HMM1414/Chapter 2

CHAPTER 2

CHEMICAL FOUNDATION OF LIFE

2.1 Properties of Water

(Refer Campbell page 47 – 53)

• Origin of life - water.

• Terrestrial organisms also tied to water:
o Cells surrounded by water.
o Cell content: 70 – 95% water.
o Reactants.

2.1.1 Polarity of Water & Hydrogen

Bonding

• Water - polar molecule

• Two hydrogen atoms form polar covalent
bond with an oxygen atom.
° Oxygen more electronegative than hydrogen.
° Oxygen – 2 partial negative charge (δ -).
° Hydrogen - partial positive charge (δ +).
• Attraction between water molecules:
° Negative regions attracted to positive regions,
forming hydrogen bonds .
° Maximum of four bonds per molecule.

(Refer Figure 3.2, Campbell page 47)

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2.1.2 Heat Capacity

• Specific heat - amount of heat that must be

absorbed or lost for 1 g of a substance to change
its temperature by 1°C.
° Specific heat of water = 1 cal/g/°C.
• Water resists/minimizes temperature
fluctuations because of its high specific heat.
° Absorbs or releases relatively large quantity of
heat for each degree of temperature change.
• Importance of high specific heat:
• Water heats up more slowly & holds its
temperature longer.
• Major component of living things.
• Medium for biochemical reactions.
• Aquatic environments have relatively stable
temperatures.

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2.1.3 Heat of Vaporization

• Heat of vaporization – quantity of heat a

liquid must absorb for 1 g of it to be converted
from liquid to gas.
° Water - high heat of vaporization
• As a liquid evaporates, the surface of the liquid
that remains behind cools - evaporative
cooling.
Importance:
• Cooling mechanism during sweating &
transpiration.
• Large amount of heat is lost during evaporation
with only minimal loss of water from body.
• Evaporative cooling moderates temperature in
lakes and ponds.

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2.1.4 Density

• Water less dense as solid than as liquid.

° Water expands as it solidifies.
° In liquid form, not all molecules are H-bonded
to 4 other molecules.
° H-bond constantly break & reform - allows
molecules some freedom of movement.
• When temperature falls below 0°C, hydrogen
bonds are locked into a crystalline lattice.
o Each molecule is hydrogen-bonded to 4
other molecules.
o Molecules are further apart from each
other.
o Fewer molecules than an equal volume of
liquid water, making ice less dense.
(Refer Figure 3.5, Campbell, page 51)
• As ice melts, some H bonds break, & water
molecules move closer to one another – density
increases.
○ Maximum density at 4°C.
○ Above 4°C, density decreases as water
expands & the molecules move faster.
• Ice is ≈ 10% less dense than water at 4°C.

• Thus, ice floats on the cool water below.

• Importance:
° If ice sank, all ponds, lakes, and ocean would
freeze solid.
° Insulation

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2.1.5 Solvent Properties

• Water - versatile solvent because of its

polarity.
° Readily forms H bonds with charged and polar
covalent molecules.
° E.g., when a crystal of salt (NaCl) is placed in
water (Refer Figure 3.6, Campbell, page 51)
 Na+ ↔ oxygen.
 Cl− ↔ hydrogen.
• Ion surrounded by a hydration shell.

• Polar molecules also soluble in water because

they form H bonds with water.
• Large molecules can dissolve in water if they
have ionic and polar regions.

• Any substance that has an affinity for water is

hydrophilic (water-loving).
° These substances are dominated by ionic or
polar bonds.
• Substances that have no affinity for water are
hydrophobic (water-fearing).
° These substances are nonionic and have non
polar covalent bonds.

