Enzymes: Biological Catalysts

• Enzymes are defined as biological catalysts or proteins that exhibit catalytic effects in reactions occurring in biological systems.
• Enzymes accelerate biological reactions that are thermodynamically possible under physiological conditions by lowering the activation energy.
• Reactions can be accelerated by 10^18 times or more.

Substrate and Product

• The molecule acted upon by an enzyme (E) is called the substrate (S).
• The molecule resulting from the reaction is called the product (P).

Enzyme Nomenclature:

• Enzymes were initially named in a non-systematic manner.
• The suffix "-in" was initially added to names like trypsin, pepsin, and ptyalin.
• Later, the suffix "-ase" was added to the end of the substrate upon which the enzymes acted.
  • Protease: An enzyme that hydrolyzes proteins
  • Urease: An enzyme that breaks down urea
  • Lipase: An enzyme that hydrolyzes lipids
• Enzymes were also named based on the type of reaction they catalyzed.
  • Examples include dehydrogenase, oxidase, and decarboxylase.
• These naming methods could lead to confusion, as one enzyme might have multiple names or different enzymes might share the same name.
• A systematic classification was later adopted to address this issue.

Systematic Classification:

• The International Union of Biochemistry and Molecular Biology (IUBMB) established enzyme nomenclature based on the type and mechanism of the reaction catalyzed (1964; last revised 1984).
• Enzymes are divided into 6 main groups, each with 4-13 subgroups.
• An enzyme commission (EC) assigns a 4-digit code number to each enzyme based on the reaction it catalyzes and its mechanism.
• The first digit in the code indicates the enzyme's main class. For example, hexokinase starts with 2 (EC 2.7.1.1).

Six Main Enzyme Classes:

• Oxidoreductases: Catalyze oxidation-reduction reactions.
• Transferases: Catalyze the transfer of chemical groups.
• Hydrolases: Catalyze the cleavage of bonds.
• Lyases: Catalyze cleavage of bonds.
• Isomerases: Catalyze the geometric isomers.
• Ligases: Catalyze the formation of bonds.

General Properties of Enzymes:

Biological Catalysts

• They are biological catalysts from biological sources.
• Their activities can be measured in vitro, using units U or katal (K).

Enzyme Activity (Unit Definition)

• One unit of enzyme activity is defined as the amount of enzyme that catalyzes the formation of 1 μmol of product (P) under standardized conditions (e.g., 25 °C, pH 7) within a specified time (e.g., 1 minute).
  • Katal: 1 mol [S]/second

Specific Catalysts

• Enzymes are specific due to their ability to catalyze only certain reactions

Not Passive

• Enzymes form an enzyme-substrate (ES) complex by binding to the substrate.
• Enzymes have specific binding regions on their surface called active sites where the substrate binds.
• After catalyzing a reaction, the enzyme is released unchanged to bind other substrates.
• Enzymes typically have high molecular weights, while substrates are relatively small molecules.

Active Site Binding Models:

• Two models are proposed for the E and S binding at the active site
  • Fisher's "lock-and-key" model: The active site of the enzyme is pre-shaped to fit the substrate.
  • Koshland's "induced fit" model: The active site changes shape upon substrate binding to fit the substrate properly.

Protein Composition:

• Enzymes are proteins, except for some RNAs that exhibit enzymatic activity.
• They exhibit all the structural properties attributed to proteins.

Simple Enzymes

• Some enzymes are made up of protein only and can exhibit catalytic activity. The best examples are digestive enzymes and the urease enzyme that break down urea.

Complex Enzymes

• Many require an additional molecule besides the protein part to exhibit activity.
  • These enzymes consist of two parts.
    • Protein part + Vitamins(B group, vitamin C)
    • Organic molecules (NAD, FAD)
    • Minerals or metal ions, are needed for the holoenzyme.
  • The protein part of these enzymes is called the apoenzyme.
  • While the non-protein part is called the coenzyme (organic group and vitamins) or cofactor (mineral and metal ions).
• Coenzymes bind loosely to the enzyme
  • If the coenzyme is tightly bound to the enzyme, it is called a prosthetic group.
    • Apoenzyme + Coenzyme = Holoenzyme

Low Concentrations

• Enzymes are present in very low concentrations in organisms.
• Their activities are measured by the rate of the reactions they catalyze.

Catalytic Effect

• Enzyme-catalyzed reactions occur more readily than non-catalyzed reactions.
  • One enzyme accelerates the conversion of a substance to a product by 10^3 - 10^18 times.
  • Enzymes enable the transformation of millions of molecules per minute.

Genetically Determined

• Genetically controlled due to protein structure.

Regulated Activity

• The enzyme that is active in the chemical reaction can be activated or inhibited depending on the cell's need for that product.

Activation Energy:

• All chemical reactions have an energy barrier between reactants and products.
• This barrier, called the free energy of activation, is the difference in energy between the reactants and the high-energy intermediate formed during the formation of products.
• For a reaction to occur, the reacting molecules must have enough energy to overcome the energy barrier of the transition state.
• Enzymes lower the reaction activation energy.

Factors Affecting Enzyme Activity:

• Incubation Time
• Temperature
• Enzyme Concentration
• pH
• Substrate Concentration
• Inhibitor Effect
• Allosteric Effect
• Covalent Modification
• Limited Proteolysis
• Enzyme Production and Degradation (Enzyme Turnover)

Incubation Time:

• The rate of an enzyme reaction is determined by the amount of product produced over a specific time.
• In region A, there is a linear relationship between time and product concentration.
• Vo (initial rate) is initially constant.
• Deviations from the constant rate can occur due to enzyme denaturation by heat, [S] decrease, or accumulation of inhibitory substances.

