INTRODUCTION TO ENZYMES : 2.

Function:
- Coenzymes often participate in enzymatic reactions by donating or accepting
GENERAL CHARACTERISTICS OF ENZYMES: chemical groups.
1. Catalytic Activity: - They serve as carriers of specific functional groups, such as electrons, atoms, or
- Enzymes act as catalysts, speeding up chemical reactions without being consumed molecules, during catalysis.
in the process. 3. Examples:
- The enzyme-substrate complex forms during the reaction, facilitating the - Nicotinamide adenine dinucleotide (NAD+)
conversion of substrates into products. - Flavin adenine dinucleotide (FAD)
2. Specificity: - Coenzyme A (CoA)
- Enzymes are highly specific, each catalyzing a particular reaction or a group of - Adenosine triphosphate (ATP)
similar reactions. - Tetrahydrofolate (THF)
- Substrate specificity ensures that enzymes bind only to specific substrates with - Coenzyme Q (ubiquinone)
complementary shapes and chemical properties. 4. Role in Metabolism:
3. Optimal Conditions: - Coenzymes play crucial roles in various metabolic pathways, including glycolysis,
- Enzymes have specific optimal conditions for activity, including temperature, pH, the citric acid cycle, and the electron transport chain.
and substrate concentration. - They facilitate the transfer of energy and chemical groups between different
- Temperature: Most enzymes function optimally within a specific temperature biochemical reactions.
range.
- pH: Enzymes have an optimal pH at which they function most effectively.
PROSTHETIC GROUPS:
- Substrate Concentration: Enzyme activity is influenced by substrate 1. Definition:
concentration, following Michaelis-Menten kinetics. - Prosthetic groups are non-protein organic or inorganic molecules that are
4. Regulation: permanently attached to the enzyme's protein structure.
- Enzyme activity can be regulated through various mechanisms, including 2. Attachment:
allosteric regulation, feedback inhibition, and covalent modification. - Prosthetic groups are covalently bound to the enzyme and are essential for the
- Allosteric Regulation: Binding of regulatory molecules at allosteric sites alters enzyme's catalytic activity.
enzyme activity. 3. Function:
- Feedback Inhibition: End product of a metabolic pathway inhibits an enzyme - Prosthetic groups often play structural or functional roles in enzymatic reactions.
earlier in the pathway. - They may assist in substrate binding, stabilize reaction intermediates, or
- Covalent Modification: Addition or removal of functional groups can alter enzyme participate directly in catalysis.
activity. 4. Examples:
Theoretical Points: - Heme in catalase and cytochrome enzymes.
 Enzymes are biological catalysts that accelerate chemical reactions. - Flavin mononucleotide (FMN) in flavoproteins.
 They lower the activation energy required for a reaction to occur. - Iron-sulfur clusters in various enzymes involved in electron transfer reactions.
- Biotin in carboxylase enzymes.
 Enzymes are specific in their action and exhibit high substrate specificity.
 Enzymes function optimally under specific conditions of temperature, pH, and Theoretical Points:
substrate concentration.  Coenzymes and prosthetic groups are essential for the catalytic activity of many
 Enzyme activity can be regulated to maintain cellular homeostasis. enzymes.
 Cofactors and coenzymes play essential roles in enzyme catalysis.  Coenzymes are loosely bound to enzymes and serve as carriers of specific
chemical groups.
COENZYMES:  Prosthetic groups are tightly bound to enzymes and play structural or functional
1. Definition: roles in catalysis.
- Coenzymes are non-protein organic molecules that bind temporarily to the active  Both coenzymes and prosthetic groups participate in various metabolic
site of an enzyme and assist in catalytic activity. pathways, facilitating biochemical reactions.
APO-ENZYMES AND HOLO-ENZYMES:
APO-ENZYMES:
1. Definition:
- Apo-enzymes are inactive enzyme precursors that lack essential cofactors or
prosthetic groups necessary for catalytic activity.
2. Characteristics:
- Apo-enzymes typically consist only of the protein component of the enzyme.