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2.1.6 Cohesion and Adhesion

•Cohesion – the binding together of like

molecules, often by hydrogen bonds.
• Cohesion among water molecules enables
transport of water against gravity in plants.
• Adhesion, attraction between different kinds
of molecules, also plays a role - enables water to
adhere to the wall of the vessels.
• Surface tension, a measure of the force
necessary to stretch or break the surface of a
liquid, is related to cohesion.
° Water has a greater surface tension because
hydrogen bonds among surface water
molecules resist stretching or breaking the
surface.
• Importance:
 Some animals can stand, walk, or
run on water.
 Capillary action

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2.1.7 Viscosity

• Viscosity – a measure of a fluid’s resistance to

flow.
• Water – low viscosity.
• Importance:
 Transport system of living organisms.
Example: blood is mostly water – can flow
easily through vessels
 Plants depend on flow of water in xylem
& phloem to transport substances.
 Less energy used by aquatic organisms
when swimming in water.

2.1.8 Other Properties

• Colorless & transparent

 Transmission of sunlight possible –
photosynthesis for aquatic plants.
• Difficult to compress
 Structural agent – hydrostatic
skeleton.
• Involved in many chemical reactions
 Raw material for photosynthesis
 Digestive reactions – breaks down
food molecules by hydrolysis

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2.2 The Chemistry of Carbon

2.2.1 Definition of Organic Compounds

• Organic compounds – compound containing

carbon.
• Cells – consist mostly of carbon-based
compounds.
•Carbon can form large, complex, and diverse
molecules.
•Accounts for diversity of biological molecules
→ great diversity of living things.
• Proteins, DNA, carbohydrates are composed of
carbon atoms bonded to each other, & to atoms
of other elements - hydrogen (H), oxygen (O),
nitrogen (N), sulfur (S), and phosphorus (P).

2.2.2 Structure of Carbon and

Formation of Covalent Bonds in
Carbon

• Carbon atom - 2 electrons in 1st electron shell

& 4 in 2nd shell.
• To complete its valence shell, it share
electrons with other atoms in four covalent
bonds.
• This tetravalence by carbon makes large,
complex molecules possible.

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• Can form covalent bonds with many different

elements.
• The valences of carbon and its partners -
building code that governs architecture of
organic molecules.

• Carbon chains form skeletons of most organic

molecules.
• Skeletons vary in length and shape.
(See Figure 4.5, Campbell, page 61)

• Skeletons may have double bonds & may have

atoms of other elements bonded to it.
• Variation in architecture of organic molecules
can also be seen in isomers.

• Distinctive properties of organic molecules

also depend on molecular components attached
to it.
 Functional groups - components of
organic molecules commonly involved in
chemical reactions.
 The number and arrangement of
functional groups help give each molecule its
unique properties.
(See Figure 4.10, Campbell, page 64 – 65)

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2.2.3 Macromolecules

• Small organic molecules are joined together to

form macromolecules.
 Consist of thousands of covalently bonded
atoms.
• Four major classes: carbohydrates, lipids,
proteins, and nucleic acids.

• Carbohydrates, proteins, and nucleic acids—

polymers.
° Polymer - long molecule consisting of many
similar or identical building blocks
(monomers) linked by covalent bonds.
• Monomers connected by covalent bonds by
condensation reaction or dehydration
reaction.
• Polymers are disassembled by hydrolysis.

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2.3 Important Organic Compounds

2.3.1 Carbohydrates
• Sugars and their polymers.
• Monosaccharides - single/simple sugars.

• Disaccharides - double sugars.

• Polysaccharides - polymers of
monosaccharides.

(a) Monosaccharides

• Molecular formula - multiple of the unit CH2O.

° For example, glucose - C6H12O6.

• Have a carbonyl group (>C=O) and multiple

hydroxyl groups (—OH).

• Classification based on:

(1) Location of carbonyl group - aldose or
ketose.
 Example: Glucose, galactose, riboses –
aldose; fructose,ribulose – ketose
(2) Number of carbons in carbon skeleton.
 Hexoses – 6C sugars - glucose.

 Pentoses – 5C sugars – ribose.

 Trioses – 3C sugars - glyceraldehyde.

(See Figure 5.3, Campbell, page 70)

• Monosaccharides in aqueous solutions form

rings.
(See Figure 5.4, Campbell, page 71)

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• Functions:
1. Major fuel for cellular work.
2. Raw material for synthesis of other monomers.

