Enzymes

Introduction to Enzymes

Enzymes are globular proteins that function as highly specific biological catalysts. They accelerate the rate of metabolic reactions by providing an alternative reaction pathway with a lower activation energy (Ea), which is the minimum energy required to initiate a reaction. They do not get used up in the reaction and do not alter the overall energy change (Gibbs free energy) of the reaction.

The specificity of an enzyme is determined by its active site, a unique three-dimensional cleft or pocket formed by the folding of the polypeptide chain. The amino acid R-groups lining the active site are precisely positioned to bind to a specific substrate molecule.

Models of Enzyme Action

1. Lock and Key Model: An early, simplified model suggesting the substrate fits perfectly into a rigid active site, like a key into a lock. This model fails to explain the flexibility of some enzymes.
2. Induced-Fit Model: The currently accepted and more accurate model. It proposes that the active site is flexible. The initial binding of the substrate induces a slight change in the shape of the active site, allowing it to fit the substrate more precisely. This conformational change strains the substrate's bonds, facilitating the reaction and forming the enzyme-substrate complex (ESC). Once the reaction is complete, the products are released, and the enzyme returns to its original shape.

Factors Affecting the Rate of Enzyme-Catalysed Reactions

The rate is typically measured as the change in concentration of product or substrate over time (e.g., mol dm⁻³ s⁻¹).

* Temperature: As temperature increases, the kinetic energy of both enzyme and substrate molecules increases. This leads to more frequent and energetic collisions, increasing the rate of reaction up to an optimum temperature. In humans, this is typically around 37°C. Beyond the optimum, the enzyme's thermal energy becomes too high, causing its atoms to vibrate excessively. This breaks the weak hydrogen and ionic bonds that maintain its specific tertiary structure. The active site changes shape permanently, a process called denaturation. The enzyme can no longer bind to its substrate, and the reaction rate plummets.

* pH: Each enzyme has an optimum pH at which its activity is maximal. Deviations from this optimum alter the ionization of the amino acid R-groups, particularly those in the active site. This disrupts the ionic bonds that maintain the enzyme's tertiary structure and can change the charges on the active site, preventing the substrate from binding. Extreme changes in pH cause irreversible denaturation.

* *Example:* Pepsin in the stomach has an optimum pH of ~2, while trypsin in the small intestine has an optimum pH of ~8.

* Substrate Concentration [S]: At a fixed enzyme concentration, increasing the substrate concentration increases the rate of reaction. This is because there are more substrate molecules available to collide with and bind to the enzyme's active sites. The rate continues to increase until the enzyme's active sites become saturated with substrate. At this point, the reaction reaches its maximum velocity (Vmax), and further increases in [S] will not increase the rate. The enzyme concentration is now the limiting factor.

* Enzyme Concentration [E]: Provided there is an excess of substrate, the initial rate of reaction is directly proportional to the enzyme concentration. More enzyme molecules mean more active sites are available to process the substrate.

Enzyme Kinetics: Vmax and Km

* Vmax (Maximum Velocity): The theoretical maximum rate of the reaction when all enzyme active sites are saturated with substrate. It is a measure of the enzyme's catalytic efficiency.

* Michaelis-Menten Constant (Km): Defined as the substrate concentration at which the reaction rate is half of Vmax (½Vmax). Km is a crucial indicator of an enzyme's affinity for its substrate.

* A low Km value indicates a high affinity. The enzyme can bind and catalyse the reaction efficiently even at low substrate concentrations.

* A high Km value indicates a low affinity. A higher substrate concentration is needed to achieve half the maximum velocity.

Enzyme Inhibition

Inhibitors are molecules that bind to an enzyme and reduce its activity. Inhibition can be reversible or irreversible.

1. Competitive Inhibition:

* Mechanism: The inhibitor molecule has a shape that is structurally similar to the substrate. It competes with the substrate for the same active site. Binding is reversible.

* Effect on Kinetics:

* Vmax is unchanged: The effect of the inhibitor can be overcome by significantly increasing the substrate concentration, as the substrate will out-compete the inhibitor, eventually allowing the reaction to reach its normal Vmax.

* Km is increased: A higher substrate concentration is required to reach ½Vmax because the substrate has to compete with the inhibitor.

1. Non-competitive Inhibition:

* Mechanism: The inhibitor binds to the enzyme at a location other than the active site, known as the allosteric site. This binding causes a conformational change in the enzyme's tertiary structure, which alters the shape of the active site. As a result, the substrate can no longer bind effectively, or the enzyme is less efficient at catalysing the reaction.

* Effect on Kinetics:

* Vmax is decreased: Since some enzyme molecules are effectively 'inactivated' by the inhibitor, the overall maximum rate of reaction is reduced. Increasing the substrate concentration cannot overcome this effect.

* Km is unchanged: The inhibitor does not affect the binding of substrate to the active sites of the *uninhibited* enzyme molecules. Therefore, the affinity of the remaining functional enzymes for the substrate remains the same.

Common Exam Traps & Misconceptions:

* Do not say enzymes are 'killed' by heat; they are proteins and undergo denaturation.

* Be precise: Denaturation is the loss of the specific 3D tertiary structure due to the breaking of hydrogen and ionic bonds.

* Clearly distinguish the effects of competitive vs. non-competitive inhibitors on Vmax and Km. A sketch graph is often the best way to explain this.