What is an Enzyme?

In biochemistry, enzymes are proteins that act as biological catalysts. Catalysis is the addition of a catalyst to a chemical reaction to speed up the pace of the reaction. Catalysis can be categorized as either homogeneous or heterogeneous, depending on whether the catalysts are distributed in the same phase as that of the reactants. Enzymes are an essential part of the cell because, without them, many organic processes would slow down and thus will affect the processes that are important for cell survival and sustenance.

What is Enzyme kinetics?

Enzyme kinetics is a study of the rate of chemical reactions that are catalyzed by enzymes. The reaction rate is evaluated in enzyme kinetics, and the effects of various reaction conditions are investigated. The catalytic mechanism of this enzyme may be revealed using this biochemical technique. Enzyme kinetics provide important details regarding the catalytic mechanism of the enzyme, its importance in metabolism, how its activity is affected inside the cell in presence or absence of certain variables and how certain substance or drugs inhibits its activity.

Enzyme co-factor

The non-protein chemical compound or metal ions required by an enzyme for its activity is called a co-factor. The enzymes remain inactive until they are bound to the co-factor. An enzyme without the co-factor is known as an apoenzyme. The enzyme becomes active "holoenzyme" when the co-factor is added. The permanently attached co-factor is known as the prosthetic group, and the temporarily attached co-factor is known as a co-enzyme. Co-enzymes are involved in oxidation and reduction in biological reactions.

Factors that affect enzyme activity

There are various factors that affect the reactions catalyzed by enzymes. These factors include:

• Enzyme concentration.
• The pH of the medium.
• Substrate concentration.
• The temperature of the medium.
• Inhibitors.

Michaelis–Menten kinetics

In biochemistry, the Michaelis-Menten is one of the best models for enzyme kinetics.  Michaelis-Menten kinetics is a broad theory that describes the speed and mechanism of enzyme-catalytic processes. It also assumes that the rate at which the product, P, is formed is equal to the enzyme substrate complex concentration. When all of the places on the enzyme molecules where catalytic activity may occur (active sites) are occupied with substrate—that is, when the substrate concentration is very high—the reaction velocity is highest. All enzyme kinetic research and calculations are related to these connections. The relationship between reaction rate and substrate concentration was derived from this theory. It is represented by the given equation, also known as the Michaelis-Menten equation.

V=d[P]/dt=Vmax[S]/(Km+[S])

Effect of substrate concentration on enzyme kinetics

When the enzyme concentration remains constant, the relationship between substrate concentration and reaction rate is rectangular hyperbolic. When the substrate concentration is low, the reaction rate increases proportionally to the substrate concentration. Following that, as the substrate concentration increases, the reaction rate gradually decreases. Finally, after a certain amount of substrate concentration has been reached, the reaction rate reaches a maximum and no longer increases with the substrate concentration.

V_max and K_m

V_max

The V_max is considered as the highest speed. Speed is so high that an increase in the substrate concentration does not affect the reaction rate catalyzed by an enzyme. This occurs because the substrate molecules saturate the active sites of the enzyme and due to which the active sites of the enzymes are not accessible to form more enzyme-substrate complexes. V_max is proportional to the catalytic rate constant and the concentration of the enzyme.

In terms of V_max, the Michalis-Menten equation is represented as

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

K_m

K_m refers to the substrate concentration at which the reaction rate is 50% of V_max.  The lower the K_m value, the more efficient the enzyme performs its activity at lower substrate concentrations. The enzyme's substrate affinity is measured by K_m. A low K_m indicates that the substrate binds strongly to the enzyme, enabling it to saturate. The km value of an enzyme-substrate system may be utilized to assess if the cell needs more enzymes or more substrate to speed up the enzymatic process.

Significance of K_m and V_max value

V_max is the maximum velocity at which an enzyme catalyzes a chemical reaction. A reaction attains maximum velocity when all the active sites of the enzymes are saturated with substrate. Because the maximum velocity is said to be proportional to enzyme concentration, it can be used to determine enzyme concentration. K_m results in a half-maximal response rate. K_m and V_max are two parameters that identify the enzyme's kinetic behavior.