(b) Disaccharides

• Form from two monosaccharides via

dehydration.
• Covalent bond - glycosidic linkage.
° Maltose - glucose + glucose. (See Figure 5.5 (a),
Campbell, page 71)
° Sucrose - glucose + fructose. (See Figure 5.5 ba),
Campbell, page 71)
° Lactose- glucose + galactose.

(c) Polysaccharides

• Polymers of many monosaccharides joined by

glycosidic linkages.
• Functions:
1. Storage.
2. Building materials/structural roles.

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(i) Storage Polysaccharides

Starch
• Storage polysaccharide of plants.
° Monomers (glucose) joined by 1–4 linkages.
° Two forms:
 Amylose - unbranched and forms a helix.

 Amylopectin - branched and more complex.

(See Figure 5.6 (a), Campbell, page 72)

• Plants store surplus glucose as starch
granules within plastids, including chloroplasts.

Glycogen
• Storage polysaccharide in
animals.
° Highly branched like amylopectin.
(See Figure 5.6 (b), Campbell, page 72)
° Stored liver and muscles.

(ii) Structural Polysaccharides

Cellulose
• Major component of plant cell
wall.
• Polymer of glucose, but with
different glycosidic linkages.
° Difference based on two slightly different ring
structures for glucose: α - and β -glucose.
(See Figure 5.7 (a), Campbell, page 73)
• Starch: α -glucose monomers. (See
Figure 5.7 (b), Campbell, page 73)

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• Cellulose: β -glucose monomers.

(See Figure 5.7 (b), Campbell, page 73)
• Starch (α 1-4 glycosidic linkages) – helical
structure
• Cellulose (β 1-4 glycosidic linkages) – straight
structures.
 H atoms on one strand can form hydrogen
bonds with OH groups on other strands.
 Plant cell walls - parallel cellulose molecules
held together into microfibrils - strong
building materials for plants. (See Figure 5.8,
Campbell, page 73)
• Enzymes that hydrolyze starch
cannot hydrolyze the β -linkages in
cellulose.
° Cellulose in human food eliminated in feces as
“insoluble fiber.”
• Some microbes use cellulase to
digest cellulose.
• Many eukaryotic herbivores, have
symbiotic relationships with
cellulolytic microbes. (See Figure
5.9, Campbell, page 74)
• Some fungi can also digest
cellulose.

Chitin
• In exoskeletons of arthropods.
° Glucose monomer has nitrogen-containing
appendage. (See Figure 5.10 (a), Campbell, page 74)
° Pure chitin is leathery but can be hardened by
the addition of calcium carbonate.
• Provides structural support for the cell walls of
many fungi.

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• Also used to make surgical threads.

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2.3.2 Lipid

• Do not form polymers.

• Hydrophobic – little or no affinity for water
 Consists mostly of hydrocarbons → form non-
polar covalent bonds.
• Most biologically important lipids: fats,
phospholipids, and steroids.

Fats
• Synthesized from glycerol and fatty acids.
° Glycerol: 3C alcohol with a hydroxyl group
attached to each carbon.
° Fatty acid: consists of a carboxyl group
attached to a long carbon skeleton.
° Non-polar C—H bonds in hydrocarbon skeleton
make fats hydrophobic.
° Fats separate from water because water
molecules hydrogen bond to one another and
exclude the fats.
• Three fatty acids joined to glycerol via ester
linkage, forming triacylglycerol, or
triglyceride.
(See Figure 5.11, Campbell, page 75)
• Fatty acids - same or different.
• Fatty acids vary in length (number of carbons)
and in the number and locations of double bonds.

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° Saturated fatty acid: no C-C double bonds;

maximum number of H atom.
° Unsaturated fatty acid: one or more C-C
double bonds.
• Saturated fatty acid - straight chain
• Unsaturated fatty acid - has a kink wherever
there is a double bond.
(See Figure 5.12, Campbell. Page 75)
• Saturated fats - made from saturated fatty
acids.
° Example, animal fats - butter.
° Solid at room temperature.
• Unsaturated fats - from unsaturated fatty
acids.
° Example: plant (olive oil) and fish fats (cod
liver oil)
° Liquid at room temperature = oils.
° Kinks prevent the molecules from packing
tightly enough to solidify at room temperature.
° “Hydrogenated vegetable oils” - unsaturated
fats synthetically converted to saturated fats
by addition of hydrogen.
 Example: Peanut butter/margarine.
° Diet rich in saturated fats - plaque deposits →
atherosclerosis.

