Course:

CELL BIOCHEMISTRY (BOM 113/4)

Type:

Minor

Total Units:

3 (3 x 1 h lecture per week)

No. of Lectures:

21 (Part II)

Lecturers:

Assoc. Prof. Dr. K. Sudesh Kumar

Room 402, Block G09A School of
Biological Sciences
Tel: 04-6534367

Email: ksudesh@usm.my

Objectives:

Introduction to cells, organelles, molecules;

structure & function relationship; working of
a cell

Course contents are divided into 4 major sections:

1. Concept of a cell: Prokaryote, Eukaryote,
Animal and Plant Cells; cellular organelles
2. Cell components (Structure & Function):
Water and buffer, amino acids and protein, fatty
acids and lipid, monosaccharide and
carbohydrate, nucleotides and DNA, RNA,
membrane and membrane transport

Objectives:

3. Enzymology: Enzyme as catalysts,

enzyme kinetics, inhibitor effects,
enzyme regulations, allosteric
enzymes

4. Energetics and metabolisms:

Bioenergetic principles,
carbohydrate metabolisms
(glycolysis), TCA cycle, pentose
phosphate pathway,
gluconeogenesis and
photosynthesis, fatty acid oxidation
3

Course evaluation: Test/Quiz/Assignment/Practical: 30%

Final Exam: 70%

Learning Objectives
The basics of metabolism, enzymes as catalyst of metabolic
pathways and energetics principles in general.
Details on vital metabolic pathways and processes such as
glycolysis, fermentation of sugars, pentose phosphate
pathway, gluconeogenesis, citric acid cycle, electron
transport chain, oxidative phosphorylation, fatty acid
oxidation and biosynthesis, photosynthesis as well as their
regulation will be covered in detail.

ENERGY FLOW

Thermodynamics
Thermodynamics is a
funny subject. The first time
you go through it, you don't
understand it at all. The
second time you go through
it, you think you understand
it, except for one or two
small points. The third time
you go through it, you know
you don't understand it, but
by that time you are so
used to it, it doesn't bother
you any more.
-Arnold Sommerfeld-

Arnold Johannes Wilhelm Sommerfeld

(December 5, 1868 April 26, 1951) was a German
theoretical physicist who pioneered
developments in atomic and quantum physics, as
well as educated and groomed a large number of
students for the new era of theoretical physics.
7 He
introduced the fine-structure constant into quantum
mechanics.

Energy
Defined as capacity to do work
Two types of energy: Kinetic (movement) and
Potential (stored) energy
To understand energy flow, we need to know two
things: the quantity of available energy and the
usefulness of the energy: these are described in
the laws of thermodynamics
emphasizes causation
emphasizes change
8

Kinds of changes made by living cells

A. Changes in chemical bonds:
1) biosynthesis
2) growth of new cells
3) maintenance of existing cells:
i) large molecules are easily damaged
ii) most large molecules are recycled
(a) protein-->amino acids-->protein-->
(b) DNA/RNA -->nucleotide-->DNA/RNA-->
iii) Synthesis of large molecules needs some energy
iv) cell needs change with time
(a) different proteins needed during mitosis
(b) food change, need different enzymes

Kinds of changes made by living cells

4) synthesis of small molecules
i) 6CO2 + 6H2O ---> glucose + 6O2
(a) requires 18 ATP (chemical energy)
(b) requires 12 NADPH (reducing power)
ii) major fraction of cell energy used here for the
synthesis of small molecules

10

Kinds of changes made by living cells

B. Mechanical work (changes in location)
1) move cell relative to environment; examples:
i) flagellated cell (sperm)
ii) ciliated epithelium in bronchial tubes
iii) muscles
http://en.wikipedia.org/wiki/Spermatozoon
iv) movement of chromosomes
v) cyclosis in plant cells
vi) movement mRNA relative to ribosome
vii) movement of plant parts (venus flytrap)

http://en.wikipedia.org/wiki/Venus_Flytrap

11

Kinds of changes made by living cells

C. Concentration work (changes in concentration across membrane)
1. move molecules or ions against chemical gradient
i) [glucose] in cell >[glucose] outside cell
(a) must pump glucose in
ii) pumping H+, Na+, Ca2+ etc
iii) eg: light driven accumulation of H+ in thylakoid lumen
(b) pumped in against concentration and electron gradient
iv) at rest, cell's energy used to pump substances

