Factors Affecting Enzyme Activity: pH, Substrates

What is Enzyme Activity?

Enzymes are protein molecules that act as catalysts. They do it in a very specific manner, recognising specific substrates, and precise mechanisms or pathways, converting the substrates into products.

Definition and Mechanism

Enzyme activity is the rate at which an enzyme converts substrates into products. It reflects the catalytic activity of enzymes.

The mechanism of enzyme activity involves an enzyme forming a temporary enzyme-substrate complex, converting substrate molecules into products and then finally releasing the products. Enzymes enhance the rate of chemical reactions by lowering the activation energy required for any reaction to proceed

Importance in Biological Processes

Almost all biochemical processes that are important for life depend on enzymes. Some examples of these processes are digestion, metabolism, DNA replication and cellular respiration. Without enzymes, these reactions would proceed too slowly for life to exist.

It is because of their unique three-dimensional shapes that enzymes obtain specificity. These unique shapes permit specific substrates to bind at the active site of an enzyme where they are converted into products. In this way, there is limited binding of the wrong substrate, which would decrease the integrity of the biological process by generating the wrong product.

Factors Affecting Enzyme Activity

The activity of enzymes is tightly regulated to provide homeostasis in cells and also to adjust to environmental changes. This may be through temperature, pH, concentration of substrate, inhibitors of enzymes, and also cofactors and coenzymes.

Temperature

Enzymes show the highest activity at optimum temperature. The activity declines progressively both above and below the temperature.

pH

Similarly, each enzyme acts best at a particular pH called optimum pH. The activity declines both above and below that pH.

Substrate Concentration

As the substrate concentration increases, the rate of reaction increases. This is because molecules will interact with enzyme molecules, the more products will be formed.

Enzyme Concentration

An increase in enzyme concentration generally speeds up the reaction rate, as more active sites become available for substrate binding.

Presence of Inhibitors

Inhibitors slow down or stop enzyme activity by binding to the enzyme’s active site, preventing substrate binding or altering enzyme structure.

Temperature and Enzyme Activity

The effect of temperature on enzyme activity is discussed below:

Optimum Temperature

• Temperature, in general, increases enzyme activity since there is heightened motion of molecules that results in increased collision.

• There exists an optimum temperature for each enzyme, where its activity is maximum. For example, most human enzymes have optimum temperatures near 37°C.

Denaturation and Industrial Relevance

• High temperatures can disrupt the enzyme structure, changing the shape of its active site, which is more important for substrate binding.

• Extreme temperatures can denature enzymes, and the molecule cannot function.

• Enzymes in brewing and baking are temperature sensitive.

• Industrial processes like pharmaceutical production are done in controlled enzyme temperatures for efficiency.

pH and Enzyme Activity

The effect of pH on enzyme activity is discussed below:

Optimum pH Range

• Each enzyme has an optimal pH at which the rate of the reaction is highest.

• At this pH value, these enzymes have their normal configurations.

Effect of pH on Active Site Ionization

• A change in pH can alter the ionization of the side chains and disrupt normal interactions. Under extreme conditions, pH, in fact, inactivates the enzyme.

• Examples include digestive enzymes, amylase present in the mouth has an optimum pH of about 7 and pepsin in the stomach has an optimum pH of about 2.

Substrate Concentration and Enzyme Kinetics

The effect of substrate concentration on enzyme activity is discussed below:

Michaelis-Menten Constant (Km)

• Generally, enzyme activity increases with an increase in substrate concentration because more and more substrate molecules collide with the enzyme.

• That is, more products are generated per unit of time as more substrate molecules occupy active sites.

• Michaelis Menten Constant, Km explains the concentration of substrate required to produce a reaction that is equal to half of the maximum velocity of the reaction.

• For example, the Km decreases when the competitive inhibitor is included.

Enzyme Saturation Point

• The enzyme saturation point is reached when all enzyme active sites are occupied by substrate molecules, and no further increase in reaction rate occurs even if substrate concentration increases.

• At this stage, the reaction proceeds at its maximum velocity (Vmax), as the enzyme is working at full catalytic capacity.

Enzyme Inhibitors and Regulation

Enzyme inhibitors are of three types:

Competitive Inhibition

• It occurs because the molecule of the inhibitor binds to the active site of the enzyme.

• An inhibitor molecule is similar enough to the substrate.

• This is known as competitive inhibition because, in effect, an inhibitor is competing with the substrate for the active binding site.

• An example of a use for a competitive inhibitor is in the treatment of influenza via the neuraminidase inhibitor, Relenza.

Uncompetitive Inhibition

• Sometimes, binding of the substrate to the active site changes the conformation of the enzyme.

• It creates an allosteric site that was not present earlier, before the binding of the substrate.

• This is the site to which an inhibitor binds and exerts its inhibitory effect.

• It cannot be overcome by increasing substrate concentration.

Non-Competitive Inhibition

• Some enzymes have permanent allosteric sites that can bind to the inhibitor.

• As the inhibitor does not compete for the active site, it is called non-competitive inhibition.

• In non-competitive inhibition, the Km does not change.

• This is because Km is a measure of the affinity of the enzyme for its substrate and this can only be measured by the active enzyme.

• The fixed amount of inactive enzyme in non-competitive inhibition does not affect the Km and, therefore, is unchanged.

• An example of a use for a noncompetitive inhibitor is in the use of cyanide as a poison (prevents aerobic respiration)

Allosteric Inhibition

• These bind at other sites of the enzyme and the altering of the shape of the enzyme in the process reduces effectiveness.

• In this binding, the activity of the enzyme in question can either be reduced or enhanced depending on the nature of the compound referred to as the allosteric modulator.