Substrate Concentration:

• Assuming the enzyme amount and other reaction conditions are constant, when the Substrate conc. is increased, the reaction rate first shows an increase.
• If the Substrate conc. continues to increase, this rapid increase slows down, eventually stabilizing at a certain level.
• The maximum rate (Vmax) is reached when the enzyme is saturated

Enzyme Concentration:

• The presence of a sufficient amount of substrate in an enzymatic reaction increases the rate of the reaction to be directly proportional to the enzyme concentration.

pH Effect:

• Enzymes have an optimum pH value or pH range at which their activities are maximum
• Activity decreases at higher or lower pH.
• Extremes in pH can cause ionization of amino and carboxyl groups, leading to protein denaturation.
• Changes in charge can cause the binding of Mg+2, Mn+2,Fe+3 to be difficult.

Temperature Effect:

• As temperature increases, the reaction rate increases until a maximum rate is reached, assuming other conditions are constant.
• The enzymes break down at high temperatures.
• There is an optimum temperature. The optimum temperature for the human body is 35-40 °C.
• Enzymes can resist to 70°C.

Substrate Concentration

• If the amount is constant, when the subs. is increased, the reaction rate will increase.
• When the enzyme saturation occurs, the catalytic activity of the enzyme will arrive at its maximum level.
• (S) versus reaction rate graph forms a hyperbola.
• Enzyme graph can plateau given full saturation of the subs.

Michaelis-Menten constant equation:

v = Vmax [S] /KM + [S]

Michealis Menten:

• Km: Indicates the affinity to the substrates. A low Km value indicates higher affinity, and a high Km value indicates lower affinity.
• Vmax: Represents the total enzyme amount able to enter rection.
• Line weaver Burk graph is often used when using the Michaelis Menten equation 1/v = Km/Vmax[S] +1/Vmax

Michaelis-Menten Equation Transformations:

• By using experimental data, enzymes Km and Vmax values can be calculated with linear graphs.
• The Line Weaver Burk equation is useful to transform MM equations.

Inhibitors:

• Substances that are not substrates of the enzyme can bind to the emzymes.
• A-Irreversible Enzyme Inhibition- when the molecule binds, it will never detach
• B-Reversible Enzyme Inhibition
  • a)Competitive Enzyme Inhibition- competes with the natural substrates
  • b)Noncompetitive Enzyme Inhibition
    • Type I - Binds to somewhere but the active site
    • Type 2 (Uncompetitive)- Only binds to enzymes-substrate complexes

A-Irreversible Inhibitors

• This type of inihibitions is when an inhibitor covalently binds at its active site,
• It inhibits the ability of enzymes to bond subs, in the active areas.
• Reactions that use serine cannot occur.
• These types of inhibitors are often found with cyanide and dinitrophenol

B-Reversible Inhibitors

competetive

• Can be reverted
• The structures are typically of similar build
• They occupy sites and dont allow enzymes to bond
• Adding more substrate can increase the rate of activity.

Non competetive

• Will bind somewhere else and change the proteins structure.
• The Km stays relatively the same
• Lower Vmax values

Type II

• Will only bond to enzyme subs complexes.
• It will alter the sites of the subs to change them for the reaction
• Vmax and KM both shift
• Drugs use Competitive inhibition*:
• sulfonamides are typically PABA analogs that act as antibacterials and stop folate synthesis
• ethanol is used to treat alcohol poisoning and methanol inhibitions. It is perferable for ethenol to bind

Allosteric Effect:

• Allosteric Enzymes: Are enzymes on top of active sites and have regulatory sections. - They all have more effactors and more modulators
  • Activator, positive catalyst
  • Inhibitor, negative catalyst
• These will alter an enzymes structure and will create different shapes
• *Allosteric enzymes do not give hyperbolas, they give sigmoidal graphs

Allosteric enzyme features:

• Sigmoidal

• • effectors decrease value
• • effectors increase value

• tend to have a protien of 4, and work well in metalolic reactions.

• Allosteric enzymes can be regulated with feedback inhibition.

• Feed back inhibition is when the amount of products alter enzyme productions.

• From A to Z, Z will inhibit some enzyme and lower its rate of production.

• multifacted chain inhibition stops chain and individual inhibition.

8-Covalent Modification:

• There are typically 2 formulas for enzyme activities in those types.
• Those acids often change their catalytic effectiveness in a reaction.
• The most likely mod is adding acid using atp.
• aside from acid, there are other reactions.

9- Limited proeolysis

• Sections of enzymes have inactive percursors synthesizers, but activate after removing a section by hydrolysis
• The enzymes are zymogen activated, using enzymes and other acids.
• Ex: digestive enzymes/ enzymes for cell death apoptosis.

10.Enzyme Production and Degradation (Turnover)

• Regulatory mechanisms also control the enzyme synthesis rate to regulate enzyme activity in the cell.
• The turnover of these reactions will determine overall time length Other mechanisms for this reactions are:
• Specific enzymes
• Genetic codes
• the amount of reactions itself

ISOENZYME WHAT???

• the same equation just with different enzymes
• can only happen under extreme conditions
• the electrical components will be different than others.
• there are enzymes within different tissues that behave differently in a reaction.
• Isozymes are tissue specific can be used to diagnose.