12

Kinds of changes made by living cells

D. Electrical work (changes in cell's potential)
i) cell membranes have electrical potential (~70mV inside neg.)
ii) move ions against electron gradient
iii) must consider both the concentration and the electron grad.
iv) electron gradient drive ATP synthesis in bacteria, chloroplasts &
mitochondria
v) ATP used to make electrical potential
(1) example: electric eel has electric organ
(a) layers of cells called electroplaxes
(b) each cell has potential of 0.15 V
(c) cells are in series
(d) total potential generated: >400 V

http://en.wikipedia.org/wiki/Electric_eel

13

Kinds of changes made by living cells

E. Heat energy
i) homeotherms (birds & mammels)
a) must maintain relatively constant temperature
b) heat production is the major use of food energy
F. Bioluminescent work
i) fireflies, some bacteria
ii) can be large energetic drain
(1) large energy output indicates important function
Which part of you consumes
the most amount of energy?

http://en.wikipedia.org/wiki/Brain#Brain_energy_consumption
14

Brain energy consumption

Although the brain represents only 2% of the body weight, it
receives 15% of the cardiac output, 20% of total body oxygen
consumption, and 25% of total body glucose utilization.
The energy consumption for the brain to simply survive is 0.1
Calories per minute, while this value can be as high as 1.5
Calories per minute (100W) during crossword puzzle-solving.
The demands of the brain limit its size in many species. Molossid
bats and the Vespertilionid Nyctalus spp. have brains that have
been reduced from the ancestral form to invest in wing-size for
the sake of maneuverability.
This contrasts with fruit bats, which require more advanced
neural structures and do not pursue their prey.
15

Using Energy
A. Environmental energy available at earths surface:

Energy Source

Cal/cm2/yr

Solar radiation

260,000

Lightening

Radioactivity

0.8

Volcanoes

0.13

Photosynthesis

100

Only source of large amounts of Energy = light

16

Our main source of

energy!

http://www.solarviews.com/eng/sun.htm

17

Using Energy
B. Many organisms capture light energy
1. Plants, Photosynthetic bacteria, Photosynthetic protists
2. Called "phototrophs"
a. use light energy to make all molecules required for
life from inorganic precursors like CO2 and H2O
b. function as chemotrophs in dark
i) use chemical energy stored in light (starch)
C. Chemotroph organisms get energy from source other than light
1. All animals, fungi, many protists and most bacteria are
chemotrophs
2. Release energy from compounds they take up (via fermentation,
glycolisis, respiration)
18

Flow of energy in biosphere

1) Begins with nuclear fusion reactions in sun
2) Light
3) Small amount captured by phototrophs and "stored" as
molecules [some energy lost as heat/entropy (S)]
4) Chemotrophs release energy captured by phototrophs
and stored as chemical [Some energy lost as
heat/entropy during conversion]
5) Flow is: from sun light (+heat/S) --> molecules of
phototrophs (+heat/S) --> molecules of chemotrophs
(+heat/S) --> heat/S

19

Flow of matter in biosphere

1) Energy usually stored as chemical bonds
2) Flow of matter accompanies energy flow
3) Energy enters without matter (light)
4) Energy leaves without matter (IR radiation/S)
5) In biosphere, energy mainly in the form of chemical bonds associated
with matter:
i) matter cycles between phototrophs and chemotrophs
a. carbon
b. oxygen
c. nitrogen
d. hydrogen
e. etc.
ii) two groups (phototrophs & chemotrophs) live in symbiotic
relationship
6) Ecology deals with macroscopic energy cycles
7) Cell biochemistry deals with microscopic energy transactions at cellular
20
and molecular level

The Laws of Thermodynamics

ALL processes that occur in the universe are subject
to the 3 basic laws of thermodynamics. These laws
define the basic properties and behavior of energy.
1st law: Energy can neither be created or destroyed (law
of conservation of energy)
2nd law: ALL naturally occurring processes proceed in a
direction that leads to a minimum potential energy
level (or maximum entropy) (means that, when energy
is converted from one form to another, the amount of
useful energy decreases)
3rd law: At absolute zero temperature (0K) the
entropy is zero
21

First Law of Thermodynamics

1.