- Without the necessary cofactors or prosthetic groups, apo-enzymes cannot
catalyze biochemical reactions effectively.
3. Formation:
- Apo-enzymes are formed either during enzyme synthesis or when cofactors or
prosthetic groups are removed from holo-enzymes.
4. Inactive State:
- In the absence of cofactors or prosthetic groups, apo-enzymes remain in an
inactive state and cannot participate in catalysis.
HOLO-ENZYMES:
1. Definition:
- Holo-enzymes are active enzyme complexes formed by the association of apo-
enzymes with their respective cofactors or prosthetic groups.
2. Characteristics:
- Holo-enzymes are fully functional and capable of catalyzing biochemical reactions
due to the presence of cofactors or prosthetic groups.
3. Formation:
- Holo-enzymes are formed when apo-enzymes bind to their specific cofactors or
prosthetic groups, resulting in the activation of enzymatic activity.
4. Active State:
- In the presence of cofactors or prosthetic groups, holo-enzymes exhibit catalytic
activity and participate in metabolic pathways.
Theoretical Points:
 Apo-enzymes are inactive enzyme precursors lacking essential cofactors or
prosthetic groups.
 Holo-enzymes are active enzyme complexes formed by the association of apo-
enzymes with cofactors or prosthetic groups.
 The presence or absence of cofactors or prosthetic groups determines the
catalytic activity of enzymes.
 Enzyme activation often involves the binding of cofactors or prosthetic groups
to apo-enzymes to form holo-enzymes.
Classification and Nomenclature of Enzymes:
CLASSIFICATION OF ENZYMES:
1. Enzyme Commission (EC) Number:
- The Enzyme Commission (EC) number system is the most widely used method for
classifying enzymes.
- It assigns a unique numerical code to each enzyme based on the reaction it
catalyzes.
- The EC number consists of four components, representing the enzyme's class,
subclass, sub-subclass, and serial number within the sub-subclass.
2. Enzyme Classes:
- Enzymes are classified into six main classes based on the type of reaction they
catalyze:
- Oxidoreductases: Catalyze oxidation-reduction reactions.
- Transferases: Transfer functional groups between molecules.
- Hydrolases: Catalyze hydrolytic cleavage reactions.
- Lyases: Catalyze the addition or removal of groups to form double bonds.
- Isomerases: Catalyze the rearrangement of atoms within a molecule.
- Ligases: Catalyze the joining of two molecules using ATP.
3. Subclasses and Sub-Subclasses:
- Each enzyme class is further divided into subclasses and sub-subclasses based on
specific characteristics of the reaction they catalyze.
- The classification system provides detailed information about the enzyme's
mechanism and substrate specificity.
NOMENCLATURE OF ENZYMES:
1. Systematic Names:
- Enzymes are often named systematically based on the reaction they catalyze and
the substrates involved.
- The systematic name typically consists of the substrate acted upon, followed by
the type of reaction and the suffix "-ase."
2. Trivial Names:
- Enzymes also have common or trivial names that are easier to remember and
pronounce.
- Trivial names are often derived from the substrate or product of the reaction, or
they may reflect the enzyme's function or source.
3. Examples:
- Systematic Name: Lactate dehydrogenase (catalyzes the conversion of lactate to
pyruvate)
- Trivial Name: Alcohol dehydrogenase (catalyzes the oxidation of alcohols)
Theoretical Points:
 Enzymes are classified based on the type of reaction they catalyze, using the
Enzyme Commission (EC) number system.
 The EC number system assigns a unique numerical code to each enzyme,
indicating its class, subclass, sub-subclass, and serial number.
 Enzymes can also be named systematically or by trivial names based on their
function, substrate specificity, or source.
 Systematic names provide detailed information about the enzyme's catalytic
activity, while trivial names are more convenient for everyday use.
ENZYME ASSAYS:
1. Definition:
- Enzyme assays are laboratory techniques used to measure the activity of enzymes
in a sample.
