Enzymes: Biological Catalysts

The Hook: Nature’s Catalysts - Millions Times Faster

Connect: Real Life → Chemistry

Without enzymes, digesting a meal would take 50 years! Enzymes speed up reactions by factors of millions. Amylase in your saliva starts breaking down starch within seconds. Lactase helps digest milk sugar. Washing powders contain enzymes to remove stains. Diabetics lack or have ineffective insulin (a hormone, not enzyme, but related concept).

Here’s the JEE question: How can enzymes speed up reactions by million-fold without being consumed? Why does high temperature destroy enzyme activity? And how can a single enzyme distinguish between mirror-image molecules?

The Core Concept

What are Enzymes?

Enzymes = Biological catalysts that speed up biochemical reactions

Key characteristics:

Proteins (almost all enzymes are proteins)
Catalysts - increase reaction rate without being consumed
Highly specific - one enzyme, one reaction (usually)
Work at body temperature (mild conditions)
pH and temperature sensitive

Etymology: From Greek “en” (in) + “zyme” (yeast)

JEE Weightage

Enzymes: 2-3 questions in JEE Main, 1-2 in JEE Advanced Focus areas: Lock-and-key model, factors affecting activity, inhibition types, specificity

Enzyme Structure

Components

Simple Enzyme:

Protein only (pure protein)
Example: Pepsin, trypsin

Conjugated Enzyme (Holoenzyme):

Apoenzyme (protein part) + Cofactor (non-protein part)
Both needed for activity

$$\boxed{\text{Apoenzyme} + \text{Cofactor} = \text{Holoenzyme (active)}}$$

Apoenzyme alone: Inactive

Cofactors

Types of cofactors:

1. Metal ions (activators):

Zn²⁺: Carbonic anhydrase, carboxypeptidase
Mg²⁺: Hexokinase, DNA polymerase
Fe²⁺/Fe³⁺: Cytochromes, peroxidase
Cu²⁺: Cytochrome oxidase
Mn²⁺: Arginase

2. Coenzymes (organic molecules):

NAD⁺, NADP⁺ (from Vitamin B₃)
FAD (from Vitamin B₂)
Coenzyme A (from Vitamin B₅)
TPP (from Vitamin B₁)

JEE Concept: Vitamins and Coenzymes

Many B vitamins are precursors of coenzymes:

Vitamin	Coenzyme	Function
B₁ (Thiamine)	TPP	Decarboxylation
B₂ (Riboflavin)	FAD, FMN	Redox reactions
B₃ (Niacin)	NAD⁺, NADP⁺	Redox reactions
B₅ (Pantothenic acid)	Coenzyme A	Acyl transfer
B₆ (Pyridoxine)	PLP	Amino acid metabolism

This is why vitamin deficiencies affect enzyme function!

Example:

No Vitamin B₁ → No TPP → Pyruvate dehydrogenase inactive → Beriberi

JEE Tip: Vitamins are essential because they form coenzymes!

Active Site

Active site = Specific region of enzyme where substrate binds

Features:

3D cleft or pocket in protein
Complementary to substrate shape
Contains catalytic residues (amino acids that perform reaction)
Relatively small (few amino acids out of hundreds)
Specific for substrate

Enzyme-Substrate Interaction

Lock-and-Key Model (Emil Fischer, 1894)

Concept: Enzyme and substrate fit like lock and key

     Substrate
        ↓
    [Substrate shape]
        ↓
    ⚬⚬⚬  ← Active site (complementary)
   /     \
  Enzyme

Features:

Rigid enzyme active site
Exact fit with substrate
Like key fitting into lock

Limitation: Doesn’t explain enzyme flexibility

Induced Fit Model (Koshland, 1958)

Concept: Enzyme changes shape upon substrate binding

  Substrate
     ↓
 Before:
   ⚬⚬⚬  ← Active site (not exact match)
  /     \
 Enzyme

