Enzyme kinetics and mechanism

Enzymes are biological catalysts that speed up chemical reactions in living organisms. Understanding how they work involves studying both their kinetics and mechanism. Enzyme kinetics refers to the rates of enzyme-catalyzed reactions and how various factors affect these rates. An understanding of enzyme mechanism requires knowledge about the specific steps of the catalytic process.

Basics of enzyme catalysis

Enzymes are proteins that act on specific substrates to catalyze a particular reaction. The place where the substrates bind is known as the active site. This binding is usually very specific, just like a key fits into a lock. The whole process can be summarized as follows:

E + S ↔ ES → E + P

Here, E is the enzyme, S is the substrate, ES is the enzyme-substrate complex, and P is the product. After the reaction, the enzyme is free to catalyze another reaction.

Michaelis-Menten kinetics

Michaelis-Menten kinetics is a model that describes the kinetic behavior of many enzymes. The rate of the enzymatic reaction depends on the concentration of the substrate. The equation representing this model is:

v = (Vmax [S]) / (Km + [S])

Here, v is the reaction rate, Vmax is the maximum reaction rate, [S] is the substrate concentration, and Km (Michaelis constant) is a measure of the substrate concentration at which the reaction rate is half of Vmax.

This model assumes that the formation and disintegration of ES complex are at equilibrium and that the product formation step is the rate-limiting step.

This illustration shows a typical Michaelis-Menten plot where the reaction rate v increases with increasing substrate concentration and eventually reaches a maximum rate Vmax.

Factors affecting enzyme activity

Enzymatic activity can be affected by several factors:

• Temperature: Increasing temperature generally increases the reaction rate up to a point called the enzyme's optimum temperature. Beyond this, enzymes can become denatured and lose their activity.
• pH: Each enzyme has an optimal pH range. Deviations can lead to decreased activity because they affect the charged groups needed for binding of the substrate.
• Substrate concentration: Higher concentrations increase rates up to the saturation point, after which further increases in substrate do not affect the rate, as shown in the Michaelis-Menten graph.
• Inhibitors: Compounds that reduce enzyme activity by interfering with substrate binding or reducing the turnover number of the enzyme. There are competitive, non-competitive, and non-competitive inhibitors.

Enzyme inhibition

Enzyme inhibition is a process in which the activity of an enzyme is slowed or stopped by another molecule. It is important for regulating enzyme activity in a pathway.

Types of prohibitions

Competitive inhibition: In this type, the inhibitor directly competes with the substrate for the active site of the enzyme. The presence of competitive inhibitor can be overcome by increasing the concentration of the substrate. The Michaelis-Menten equation for competitive inhibition is modified as follows:

v = (Vmax [S]) / (αKm + [S])

where α = 1 + [I]/Ki and Ki is the inhibitor constant, and [I] is the inhibitor concentration.

Non-competitive inhibition: The inhibitor binds to a site other than the active site, causing a decrease in the maximum reaction rate Vmax, but has no effect on the binding of the substrate. The equation becomes:

v = (Vmax/α) [S] / (Km + [S])

Here, α directly affects Vmax.

Enzyme mechanism

Understanding enzyme mechanism involves knowing how enzymes stabilize transition states, their active site architecture, and the kinetics of substrate conversion into product. There are several ways to do this:

Covalent catalysts

In covalent catalysis, the enzyme forms a transient covalent bond with the substrate, thereby facilitating the catalytic process. This involves an additional step where a covalent intermediate is formed before being broken down into the product:

E + S ↔ ES ↔ EX → E + P

Here, EX denotes covalent intermediate.

Acid-base catalyst

This mechanism involves the transfer of protons between the enzyme and the substrate. Enzymes such as lysozyme use acidic and basic residues to stabilize the transition states and facilitate the scission of bonds.

Metal ion catalysis

Some enzymes require metal ion cofactors for their activity. Metal ions can stabilize negatively charged intermediates, act as electrophilic catalysts, or bridge substrates to enzymes.

Multisubstrate reactions

Many enzymes catalyze reactions involving two or more substrates. These enzymes can work through one of two mechanisms:

Sequential reactions: Both substrates must bind to the enzyme before any products are released. Ordered sequential mechanisms require the substrates to bind in a specific order.

E + S1 → ES1 ↔ ES1S2 → E + P1 + P2

Ping-pong reactions: One or more products are released before all the substrates have attached to the enzyme. The enzyme switches between two states during the process.

E + S1 ↔ E'P1 → E' → E + P2

Conclusion

Understanding the dynamics and mechanisms of enzymes is important in biochemistry because enzymes are vital to life. They are involved in digestion, metabolism, DNA replication, and many other processes. By studying enzymes, scientists can develop drugs that inhibit or activate enzymes, thereby improving human health.

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