- These assays provide quantitative or qualitative information about the enzyme's
catalytic activity under specific conditions.
2. Importance:
- Enzyme assays are crucial for studying enzyme kinetics, determining enzyme
activity levels, and characterizing enzyme properties.
- They are used in basic research, drug discovery, clinical diagnostics, and industrial converted to product per minute (U/min) or micromoles of substrate converted to
processes.
3. Principles:
- Enzyme assays are based on monitoring changes in substrate concentration,
product formation, or other reaction parameters over time.
- The rate of substrate conversion or product formation is directly proportional to
enzyme activity and can be measured using various methods.
4. Types of Assays:
- Continuous Assays: Measure changes in substrate or product concentration
continuously over time using spectrophotometry, fluorometry, or chromatography.
- Endpoint Assays: Measure the amount of product formed or substrate remaining
at a specific time point using colorimetric, fluorometric, or radioactive detection
methods.
- Kinetic Assays: Determine enzyme kinetic parameters such as Michaelis-Menten
constant (Km) and maximum velocity (Vmax) by varying substrate concentration and
measuring initial reaction rates.
5. Assay Conditions:
- Enzyme assays are performed under controlled conditions of temperature, pH,
substrate concentration, and cofactor presence to ensure accurate measurement of
enzyme activity.
- Optimal assay conditions may vary depending on the specific enzyme and its
physiological environment.
Theoretical Points:
 Enzyme assays are laboratory techniques used to measure the activity of
enzymes.
 These assays provide quantitative or qualitative information about enzyme
catalysis under specific conditions.
 Enzyme assays are essential for studying enzyme kinetics, characterizing
enzyme properties, and assessing the effects of inhibitors or activators.
 Various types of enzyme assays exist, including continuous assays, endpoint
assays, and kinetic assays, each with its advantages and applications.
Enzyme Activity and Specific Activity:
ENZYME ACTIVITY:
1. Definition:
- Enzyme activity refers to the catalytic effectiveness of an enzyme, measured by the
rate at which it catalyzes a specific chemical reaction.
- It represents the amount of substrate converted to product per unit time under
specified conditions.
2. Units of Measurement:
- Enzyme activity is commonly expressed in units such as moles of substrate
product per minute (µmol/min).
- Other units include international units (IU) or katal (kat), which are standardized
measures of enzyme activity.
3. Factors Influencing Activity:
- Enzyme activity is influenced by various factors, including temperature, pH,
substrate concentration, enzyme concentration, and the presence of cofactors or
inhibitors.
- Optimal conditions for enzyme activity vary depending on the specific enzyme and
its physiological environment.
4. Measurement:
- Enzyme activity is determined experimentally using enzyme assays, which
measure changes in substrate concentration, product formation, or reaction kinetics
over time.
- The rate of substrate conversion or product formation is proportional to enzyme
activity and can be quantified using spectrophotometry, fluorometry, or other
detection methods.
SPECIFIC ACTIVITY:
1. Definition:
- Specific activity is a measure of the purity of an enzyme preparation, calculated as
the ratio of enzyme activity to total protein concentration.
- It represents the amount of enzyme activity per unit of protein and is expressed in
units such as U/mg or U/g of protein.
2. Importance:
- Specific activity provides information about the relative purity and efficiency of an
enzyme preparation.
- It is used to assess the success of enzyme purification procedures and to compare
the activities of different enzyme preparations.
3. Calculation:
- Specific activity is calculated using the formula:
Specific activity = Enzyme activity (U) / Total protein concentration (mg or g)
- It indicates the amount of enzyme activity per unit mass of protein in the sample.
4. Interpretation:
- Higher specific activity values indicate greater purity and efficiency of the enzyme
preparation, as more active enzyme is present relative to the total protein content.
- Lower specific activity values may indicate contamination with other proteins or
inactive forms of the enzyme.
Theoretical Points:
 Enzyme activity refers to the catalytic effectiveness of an enzyme in converting
substrate to product.
 It is influenced by various factors such as temperature, pH, substrate
concentration, and enzyme concentration.
