The Story of Catalysis

Why Catalysis? What is a Catalyst? A Petrochemical Application How Catalysts Work

Why Catalysis?
Need to make chemicals faster
Most Reactions are too slow to be useful...

What is a Catalyst?
Catalysts speed up a chemical reaction without being used up...
Catalyst-Reactants

Catalyst + Reactants
Catalyst + Products

The Nature of Enzyme Catalysis

Enzyme provides a catalytic surface This surface stabilizes transition state Transformed transition state to product
B A A B
Catalytic surface
Juang RH (2004) BCbasics

Stickase
Substrate Transition state Product

If enzyme just binds substrate then there will be no further reaction

Enzyme not only recognizes substrate, but also induces the formation of transition state
Adapted from Nelson & Cox (2000) Lehninger Principles of Biochemistry (3e) p.252

Examples of Reactions w/o Catalysis

Plants & animals decompose to oil & coal Oxygen in the air mixes with iron to form rust

1 week to several months

Hydrogen Peroxide forms a chemical reaction with your body

Millions of years

Seconds

Ways to Make Chemicals Faster

Temperature
Disadvantage--Too hot!

Pressure
Disadvantage--Cause Explosions

Add other Chemicals

Disadvantage--Separate chemicals

Catalysts!!!!
Disadvantage--Costly

Enzyme Stabilizes Transition State

Energy change
Energy required (no catalysis)

ST
Energy decreases (under catalysis)

S
ES

EST

EP
Reaction direction T = Transition state

Whats the difference?

Adapted from Alberts et al (2002) Molecular Biology of the Cell (4e) p.166

Active Site Is a Deep Buried Pocket

Why energy required to reach transition state is lower in the active site?

It is a magic pocket

+
CoE (1) (4)
(2)

(3)

(1) Stabilizes transition (2) Expels water (3) Reactive groups (4) Coenzyme helps
Juang RH (2004) BCbasics

Enzyme Active Site Is Deeper than Ab Binding

Instead, active site on enzyme also recognizes substrate, but actually complementally fits the transition state and stabilized it.

Ag binding site on Ab binds to Ag complementally, no further reaction occurs.

Adapted from Nelson & Cox (2000) Lehninger Principles of Biochemistry (3e) p.252

Active Site Avoids the Influence of Water

+ -

Preventing the influence of water sustains the formation of stable ionic bonds
Adapted from Alberts et al (2002) Molecular Biology of the Cell (4e) p.115

Reaction Rates
9

R e a c tio n R a te s

E n e rg y

W ith o u t C a ta ly s t W ith C a ta ly s t

0 0 2 4 6 8 10 12 14

T im e

Cyclar... An Example of A Catalytic Process

2 ( C - C - C ) + Catalyst Propane + Catalyst

Benzene

Propane is unreactive Benzene is in higher demand than propane

How Does Cyclar Work?

Surface Area!!
Catalysts are not solid Catalysts are porous

What Does Surface Area Do?

Increases the amount of available reaction sites

2 propanes

Benzene Reactive Sites

Benzene product

Do Catalysts Live Forever?

NO!!!
Catalysts can last from 2 hours to 2 years
They can die from:
--poisons which contaminate the catalyst --large molecules which cover the catalyst --over heating, over pressurizing --crumbling/crushed

Conclusions
Catalysts are reusable u Catalysts help increase reaction rates u Catalysts work using surface area
u

Who Uses Catalyst?

Oil Industries
Your body (Enzymes) Chemical/Pharmaceutical Companies Catalytic Converters in your car

Examples of Reactions w/Catalysts

Propane + Catalyst = Benzene + Catalyst
C-C-C + = +

Food

+ Enzymes = Small Molecules + Enzymes Proteins + Fats Simple Sugars

Examples of Catalysis
Basic substances like ashes catalyze the burning of sugar

A non-reactive substance like water turns reactive when mixed with catalyst

What Makes a Catalyst Work?

Surface Area!!!!

Steel Wool has a large surface area, so it oxidizes easily

Enzyme Inhibition (Mechanism)

I

Competitive
Substrate

Non-competitive
S

Uncompetitive
E
I

Cartoon Guide

E
S
I

E
Compete for Inhibitor active site

Different site

Equation and Description

E + S ES E + P + I EI
[I] binds to free [E] only, and competes with [S]; increasing [S] overcomes Inhibition by [I].

E + S ES E + P + + I I EI + S EIS
[I] binds to free [E] or [ES] complex; Increasing [S] can not overcome [I] inhibition.