 After binding:
 Substrate
    ↓
   ⚬⚬⚬  ← Active site molds around substrate
  /     \
 Enzyme

Features:

Flexible enzyme
Conformation change upon substrate binding
Better fit AFTER binding
Like glove fitting hand (glove molds to hand shape)

Advantage: Explains:

How enzymes can accommodate similar substrates
Why transition state is stabilized
Enzyme specificity

Memory Trick: Models

Lock-and-Key:

Rigid (lock doesn’t change)
Pre-formed fit
Older model

Induced Fit:

Flexible (enzyme changes)
Induced by substrate
Modern, accepted model

Analogy:

Lock-and-Key = Key and lock (rigid)
Induced Fit = Glove and hand (flexible)

JEE Tip: Both models explain specificity, but induced fit is more accurate!

Mechanism of Enzyme Action

Steps in Enzyme Catalysis

Step 1: Substrate binds to active site

$$\text{E} + \text{S} \rightarrow \text{ES}$$

Step 2: Enzyme-substrate complex forms

$$\text{ES} \rightarrow \text{EP}$$

(transition state)

Step 3: Product forms and releases

$$\text{EP} \rightarrow \text{E} + \text{P}$$

Overall:

$$\boxed{\text{E} + \text{S} \leftrightarrow \text{ES} \rightarrow \text{E} + \text{P}}$$

Enzyme is regenerated - acts as catalyst!

Interactive Demo: Visualize Enzyme Catalysis

See how enzymes lower activation energy and facilitate reactions step-by-step.

Energy Diagram

Energy
  ↑
  |        Without enzyme
  |           •
  |         /   \
  |        /     \
  |       /       \
  |      /         \
  |     /  With      \
  |    /  enzyme  •   \
  |   /         /  \   \
  |  /    ES  /    \EP  \
  |_/__________\____\____\____
     E+S              E+P

  Activation energy (Ea):
  - Without enzyme: High
  - With enzyme: LOW (ES complex)

Key point:

Enzyme lowers activation energy (Ea)
Does NOT change ΔG (overall energy change)
Makes reaction faster (more molecules have required energy)

Enzyme Specificity

Types of Specificity

1. Absolute Specificity:

Acts on ONE substrate only
Very strict
Example: Urease - only urea (nothing else)

2. Group Specificity:

Acts on molecules with specific functional group
Example: Alcohol dehydrogenase - all alcohols

3. Linkage Specificity:

Acts on specific type of bond
Example:
- Proteases - peptide bonds
- Lipases - ester bonds
- Amylases - α(1→4) glycosidic bonds

4. Stereochemical Specificity:

Distinguishes between stereoisomers
Example:
- L-amino acid oxidase - only L-amino acids (not D)
- Maltase - α-glucose linkages (not β)

JEE Question: Specificity

Q: Explain why maltase can hydrolyze maltose but not cellobiose, even though both are disaccharides of glucose.

Solution:

Maltose:

α-D-Glucose (C1) — O — β-D-Glucose (C4)
        α(1→4) linkage

Cellobiose:

β-D-Glucose (C1) — O — β-D-Glucose (C4)
        β(1→4) linkage

Key difference: α vs β glycosidic linkage

Maltase enzyme:

Specific for α(1→4) linkages
Active site complementary to α-linkage geometry
Cannot accommodate β-linkage (wrong orientation)

Result:

Maltase hydrolyzes maltose (α-linkage) ✓
Maltase does NOT hydrolyze cellobiose (β-linkage) ✗

Stereochemical specificity:

Even though both are Glu-Glu disaccharides
Different 3D geometry of linkage
Enzyme distinguishes α from β

This is why:

We can digest starch (α-linkages, maltase works)
We cannot digest cellulose (β-linkages, no enzyme)

JEE Principle: Enzyme specificity depends on 3D shape of substrate!