 Enzyme activity is measured experimentally using enzyme assays, which
quantify changes in substrate or product concentrations over time.
 Specific activity is a measure of enzyme purity, calculated as the ratio of enzyme
activity to total protein concentration.
SPECIFICITY OF ENZYMES:
1. Definition:
- Enzyme specificity refers to the ability of enzymes to selectively bind to specific
substrates and catalyze particular chemical reactions.
- It ensures that enzymes interact only with their target substrates, leading to
precise and efficient catalysis.
2. Lock-and-Key Model:
- The lock-and-key model of enzyme specificity describes the complementary fit
between the enzyme's active site and the substrate.
- According to this model, the active site of the enzyme has a specific shape and
chemical environment that matches the structure of the substrate.
3. Induced Fit Model:
- The induced fit model suggests that the active site of the enzyme undergoes
conformational changes upon substrate binding to achieve optimal interaction.
- Substrate binding induces structural changes in the enzyme, leading to the
formation of the enzyme-substrate complex.
4. Types of Specificity:
- Substrate Specificity: Enzymes exhibit specificity for certain substrates based on
their chemical structure and functional groups.
- Stereochemical Specificity: Enzymes distinguish between stereoisomers and
selectively bind to one stereoisomer over another.
- Reaction Specificity: Enzymes catalyze specific types of chemical reactions, such as
oxidation-reduction, hydrolysis, or isomerization.
CONCEPT OF ACTIVE SITE:
1. Definition:
- The active site of an enzyme is a region on the enzyme's surface where the
substrate binds and catalysis occurs.
- It contains amino acid residues that directly participate in substrate binding and
catalytic activity.
2. Key Features:
- The active site is typically a small, well-defined pocket or crevice on the enzyme
surface.
- It has a specific three-dimensional structure and chemical environment that
complement the structure of the substrate.
- Amino acid residues within the active site form hydrogen bonds, ionic
interactions, and hydrophobic interactions with the substrate.
3. Role in Catalysis:
- The active site provides a microenvironment that stabilizes the transition state of
the reaction, lowering the activation energy required for the reaction to occur.
- It facilitates the formation of enzyme-substrate complexes and the catalytic
conversion of substrates to products.
4. Flexibility and Adaptability:
- The active site may exhibit flexibility and adaptability to accommodate substrates
of varying sizes and shapes.
- Conformational changes in the active site may occur during substrate binding and
catalysis, contributing to enzyme specificity and efficiency.
Theoretical Points:
 Enzyme specificity ensures that enzymes selectively bind to specific substrates
and catalyze particular chemical reactions.
 The lock-and-key and induced fit models describe the complementary fit
between the enzyme's active site and the substrate.
 The active site is a region on the enzyme's surface where substrate binding and
catalysis occur, containing amino acid residues that participate in substrate
recognition and catalysis.
 Flexibility and adaptability of the active site contribute to enzyme specificity and
efficiency by allowing for conformational changes during substrate binding and
catalysis.
FISCHER'S LOCK AND KEY HYPOTHESIS:
1. Definition:
- Fischer's lock and key hypothesis, proposed by Emil Fischer in 1894, describes the
mechanism of enzyme-substrate interaction based on the complementarity between
the enzyme's active site and the substrate.
- According to this hypothesis, the active site of the enzyme has a specific shape and
chemical environment that precisely fits the structure of the substrate, similar to a
lock and key.
2. Key Features:
- Enzymes have a rigid active site with a specific three-dimensional structure that is
complementary to the structure of the substrate.
- The substrate fits into the active site like a key into a lock, forming an enzyme-
substrate complex.
 - The active site provides a microenvironment that stabilizes the transition state of
the reaction, facilitating catalysis.
3. Implications:
- Fischer's lock and key hypothesis suggests that enzyme specificity is determined
by the precise fit between the enzyme's active site and the substrate.
- Only substrates with complementary structures can bind to the active site and
undergo catalysis, ensuring specificity of enzyme-substrate interactions.