E + S ES E + P + I EIS
[I] binds to [ES] complex only, increasing [S] favors the inhibition by [I].
Juang RH (2004) BCbasics

Enzyme Inhibition (Plots)

I
vo

Competitive
Vmax

Non-competitive
Vmax

Uncompetitive
Vmax

Direct Plots

vo

Vmax

Vmax

Km Km

[S], mM

Km = Km

[S], mM

Km Km

[S], mM

Double Reciprocal

Vmax unchanged Km increased 1/vo

Intersect at Y axis

Vmax decreased Km unchanged 1/vo

Both Vmax & Km decreased 1/vo

Two parallel lines

1/ Vmax 1/[S]

Intersect at X axis

1/ Vmax 1/[S] 1/Km

1/ Vmax 1/[S]

1/Km

1/Km

Juang RH (2004) BCbasics

Competitive Inhibition
Product Substrate Succinate C-OOC-H C-OOH-C-H Competitive Inhibitor Glutarate C-OOH-C-H Malonate C-OOH-C-H Oxalate C-OOC-OO-

C-H
C-OO-

H-C-H
C-OO-

H-C-H
H-C-H C-OO-

C-OO-

Succinate Dehydrogenase
Adapted from Kleinsmith & Kish (1995) Principles of Cell and Molecular Biology (2e) p.49

Sulfa Drug Is Competitive Inhibitor

Domagk (1939)

Para-aminobenzoic acid (PABA)

H2NPrecursor

-COOH

Bacteria needs PABA for the biosynthesis of folic acid

Folic acid

Tetrahydrofolic acid

H2N-

-SONH2

Sulfa drugs has similar structure with PABA, and inhibit bacteria growth.

Sulfanilamide Sulfa drug (anti-inflammation)

Adapted from Bohinski (1987) Modern Concepts in Biochemistry (5e) p.197

Enzyme Inhibitors Are Extensively Used

Sulfa drug (anti-inflammation)

Pseudo substrate competitive inhibitor

Protease inhibitor

Alzheimer's disease

Plaques in brain contains protein inhibitor

HIV protease is critical to life cycle of HIV

HIV protease (homodimer): inhibitor is used to treat AIDS
subunit 1 subunit 2

Human aspartyl protease:

(monodimer)

As p

As p

Symmetry

domain 1 domain 2 Not

As p

As p Juang RH (2004) BCbasics

symmetry

HIV protease vs Aspartyl protease

HIV Protease inhibitor is used in treating AIDS

HIV protease (homodimer)

subunit 1
Asp

subunit 2
Asp

Symmetric dimer

domain 1
Asp

domain 2
Asp

Asymmetric monomer

Aspartyl protease (monomer)

Juang RH (2004) BCbasics

Chymotrypsin Has A Site for Specificity

O O N C C N C C N C C N C C R H R
OC Ser

Specificity Catalytic Site Site

Active Site
Juang RH (2004) BCbasics

Specificity of Ser-Protease Family

Trypsin
cut at Lys, Arg O O CNCCN C C C C NH3 + COOC Asp

Chymotrypsin
cut at Trp, Phe, Tyr
O O CNCCN C

Elastase
cut at Ala, Gly
O O CNCCN CH3
Shallow and non-polar pocket

Deep and negatively charged pocket

Non-polar pocket

Active Site

Juang RH (2004) BCbasics

Stereo Specificity

A
sp3

B
Enzyme surface

The tetrahedral structure of carbon orbital has rigid steric strain which makes the basic building unit of protein conformation

These two triangles are not identical

Juang RH (2004) BCbasics

Control Points of Gene Regulation

Transcription DNA
ribosome mRNA RNA Processing RNA Transport RNA Degradation

DNA
5 process mRNA 3 mature mRNA 3 tail proteins

Translation
proteins

cap 5

Activity
Proteolysis

Prokaryotics

Post-translational control

Eukaryotics
Juang RH (2004) BCbasics

Regulation of Enzyme Activity

Inhibitor
or

Proteolysis

o
S

I
inhibitor

x
I

proteolysis

Feedback regulation

Phosophorylation

o
S

R regulator effector

x
R

P
S

o
(+) P

phosphorylation
Juang RH (2004) BCbasics

Signal transduction

x
(-)

A or
Regulatory subunit
S

cAMP or calmodulin

Cascade Amplification of Signals

Cascade

nS

1 Enzyme

nP

Juang RH (2004) BCbasics

How to Separate These Objects

Shape Size Density

2 3 4 5 6

7 8 9

10 11

12

wood stone cotton wood wood cotton stone wood stone cotton stone cotton

1
Size

2 3
6
7 8
Density
cotton

Shape

4
8

4 5 6
9
10 11

wood

7
12

stone

Sieving different sizes

Different sedimentation

Different rolling speed

Juang RH (2004) BCbasics

Basic Principles of Protein Purification

Cell
Small molecule
Amino acid, Sugar, Nucleotides, etc

Organelle
Homogenization

Macromolecule
Nucleic acid

Protein

Carbohydrate

(Lipid)

Cell Debris

Ammonium sulfate
Size Gel filtration, SDS-PAGE, Ultrafiltration Charge

fractionation
Polarity Affinity

Ion exchange, Reverse phase Affinity Chromatofocusing, chromatography, chromatography, HIC, Disc-PAGE, Hydroxyapatite Salting-out Isoelectric focusing
Juang RH (2004) BCbasics