Energy cannot be created or destroyed, only converted

from one form to another
a. for a given system:
(1) energy out = energy in - energy stored, or
(2) energy stored = energy in - energy out
b. amount energy stored in a system/molecule = E
c. not concerned with actual value of E
d. very concerned with "changes in E"
e. E = E2 - E1 = Ein - Eout

22

First Law of Thermodynamics

2. Example:
a. C6H12O6 + 6O2 --> 6CO2 + 6H2O + energy
b. Burn 1 mole glucose; 673 kcal energy released
i) 1 mole glucose has 673 more kcal energy than 6 moles
of CO2 + 6 moles of H2O
ii) reaction has E = -673 kcal
iii) negative E => a decrease in internal energy of
molecules as glucose & O2 converted to CO2 & H2O

23

First Law of Thermodynamics

c. The reverse reaction
i) 6CO2 + 6H2O + energy --> C6H12O6 + 6O2
ii) this reaction: E = +673 kcal
iii) positive E => internal energy when CO2 & H2O converted to
glucose & O2
(a) glucose + O2 has internal energy released to CO2 + H2O
d. So: amount of internal energy released when glucose is burned is
exactly equal to amount of energy stored in glucose when
the sugar is synthesized (energy neither created nor
destroyed = 1st Law of Thermodynamics)

24

Second Law of Thermodynamics

1. The value of S (entropy) in the universe is positive for
every real process or reaction
2. Universe becomes more random with every reaction
that occurs
3. But, not always convenient to deal with predicting the
spontaneity of reaction in the test tube in terms of
universal entropy
4. Use G (free energy) to predict spontaneity of reaction in
closed systems
Lets look at the concept of entropy first
25

The concept of entropy, S

Entropy, represented by the symbol S, is a measure
of the randomness or orderliness of the energy and
matter in a system.
The more random, disordered, disorganized, or
chaotic the system, the higher the entropy.
Only organized, non-random/focused/concentrated
energy is useful (can be made to do work)
Entropy of a substance increases with its volume. (eg.
gas molecules in occupying all the volume available
to it maximizes its entropy) (What will happen to our
universe?)
26

Entropy and the 2nd law of

thermodynamics
Disorder spreads
through the universe,
and life alone battles
against it

G. Evelyn Hutchinson (1903-1991) 27

The Concept of Free Energy Arises from

a Combination of the First & Second
Laws
Free energy (G) - energy available to do work; defined
by J. Willard Gibbs, American chemist (1878)

Together the 1st & 2nd laws of thermodynamics show that the
energy of the universe is constant, but that entropy (S)
continues to increase toward a maximum

Gibbs combined concepts inherent in 1st & 2nd Laws to

get equation:H = G + TS where:
1. G is the change in free energy (the change during a
process in energy available to do work)
2. H - change in enthalpy (total energy content of system;
equivalent to E for our purposes)
3. T - absolute temperature (K; K = C + 273)
4. S - change in entropy of system
28

H = G + TS
Total energy change = sum of changes in useful energy
available (G) & unavailable (TS) for work
Rearrange to G = H - TS; can predict direction in
which, process will proceed & the extent to which the
process will occur
Spontaneous process has - G (exergonic) & proceeds
toward state of lower free energy; such a process is
thermodynamically favored
Non-spontaneous process, +G (endergonic); cannot
occur spontaneously; it is thermodynamically
unfavorable; make it go by coupling to high -G (energyreleasing) reaction.
29