Factors Affecting Enzyme Activity

1. Temperature

Effect:

Low temperature: Slow reaction (low kinetic energy)
Optimum temperature: Maximum activity (usually 37°C for human enzymes)
High temperature: Denaturation (protein unfolds, loses activity)

Graph:

Activity
  ↑
  |        •
  |       / \
  |      /   \
  |     /     \  ← Denaturation
  |    /       \
  |   /         \___
  |__/______________\____
     0°   37°   60°  Temp (°C)
         (opt)

Why denaturation?

Heat breaks H-bonds in protein
Enzyme unfolds
Active site destroyed
Irreversible loss of activity

Q10 value:

Reaction rate doubles for every 10°C rise
Valid only before denaturation

2. pH

Effect:

Each enzyme has optimum pH
Activity decreases above or below optimum
Extreme pH: denaturation

Examples:

Enzyme	Optimum pH	Location
Pepsin	1.5-2.0	Stomach
Amylase	6.7-7.0	Saliva, pancreas
Trypsin	7.5-8.5	Small intestine
Arginase	9.5-10.0	Liver

Why pH matters:

1. Active site ionization:

Amino acids in active site have ionizable groups
pH changes their charge
Wrong charge → substrate can’t bind

2. Substrate ionization:

Substrate charge changes with pH
Affects binding to enzyme

3. Protein structure:

Extreme pH disrupts ionic bonds
Protein denatures

3. Substrate Concentration [S]

Effect:

Low [S]:

Rate proportional to [S]
Rate = k[S]
First order kinetics

High [S]:

Enzyme saturated
Rate constant (maximum)
Zero order kinetics

Michaelis-Menten Graph:

Velocity (V)
  ↑
  |          Vmax ─────────
  |               /
  |             /
  |           /
  |         /
  |       /  ← Vmax/2
  |     /
  |___/__________________
         Km        [S]

Km (Michaelis constant):

[S] at which V = Vmax/2
Measure of enzyme-substrate affinity
Low Km = high affinity (tight binding)
High Km = low affinity (weak binding)

4. Enzyme Concentration [E]

Effect:

At constant [S], rate proportional to [E]
More enzyme → more active sites → faster reaction
Linear relationship (until [S] becomes limiting)

Enzyme Inhibition

Competitive Inhibition

Inhibitor resembles substrate:

Competes for same active site
Reversible binding

Mechanism:

  Substrate         Inhibitor
      ↓                ↓
   ⚬⚬⚬          or  ⚬⚬⚬
  /     \            /    \
 Enzyme             Enzyme
 (active)          (inactive)

Effect:

Km increases (appears weaker substrate binding)
Vmax unchanged (can be overcome by high [S])

Example:

Malonate inhibits succinate dehydrogenase
Malonate resembles succinate (substrate)

Overcoming:

Increase [S]
High substrate outcompetes inhibitor

Non-competitive Inhibition

Inhibitor binds to different site:

Allosteric site (not active site)
Changes enzyme shape
Active site distorted

Mechanism:

  Inhibitor
      ↓
   ⚬⚬⚬  ← Distorted active site
  /  |  \
 Enzyme

Effect:

Vmax decreases (fewer functional enzymes)
Km unchanged (substrate binding not affected)

Example:

Heavy metals (Hg²⁺, Pb²⁺)
React with -SH groups
Distort enzyme structure

Cannot be overcome by increasing [S]

Irreversible Inhibition

Inhibitor permanently inactivates enzyme:

Covalent binding to active site
Enzyme destroyed

Examples:

Nerve gases (DFP, Sarin)
- Inhibit acetylcholinesterase
- Covalent bond with serine in active site
- Fatal (nerve signals disrupted)
Penicillin
- Inhibits bacterial cell wall synthesis enzyme
- Binds covalently
- Bacteria die

JEE Concept: Inhibition Types

Comparison:

Type	Binding Site	Reversible?	Effect on Vmax	Effect on Km	Overcome by [S]?
Competitive	Active site	Yes	Unchanged	Increases	Yes
Non-competitive	Allosteric	Yes	Decreases	Unchanged	No
Irreversible	Active site (usually)	No	Decreases	Variable	No

Memory:

Competitive:

“Competes for site, overcome by Concentration (of substrate)”
Km up, Vmax same

Non-competitive:

“No competition, No overcoming”
Vmax down, Km same

JEE Tip: Competitive changes Km, Non-competitive changes Vmax!