Theoretical Points:
 Fischer's lock and key hypothesis proposes that enzyme-substrate interaction is
based on the complementarity between the enzyme's active site and the
substrate, similar to a lock and key.
 The active site of the enzyme has a specific three-dimensional structure that
precisely fits the structure of the substrate, allowing for selective binding and
catalysis.
 The hypothesis implies that enzyme specificity is determined by the precise fit
between the active site and the substrate, ensuring efficient and specific
catalysis.
KOSHLAND'S INDUCED FIT HYPOTHESIS:
1. Definition:
- Koshland's induced fit hypothesis, proposed by Daniel E. Koshland Jr. in 1958,
describes the mechanism of enzyme-substrate interaction based on dynamic
conformational changes in both the enzyme and the substrate upon binding.
- According to this hypothesis, the active site of the enzyme is flexible and can
undergo structural changes upon substrate binding, leading to the optimal alignment
of the enzyme and substrate for catalysis.
2. Key Features:
- Enzymes and substrates do not have rigid structures but instead possess some
degree of flexibility.
- When a substrate binds to the enzyme's active site, both the enzyme and the
substrate undergo conformational changes.
- The induced conformational changes in the enzyme's active site optimize the fit
between the enzyme and the substrate, enhancing catalytic efficiency.
3. Implications:
- Koshland's induced fit hypothesis suggests that enzyme specificity is not solely
determined by the static structure of the active site but also by the dynamic
interactions between the enzyme and the substrate.
- The flexibility of the active site allows enzymes to accommodate substrates of
varying sizes and shapes, contributing to their catalytic versatility.
Theoretical Points:
 Koshland's induced fit hypothesis proposes that enzyme-substrate
interaction involves dynamic conformational changes in both the enzyme and
the substrate upon binding.
 The active site of the enzyme is flexible and can undergo structural changes
to optimize the fit with the substrate.
 The hypothesis emphasizes the dynamic nature of enzyme-substrate
interactions and the role of induced conformational changes in catalysis.
CATALYTIC POWER:
1. Definition:
- Catalytic power refers to the ability of an enzyme to enhance the rate of a chemical
reaction compared to the same reaction in the absence of the enzyme.
- It quantifies the efficiency of enzyme catalysis in accelerating the conversion of
substrate into product.
2. Factors Influencing Catalytic Power:
- Active Site: The structure and chemical environment of the active site determine
the enzyme's ability to bind to the substrate and facilitate catalysis.
- Substrate Specificity: The enzyme's specificity for its substrate influences its
catalytic power, as it ensures selective binding and efficient catalysis.
- Transition State Stabilization: Enzymes stabilize the transition state of the
reaction, lowering the activation energy barrier and enhancing catalytic efficiency.
- Proximity and Orientation Effects: Enzymes bring reactants into close proximity
and proper orientation, facilitating the formation of the transition state and
increasing the reaction rate.
- Cofactors and Coenzymes: Cofactors and coenzymes assist enzymes in catalysis by
providing additional chemical groups or facilitating electron transfer reactions.
3. Measurement:
- Catalytic power is typically quantified by comparing the rate of the enzymatic
reaction to the rate of the same reaction in the absence of the enzyme.
- It is expressed as the ratio of the rate constant (k_cat) for the enzyme-catalyzed
reaction to the rate constant for the uncatalyzed reaction.
4. Importance:
- Catalytic power reflects the efficiency and specificity of enzyme catalysis, which
are essential for the proper functioning of biological systems.
- Understanding the factors that influence catalytic power helps in the design of
enzyme inhibitors, drugs, and industrial catalysts.
Theoretical Points:
 Catalytic power measures the ability of an enzyme to accelerate a chemical
reaction compared to the uncatalyzed reaction.
 Factors such as active site structure, substrate specificity, transition state
stabilization, and cofactor involvement influence the catalytic power of
enzymes.
 Catalytic power is quantified by comparing the rate constant of the enzyme-
catalyzed reaction to that of the uncatalyzed reaction.