Josiah Willard Gibbs

Josiah Willard Gibbs
(February 11, 1839 April 28,
1903) was an American
theoretical physicist, chemist,
and mathematician. One of
the greatest American
scientists of all time, he
devised much of the
theoretical foundation for
chemical thermodynamics as
well as physical chemistry.
http://en.wikipedia.org/wiki/Josiah_Willard_Gibbs

30

Gibbs free energy (G) - energy available to do

work at constant temperature and pressure
High Energy

- G

+ G

High Energy

- G

Low Energy

+ G

- G

+ G

Low Energy

The coupling of energy-producing reactions and energy-requiring

reactions is a central feature in the metabolism of all organisms. In
catabolism, oxidative reactions are coupled to the endergonic
production of ATP by phosphorylation of ADP.
In anabolism, the exergonic hydrolysis of the high-energy bond of
ATP releases the energy needed to drive endergonic reductive
31
reactions.

ATP functions as
energy currency
in processes that
release energy
and processes
that use energy

32

Free energy changes in

chemical reactions
All cell chemical reactions are reversible, so
one must consider forward & reverse reactions
at the same time
Law of Mass Action - reaction rate
proportional to reactant concentration
(Ex.: A + B <> C + D)
1. Forward reaction rate - proportional to A & B
concentrations (k1[A][B]); k1 : forward rate
constant
2. Reverse reaction rate - proportional to C & D
concentrations (k2[C][D]); k2 : reverse rate
33
constant

 All reactions proceed toward equilibrium (can

be slow), where forward & reverse rates are
equal
1. At equilibrium, the same number of A & B
molecules are converted into C & D molecules
per unit time as are formed from them
2. k1[A][B] = k2[C][D] at equilibrium - allows
calculation of equilibrium constant (Keq) by
combining k1 & k2 into Keq (Keq = k1/k2 =
[C][D]/[A][B])
3. Gives predictable ratio of product to reactant
concentrations at equilibrium & allows
prediction of reaction direction (forward or
reverse) under given set of conditions
34

4. Ratio can be determined experimentally by

measuring product & reactant concentrations
at equilibrium
5. If Keq > 1 & initial concentration ratios = 1 >
products increase at expense of reactants
6. If Keq < 1 & initial concentration ratios = 1 >
reactants increase at expense of products
7. Net direction in which, reaction is proceeding
at any moment depends on relative
concentrations of all participating molecules
& can be predicted if Keq is known.
35

Free energy changes in

chemical reactions
-G favors product formation; +G favors
reactant formation; a greater - G reaction is
farther from equilibrium & the system can
perform more work
- G when total free energy of reactants >
total free energy of products (+G - vice versa)
As reaction proceeds, difference in free energy
content between reactants & products
decreases (G gets less negative)
At equilibrium, no further work can be obtained
& the free energy difference, G is 0
36

Standard States for Free-Energy Changes

1.

2.

3.

4.

5.

G for a given reaction depends on the reaction mixture

present at a given time, thus it is not useful for comparing
the energetics of various reactions
To allow such comparisons & various types of calculations,
scientists began to consider reactions under standard
conditions
G0: G at standard conditions, in which the concentration
of all reactants in solution is set at 1 M. Can be related to the
equilibrium constant of a reaction by the equation:
G0 = -RT ln Keq
G0 - modified standard states (for biochemical
applications) ; 25C [298K], 1 atm pressure,
reactant/product concentrations, 1 M, pH 7 [H+ ion
concentration at 10-7 M])
prime indicates a modified standard state, where [H] is
37
changed from 1M to 110-7M)

Relationship between Keq & G0

(G0: spoken as delta G zero prime)

G0 = -RT ln Keq = - 2.3 RT log Keq

1. R = gas constant (1.987 cal/mol/K or 8.31
J/mol/K); T = absolute temperature (K)
2. G0 & K'eq indicate that standard conditions include
pH7 (for biological conditions), while G0 & Keq indicate
standard conditions in 1 M H+ (pH = 0.0!)
3. If Keq > 1, then G0 negative (spontaneous) & if Keq < 1,
then G0 positive (nonspontaneous) under standard
conditions
4. So, G predicts the spontaneity of a reaction.
5. G predicts only direction of reaction, NOT reaction rate!
6. G says nothing about how fast a reaction will go.
38