Nomenclature of Enzymes

Common Names (Old System)

Based on substrate or function:

Pepsin (digests proteins in stomach)
Trypsin (digests proteins)
Urease (hydrolyzes urea)
Amylase (breaks down amylose/starch)

Problem: No systematic pattern

Systematic Names (IUPAC)

Format: Substrate + type of reaction + -ase

6 classes based on reaction type:

1. Oxidoreductases:

Transfer electrons (oxidation-reduction)
Example: Alcohol dehydrogenase

2. Transferases:

Transfer groups (methyl, acyl, phosphate)
Example: Aminotransferases

3. Hydrolases:

Hydrolysis reactions (add water, break bonds)
Example: Esterases, peptidases, lipases

4. Lyases:

Add/remove groups to form double bonds
Example: Decarboxylases

5. Isomerases:

Rearrangement (isomerization)
Example: Glucose-6-phosphate isomerase

6. Ligases:

Join molecules (using ATP energy)
Example: DNA ligase

Common Mistakes to Avoid

Mistake #1: Enzyme Consumed in Reaction

Wrong: “Enzyme is used up in reaction”

Correct: Enzyme is catalyst

Not consumed
Regenerated after each cycle
Can work on many substrate molecules

JEE Fact: One enzyme molecule can process thousands of substrate molecules per second!

Mistake #2: Enzymes Change ΔG

Wrong: “Enzymes make unfavorable reactions favorable”

Correct: Enzymes:

Lower activation energy (Ea)
Speed up reaction
Do NOT change ΔG (equilibrium position)
Cannot make thermodynamically unfavorable reaction occur

JEE Principle: Enzymes affect rate, not equilibrium!

Mistake #3: All Enzymes are Proteins

Wrong: “All enzymes are proteins”

Correct: Almost all enzymes are proteins

Exception: Ribozymes (RNA enzymes)
Example: Ribosomal RNA has catalytic activity
But for JEE: assume enzymes are proteins

JEE Tip: If question says “enzyme,” think protein!

Practice Problems

Level 1: Foundation (NCERT)

Problem 1: Definition

Q: What are enzymes? How do they differ from inorganic catalysts?

Solution:

Enzymes:

Biological catalysts (speed up biochemical reactions)
Protein in nature
High specificity

Comparison:

Feature	Enzymes	Inorganic Catalysts
Nature	Protein (biological)	Metal, metal oxide
Specificity	Very high (one substrate)	Low (many substrates)
Conditions	Mild (37°C, pH 7)	Harsh (high T, P)
Efficiency	Very high (10⁶-10¹² times)	Moderate
Sensitivity	Sensitive (denatured by heat)	Stable
Regulation	Can be regulated (inhibitors)	No regulation

JEE Example:

Enzyme (sucrase) breaks sucrose at 37°C
Inorganic catalyst (H₂SO₄) needs heating

Problem 2: Active Site

Q: What is an active site? Why is it important?

Solution:

Active site:

Specific region of enzyme where substrate binds
3D pocket/cleft in protein structure
Contains catalytic amino acids

Characteristics:

Complementary shape to substrate
Small region (few amino acids)
Flexible (induced fit model)
Contains residues for catalysis

Importance:

1. Specificity:

Shape determines which substrate fits
Like lock-and-key
Wrong substrate won’t fit

2. Catalysis:

Amino acids in active site perform reaction
Stabilize transition state
Lower activation energy

3. Regulation:

Inhibitors block active site
Controls enzyme activity

Damage to active site = loss of activity!

