator binding sites and are variable-rate enzymes.
• Feedback inhibition involves an end product inhibiting an earlier step to maintain product levels; this allows for stable concentration of the product to be synthesised.
• Synthesis of end products such as isoleucine from threonine is an example.
• When there is high isoleucine levels it binds to the enzyme threonine deaminase.
• This changes the configuration and then threonine binds poorly to receptor, slowing the production of this enzyme.
• This maintains a balanced level of products to be synthesised.

Genetic Control

• Proteins and enzymes synthesis is under genetic control by nucleic acids.
• Enzyme induction happens when enzymes are synthesized in response to cell need.
• Genetic control and allosteric regulation offer close control of cellular processes.
• A significant role of genetic control is b-galactosidase, an enzyme for lactose hydrolysis in Escherichia coli. Lactose is split into D-galactose and D-glucose.
• With no lactose, there are less b-galactosidase molecules.
• Thousands of enzyme molecules appear upon the presence of lactose.
• Enzyme creation ceases if it's removed.

Enzymes in Clinical Diagnosis

• Enzymes in the bloodstream can indicate tissue damage.
• Measuring enzyme concentrations is a diagnostic tool for conditions like heart, liver, pancreas, and prostate illnesses.
• Isoenzymes are different forms of the same enzyme made by different tissues.
• The enzyme lactate dehydrogenase (LDH) is made of four subunits with 2 different subunit structures. Subunit H is found in heart muscle cells; and subunit M subunit in skeletal muscle cells. LDH has 5 possible subunit combinations which have different properties and are identified separately.
• Each tissue has a unique isoenzyme expression pattern.

Isoenzymes Can Be Used In Diagnostics

• They diagnose a wide range of illnesses.
• Anemias can be identified through the identification of LDH.
• Acute liver conditions: congestive heart condition such as failure, and skeletal and muscle weakness.
• Myocardial infarction can be caused by higher levels of LDH1 and LDH2.
• Possible liver damage can be identified through a high LDH5 count.