Sample calculation 1

Hydrolysis of ATP at pH 7, 25C, yields ADP,

monohydrogen phosphate ion (Pi) and H+
ATP + H2O ADP + Pi + H+

The value for Keq for this reaction is 2.23 105

G0 = -RT ln Keq
= - (8.31 J/mol/K)(273+25K)(12.32)
= - 3.05 104 J/mol
= - 30.5 kJ/mol

= - 7.29 kcal/mol (because 1 kJ = 0.239 kcal)

- G0 indicates that the hydrolysis reaction of ATP
to ADP is a spontaneous process in which energy is
39
released.

Sample calculation 2
The G0 for a reaction at 25C is -15 kJ/mol. What is
the equilibrium constant for the reaction? (R=8.314;
T=273)

Keq = e-G/RT
= e-[-15,000/(8.314)(298)]
= e6.05
= 426

40

Facts of Origins of Life

Life originated 600 million years ago and continental drift occurred
200 million years ago creating five continents.
Mammals evolved 140 million years ago, Hominids that is the
human type, evolved 26 million years ago but modern humans only
arrived on the scene some 200,000 years ago.
Humans migrated and colonized the world only in the last 50,000
years.
The spoken language is some 10,000 years old while writing
evolved only a few thousand years ago.
All this phenomenal progress has been achieved only within the
short span of 200 to 400 generations, that is in just 10,000 to 5,000
years.
from Dr. APJ Abdul Kalams Convocation Acceptance Speech at the
Universiti Sains Malaysia, Penang 30/Aug/2008 : Pinang, MalaysiaEvolution of
Enlightened citizenship
41

B
I
O
E
N
E
R
G
E
T
I
C
S

B
I
O
E
N
E
R
G
E
T
I
C
S

The Cell is a Open System

No exchange of matter
or energy

Exchange of energy
may occur

Exchange of matter and/or

energy may occur

43

44

45

Bioenergetics
LIFE requires constant supply of ENERGY &
MATERIAL
Derived from the food that we consume
Involves a series of biochemical reactions
Electron donors transfer energy to electron
acceptors
Oxidation-reduction reactions are
fundamental to the extraction of energy from
our food
46

METABOLISM
The molecules of carbohydrates, fats,
and proteins taken into an organism are
processed in a variety of ways.
This is called METABOLISM, which is
the sum total of the chemical reactions
of biomolecules in an organism.
Metabolism consists of two processes:
CATABOLISM and ANABOLISM
47

CATABOLISM

The breakdown of large molecules to smaller

ones
Is an oxidative process that releases energy
Free energy in substrate/reactant is greater
than the free energy in product (exergonic)
Electrons are transferred to various acceptor
molecules
3 stages:
1) Macromolecule-->monomer (eg. Starch to
glucose)
Energy is produced!
2) Monomer --> acetyl-CoA
48
3) Acetyl-CoA --> CO2 and H2O

ANABOLISM
The process of building macromolecules from
small molecules
Is a reductive process that requires energy
The free energy in reactants are less than the
free energy in product (endergonic)
Involves acceptance of electrons from a variety
of donors
3 stages:
1. CO2 + H2O + energy --> acetyl-CoA
2. Acetyl-CoA --> monomers (fatty
acid/sugar/amino acid)
Energy is used!
3. Monomers --> macromolecules (eg. Polymers 49
and complex molecules)

Nutrients (fats, carbohydrates, proteins) taken into an

organism are processed in a variety of ways

50

Oxidation and Reduction Transfer of

Electrons
Oxidation - reduction (redox) reactions
must occur simultaneously
Involve change in the electronic state of
reactants
Both oxidation & reduction are
reversible; both anabolic & catabolic
pathways involves redox reactions

51

Oxidation
Oxidation - reaction involving loss of one
or more electrons; ex.: metallic iron (Fe0)
to ferrous state (Fe2+) via loss of electron
pair; since an electron acceptor is
needed, it is accompanied by reduction
The molecule oxidized is called a
reducing agent & becomes more positive
Reaction is reversible; oxidized molecule
can pick up electrons & return to earlier
state (Fe2+ -> Fe0)
52