Enzyme inhibitor

Enzyme inhibitors are molecular agents that interfere with catalysis, slowing or stopping enzymatic reactions. They inhibit the process of substrate integration into the active site of the enzyme.

Enzyme inhibition

In biochemistry, enzymes are essential for the majority, if not all, of life's functions. Enzymes catalyze reactions by reducing the activation energy of the reaction. Enzymes, on the other hand, must be closely controlled to verify that product levels do not escalate to unacceptably high levels and inhibition of enzyme is used to accomplish this.

Class of enzyme inhibition

There are two types of enzyme inhibition:

• Reversible inhibition.
• Irreversible inhibition.

Reversible inhibition

Inhibition of enzyme activity in which the inhibiting molecule can associate and dissociate from the enzyme's active site reversibly. Reversible inhibition includes three kinds of inhibition, they are:

• Competitive inhibition.
• Non-competitive inhibition.
• Uncompetitive inhibition.

Competitive inhibition

In competitive inhibition, the competitive inhibitor competes with the substrate for the enzyme's active site, resulting in a rise in the Km. Vmax, on the other hand, remains constant since the reaction may still be completed with enough substrate concentration. Due to the increased Km, the graph of enzyme activity versus substrate concentration moves towards the right.

Non-competitive inhibition

In non-competitive inhibition, the inhibitor attaches to a site other than the active site of the enzyme and lowers the Vmax. The site is known as an allosteric site. Km, on the other hand, is unaffected. When compared to the reaction without any inhibitor, the rate of reaction is lower in the presence of a non-competitive inhibitor.

Uncompetitive inhibition

When the inhibitor mainly interacts with enzyme-subtract complex rather than enzyme alone, it is referred to as uncompetitive inhibition. It affects the reaction only at high substrate concentrations, which results in the formation of several enzyme-substrate complexes. This indicates that increasing the substrate concentration (S) does not prevent the inhibitor from binding.

Irreversible inhibition

During irreversible inhibition, inhibitors bind covalently to the enzyme's active site and affect the functional group that is essential for its activity. Irreversible inhibitors can be divided into two types: competitive and allosteric. Competitive inhibitors permanently bind to the enzyme's active site. Allosteric inhibitors permanently bind to an enzyme's allosteric site, and they change the shape of the enzyme.

Context and Applications

This topic is essential in the professional exams for both school level, undergraduate and postgraduate courses, especially for bachelors and masters in biochemistry, bachelors in molecular biology, and masters in molecular biology.

Practice problems

Question 1: When the velocity of enzyme activity against substrate concentration is shown, which of the following is the by-product?

1. Hyperbolic curve
2. Parabola
3. A straight line with a positive slope

Answer: Option 1 is correct.

Explanation: The rate of formation of an enzyme-substrate complex determines the rate of a reaction at low substrate concentrations.

Question 2: In Michaelis-Menten kinetics, rate-determining step is____________

1. The complex formation steps
2. The production formation step
3. The complex dissociation step is to produce products

Answer: Option 1 is correct.

Explanation: The breakdown of the enzyme-subtract complex is the rate-determining step of Michaelis-Menten’s kinetics.

Question 3: The chemical that directly affects an enzyme to reduce its catalytic rate is known as________

1. Repressor
2. Regulator
3. Inhibitor

Answer: Option 3 is correct.

Explanation: An inhibitor is a chemical that lowers the catalytic rate by interfering with the substrate-active site binding.

Question 4: Which of the following factors may be used to compare the catalytic efficiency of two different enzymes?

1. A pH of the optimum value
2. Km
3. Size of the enzyme

Answer:  Option 2 is correct.

Explanation: The substrate concentration is K_m. The rate of reaction increases as the substrate concentration rises.

Question 5: K_m is called as _________________

1. Maximum velocity
2. Michaelis constant
3. None of the above

Answer: Option 2 is correct.  

Explanation: Km is also called the Michaelis constant. Km measures an enzyme's affinity for its substrate; the lower the km value, the more effective the enzyme is in performing its activity and reducing the substrate concentration.