Level 2: JEE Main

Problem 3: Temperature Effect

Q: Explain why enzyme activity increases with temperature initially but decreases at high temperature.

Solution:

Initial increase (0-37°C):

Kinetic theory:

Higher temperature → more kinetic energy
More collisions between E and S
More ES complexes form
Rate increases

Rule: Rate doubles for every 10°C rise (Q₁₀ = 2)

Maximum at optimum (37°C for human enzymes):

Highest activity
Best balance of speed and stability

Decrease at high temperature (>40-50°C):

Denaturation occurs:

Heat breaks H-bonds in protein
Enzyme unfolds
Active site destroyed
Cannot bind substrate
Activity lost

Graph:

Activity
  ↑
  |        •  ← Optimum (37°C)
  |       / \
  |      /   \
  |     /     \___  ← Denaturation
  |____/___________\____
     0°    37°   60°  Temp

Key point:

Low temp: Reversible (enzyme reactivated on warming)
High temp: Irreversible (permanent damage)

JEE Application:

Cooking destroys enzymes (why food doesn’t spoil when cooked)
Fever affects enzyme function (why high fever is dangerous)

Problem 4: Inhibition Type

Q: A competitive inhibitor increases Km but doesn’t change Vmax. Explain.

Solution:

Competitive inhibitor:

Resembles substrate
Competes for same active site
Reversible binding

Effect on Km:

Km = apparent affinity for substrate

Without inhibitor:

E + S ⇌ ES → Products

With inhibitor:

E + S ⇌ ES → Products
  +
  I
  ↓
 EI (inactive)

Less free enzyme available for substrate:

Appears as if substrate binds weaker
Km increases (need more [S] to reach half Vmax)

Effect on Vmax:

At very high [S]:

Substrate outcompetes inhibitor
All enzyme eventually as ES
Same maximum rate achieved
Vmax unchanged

Conclusion:

Competitive inhibition is overcome by high [S]
Km increases (apparent affinity decreases)
Vmax stays same (eventually reaches max)

JEE Analogy:

Parking lot with limited spots
Cars (S) and trucks (I) compete for spots
More cars → eventually all spots filled with cars (Vmax reached)
But takes more cars to fill half the spots (Km increased)

Level 3: JEE Advanced

Problem 5: Multi-concept

Q: Explain why: (a) Enzymes are specific for substrates (b) Heavy metals are toxic (c) Vitamin deficiencies affect enzyme function

Solutions:

(a) Enzyme specificity:

Lock-and-key / Induced fit:

Active site has specific 3D shape
Complementary to substrate
Wrong substrate doesn’t fit

Example: Maltase

Specific for α(1→4) linkage (maltose)
Cannot hydrolyze β(1→4) (cellobiose)
Even tiny difference in geometry prevents binding

Reasons:

Shape complementarity
Charge distribution (electrostatic interactions)
H-bonding pattern
Hydrophobic/hydrophilic regions

Result: High substrate specificity

(b) Heavy metals toxicity:

Mechanism:

Heavy metals (Hg²⁺, Pb²⁺, Cd²⁺):

React with -SH groups (cysteine residues)
Form covalent bonds
Disrupt disulfide bridges

Consequences:

Enzyme denaturation
- Protein structure distorted
- Active site destroyed
- Loss of activity
Cofactor displacement
- Heavy metals replace Mg²⁺, Zn²⁺
- But don’t function properly
Irreversible binding
- Permanent damage
- Cannot be easily removed

Examples:

Mercury poisoning (Minamata disease)
Lead poisoning (affects brain enzymes)

Treatment:

Chelating agents (EDTA, dimercaprol)
Bind heavy metals, allow excretion

(c) Vitamin deficiency affects enzymes:

Many vitamins are coenzyme precursors:

Examples:

Vitamin B₁ (Thiamine) → TPP (coenzyme)