• Allosteric regulation is reversible and depends on the signal in the cell or something that the cell requires.

Role of Cofactors and Coenzymes

Many enzymes require cofactors and coenzymes to facilitate biochemical reactions.

Inorganic Cofactors (Zn²⁺, Mg²⁺)

• Cofactors can be either inorganic, that is, a metal ion, or organic molecules, but they are not proteins.

• Some examples of cofactors are the zinc ion Zn²⁺ in carbonic anhydrase and the magnesium ion Mg²⁺ in hexokinase.

Organic Coenzymes (NAD⁺, FAD, CoA)

• Coenzymes are organic molecular helpers, especially vitamin derivatives.

• They can work along with an enzyme to serve in a reaction, but they can work independently in the cell.

• Examples of coenzymes include NAD⁺, FAD, and Coenzyme A. Enzymes can best conduct catalytic activities using such molecules, therefore allowing the transfer of chemical groups during a reaction.

Regulation of Enzyme Activity

Enzyme activity can be regulated by various mechanisms:

Feedback Inhibition

• One of the end products in a metabolic pathway can inhibit enzymes in earlier steps in the pathway to prevent too much of the end product from being made.

• The cell is self-regulating and efficiently uses its resources through these types of mechanisms.

Enzyme Induction and Repression

• Enzyme induction is the process where the synthesis of enzymes is increased by a substrate or environmental change.

• Enzyme repression is when the product is in excess or when there is a high abundance of a repressor molecule, enzyme synthesis is repressed, thus conserving energy and resources.

Enzyme Cascades

• Enzyme cascades include several metabolic pathways in cells mediated by cascades of enzymatic reactions that could potentially amplify signals greatly.

Allosteric Regulation (ATP and PFK example)

• Molecules can bind to allosteric sites on enzymes, inducing a conformational change that either increases or decreases enzyme activity.

• Such regulation allows for fine control of metabolic pathways in response to cellular demands.

• In allosteric regulation in glycolysis, ATP is an allosteric inhibitor of phosphofructokinase. Whenever energy levels in the cell are high, ATP binds to phosphofructokinase and inhibits it, and that is a turn off this pathway.

Applications of Enzyme Activity

The application of enzyme activity includes:

Industrial Enzyme Optimization

Through enzyme optimization, conditions like pH, temperature, and concentration are adjusted to achieve maximum activity. Industrial enzymes are used in food processing, textile manufacturing, biofuel production, and detergent formulation for faster and more eco-friendly processes.

Pharmaceutical and Medical Uses

In medicine, enzymes are used for diagnosis and therapy. For example, streptokinase helps dissolve blood clots, lactase is used to treat lactose intolerance, and enzymes like DNA polymerase are key in molecular diagnostics. Enzyme-based assays such as ELISA are also essential for detecting diseases and monitoring health conditions.

Enzyme Activity NEET MCQs (With Answers & Explanations)

Important topics for NEET exam are:

• Factors affecting enzyme activity

• Regulation of Enzyme Activity

Practice Questions for NEET

Q1. Which combination of relative values of km (Michaelis constant) and kcat (catalytic rate constant) would indicate a high catalytic efficiency for an enzyme?

1. Low km and low kcat

2. High km and high kcat

3. High km and low kcat

4. Low km and high kcat

Correct answer: 4) Low km and high kcat

Explanation:

The combination of low km (Michaelis constant) and high kcat (catalytic rate constant) indicates a high catalytic efficiency for an enzyme.
A low km value indicates that the enzyme has a high affinity for its substrate, meaning it can effectively bind to the substrate at relatively low concentrations.
On the other hand, a high kcat value indicates that the enzyme has a high turnover rate, meaning it can efficiently convert substrate molecules into product molecules per unit time.

Hence, the correct answer is option 4). Low km and high kcat.

Q2. Choose the option that does not accurately describe the characteristics of enzyme catalase.

1. The cofactor exhibits an inorganic and proteinaceous nature.

2. Haem serves as the necessary prosthetic group for the enzyme.

3. The cofactor necessary for optimal activity must be firmly bound to the apoenzyme.

4. It facilitates the decomposition of hydrogen peroxide into water and oxygen.

Correct answer: 1) The cofactor exhibits an inorganic and proteinaceous nature.

Explanation:

Option 1 does not accurately describe the characteristics of enzyme catalase. While catalase does require a cofactor for its activity, the cofactor is not both inorganic and proteinaceous. The cofactor for catalase is a heme group, which is an iron-containing prosthetic group. The heme group is essential for catalase's function in facilitating the decomposition of hydrogen peroxide into water and oxygen.

Hence, the correct answer is option 1) The cofactor exhibits an inorganic and proteinaceous nature.

Q3. In the context of its catalytic function in the conversion of glucose-1-phosphate to glucose-6-phosphate, which enzyme classification is most appropriate for phosphoglucomutase?

1. Oxidoreductase

2. Isomerase

3. Kinase

4. Lyase

Correct answer: 2) Isomerase

Explanation:

Phosphoglucomutase is classified as an isomerase enzyme. Isomerases are a class of enzymes that catalyze the interconversion of isomers, which are molecules with the same molecular formula but different structural arrangements. In the case of phosphoglucomutase, it facilitates the conversion of glucose-1-phosphate into glucose-6-phosphate by rearranging the positions of phosphate groups within the molecule. This rearrangement is a structural isomerization reaction, and thus, phosphoglucomutase is classified as an isomerase enzyme.

Hence, the correct answer is option 2)Isomerase.

Also Read-

• MCQs on Factors Affecting Enzyme Activity

• Biomolecules

• Metabolites