Reduction
Reduction - a reaction involving a gain
of one or more electrons; ex.: Fe2+ to
Fe0;
The reduced molecule is called an
oxidizing agent;
Since electron donor is needed, it is
accompanied by oxidation
Example: Fe0 + Cu2+ <> Fe2+ + Cu0;
Cu2+ is oxidizing agent, Fe0 is reducing
agent
53

Redox of organic compounds

The oxidation & reduction of organic compounds
during cellular metabolism involve carbon atoms
that are covalently bonded to other atoms
When a pair of electrons is shared by 2 different
atoms, the electrons are more strongly attracted
to one of the 2 atoms of the polarized bond
In CH bonds, C pulls electron more strongly &
is in reduced state
In CO & CN bonds, electrons are attracted by
electronegative atom, thus C is in oxidized state
54

Oxidation state of carbon atom

Carbon atom has a number of oxidation
states, since it can share a number of
valence electrons (4)
Ranging from CH4 (fully reduced) to CO2
(fully oxidized);
Oxidation state of a carbon atom in an
organic molecule is a measure of the
molecule's free energy content
One can roughly determine oxidation
state by counting number of H vs. O & N
atoms per C atom
55

Capture & utilization of energy

Chemical fuels (hydrocarbons) are highly
reduced organic compounds; energy is
released when they are burned in presence of
O2 converting them to more oxidized states
(CO, CO2)
As carbon is more reduced, more chemical
work can be done in cell - fats (HCH) have
more energy per unit weight than
carbohydrates (HCOH)
The more H atoms that can be stripped from a
"fuel" molecule, the more ATP can be made
56

The role of coenzymes in biologically

important oxidation-reduction reactions
NADH: the reduced form of nicotinamide adenine
dinucleotide
NADH is oxidized to NAD+, which is an electron
acceptor in redox reactions
Important coenzyme in many reactions
Is a derivative of nicotinic acid (also called niacin),
one of the B-complex vitamins
Its structure has 3 parts: a nicotinamide ring, an
adenine ring and two sugars. The phosphate groups
of the two sugars are linked together.
57

NAD+

NADH

The functional
group involved in
the reaction

A similar compound is NADPH

(the oxidized form is NADP+).
It differs from NADH by having
an additional phosphate group
attached to one of the58ribose
sugars

 Another important electron acceptor is FAD (flavin

adenine dinucleotide), which is the oxidized form of
FADH2. They contain the flavin group which are
derived from vitamin riboflavin B2.
Oxidation of nutrients to provide energy for an
organism cannot take place without reduction of
some electron acceptor such as NAD+ and FAD.
The ultimate electron acceptor in aerobic oxidation is
oxygen.

59

60

NAD+,
FAD and
ATP are
constantly
recycled

61

62

Activation of metabolic pathways

and the role of coenzyme A
In order to produce and use energy efficiently and in
a controlled manner, cells carry out multistep
metabolic processes
We will see later that the partial anaerobic breakdown
of glucose (glycolysis) and the complete aerobic
breakdown of glucose to CO2 and H2O requires many
steps
A step frequently encountered in metabolism is the
process of activation. This is to make the
substrate/metabolite more reactive.
Coenzyme A is a compound that is used for this
purpose.
63

Thiol group is the

reactive portion of
CoA

64

High energy thioester/

covalent bond

Acetyl-CoA

65

Energy Changes and Electron Transfer

in Metabolism
Learning Outcome: By now you should
Know What Are Standard States for Free-Energy Changes
Know What Is a Modified Standard State for Biochemical
Applications
Know What Is Metabolism
Know How Are Oxidation and Reduction Involved in
Metabolism
Know How Are Coenzymes Used in Biologically Important
OxidationReduction Reactions
Know How Are the Production and Use of Energy Coupled?
To Know How Is Coenzyme A Involved in Activation of
Metabolic Pathways?

66