Needed for pyruvate dehydrogenase
Deficiency → Enzyme inactive → Beriberi

Vitamin B₂ (Riboflavin) → FAD (coenzyme)

Needed for many redox enzymes
Deficiency → Energy metabolism affected

Vitamin B₃ (Niacin) → NAD⁺/NADP⁺

Essential for redox reactions
Deficiency → Pellagra

Without coenzyme:

$$\text{Apoenzyme} + \text{Cofactor} = \text{Holoenzyme (active)}$$

Missing cofactor:

$$\text{Apoenzyme} \text{ (alone)} = \text{INACTIVE}$$

Conclusion:

Vitamins essential for enzyme function
Vitamin deficiency = enzyme deficiency
Disrupts metabolism → disease

JEE Insight: This connects vitamins, enzymes, and disease - high-yield concept!

Quick Revision Box

Topic	Key Points	JEE Fact
Definition	Biological protein catalysts	Speed up reactions, not consumed
Active Site	Substrate binding region	Determines specificity
Models	Lock-and-key, Induced fit	Induced fit more accurate
Cofactors	Metal ions or coenzymes	Many from vitamins
Temperature	Optimum ~37°C	Denatures at high temp
pH	Each enzyme has optimum	Affects ionization
Competitive	Inhibitor at active site	Km↑, Vmax same, overcome by [S]
Non-competitive	Inhibitor at allosteric site	Vmax↓, Km same, can’t overcome
Specificity	High substrate specificity	Shape, charge complementarity

Teacher’s Summary

Key Takeaways

1. Enzymes as Catalysts:

Biological catalysts (proteins)
Not consumed in reaction
Lower activation energy (Ea)
Highly specific (one enzyme, one reaction usually)
Work at mild conditions (37°C, pH 7)

2. Structure-Function:

Active site = substrate binding region
Induced fit model (flexible, molds to substrate)
Holoenzyme = Apoenzyme + Cofactor

3. Factors Affecting Activity (HIGH-YIELD):

Temperature:

Optimum: 37°C
High temp: Denaturation (irreversible)

pH:

Each enzyme has optimum
Extreme pH: denaturation

[S] and [E]:

Low [S]: rate ∝ [S]
High [S]: rate constant (Vmax)

4. Enzyme Inhibition (MASTER THIS):

Competitive:

At active site
Km increases, Vmax unchanged
Overcome by high [S]

Non-competitive:

At allosteric site
Vmax decreases, Km unchanged
Cannot overcome

5. Specificity:

Lock-and-key / Induced fit
Shape complementarity
Stereochemical specificity (L vs D, α vs β)

6. Connections:

Vitamins → Coenzymes
Heavy metals → Toxicity (-SH binding)
Temperature/pH → Denaturation

“Enzymes are nature’s catalysts - exquisitely specific, incredibly efficient, but delicate!”

This completes the Biomolecules chapter!

The Hook: Nature’s Catalysts - Millions Times Faster

The Core Concept

What are Enzymes?

Enzyme Structure

Components

Cofactors

Active Site

Enzyme-Substrate Interaction

Lock-and-Key Model (Emil Fischer, 1894)

Induced Fit Model (Koshland, 1958)

Mechanism of Enzyme Action

Steps in Enzyme Catalysis

Interactive Demo: Visualize Enzyme Catalysis

Energy Diagram

Enzyme Specificity

Types of Specificity

Factors Affecting Enzyme Activity

1. Temperature

2. pH

3. Substrate Concentration [S]

4. Enzyme Concentration [E]

Enzyme Inhibition

Competitive Inhibition

Non-competitive Inhibition

Irreversible Inhibition

Nomenclature of Enzymes

Common Names (Old System)

Systematic Names (IUPAC)

Common Mistakes to Avoid

Practice Problems

Level 1: Foundation (NCERT)

Level 2: JEE Main

Level 3: JEE Advanced

Quick Revision Box

Teacher’s Summary

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