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• Function of fats
1)Energy storage.
° 1 g of fat stores more than twice as much
energy as 1 gram of a polysaccharide.
° 38kJ per g.
° Fats – compact energy storage as compared
starch which is bulky.
° Plants store fats (oils) in seeds; human & other
mammals – adipose cells.

2) Adipose tissues cushion (shock absorber) vital

organs - kidneys.

3) Fat (subcutaneous) layer functions as

insulation.

4) Buoyancy for aquatic organisms.

° Fats – relatively low density

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Phospholipids
• Consists of two fatty acids attached to
glycerol and a phosphate group.
(See Figure 5.13, Campbell, page 76)
° Phosphate group - negative charge.
° Additional groups may be attached to the
phosphate groups.
• Fatty acid – hydrophobic tails.
• Phosphate group + attachments - hydrophilic
head.
• In water, they self-assemble into bilayers with
hydrophobic tails pointing toward interior.
(See Figure 5.14, Campbell, page 77)
° This type of structure = micelle.
• Major component of all cell membranes.
 Example:
Lecithin
Steroids
• Consist of carbon skeleton with four fused
rings.

(See Figure 5.15, Campbell, page 77)

• Cholesterol
 Precursor from which other steroids are
synthesized.
° Example hormones, including vertebrate sex
hormones.
 High levels in blood - cardiovascular disease.

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 Importance:
1. Raw material for manufacture of vitamin D.
2. Component of mammalian cell membranes –
strengthen membranes at high body
temperatures.

Steroid abuse:

• Example: anabolic steroids

• Synthetic androgens (male reproductive
hormones)
• Used by sportsmen to increase:
 Muscle mass (to “bulk up”)

 Physical strength

 Endurance

 Aggressiveness

• Signs of abuse:
 Mood swings

 Suicidal thoughts/suicide attempts

 Restlessness

 Fatigue & sleeplessness

 Loss of appetite

• Change in physical appearance:

 Males – baldness, impotence, shrinking testes,

& development of breasts.

 Females – growth of facial hair, deepening of

voice, & reduction in breast size.

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 Both sexes – acne, oily hair & skin, cysts,

jaundice (yellowing of skin), swelling of feet &
ankles, aching joints, bad breath, nervousness,
& trembling.

• Risk from taking steroids:

1. Possibility of heart attack & stroke.
2. Increase in anger, hostility, and violent
behavior.
3. Increase risk of getting AIDS from sharing
needles with people that use injectable
steroids.

2.3.3 Protein

• 50% of dry mass of most cells.

• Functions:
1) Structural support
2) Storage
3) Transport
4) Cellular signaling
5) Movement
6) Defense against foreign substances
7) Enzymes - catalysts.

(See Table 5.1, Campbell, page 78)

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• Structurally complex molecules: 3-D shape or

conformation.
• Built using same set of 20 amino acid
monomers.
• Consists of one or more polypeptides.
• Polypeptides – polymers of amino acids.

Amino acids

• Have carboxyl and amino groups.

• Four components attached to asymmetric α -
carbon atom of amino acid:

(4)
R α -carbon
|
H–N–C– C=O
| | |
H H H
(3) (1) (2)

1. A hydrogen atom
2. A carboxyl group
3. An amino group
4. A variable R group (side chain)
 Different R groups characterize the 20
different amino acids.

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• Physical and chemical properties of R group

determine type of amino acid.
(See Figure 5.17, Campbell, page 79)
• Example:
1. Non-polar (hydrophobic) – glycine.
2. Polar (hydrophilic) – serine.
3. Charged (ionized):
a)Acidic (negative in charge) – Aspartic acid.
b)Basic (positive in charge) – Lysine.

• Properties of amino acid:

1. Amphoteric – has both acidic & basic
properties.
2. Form zwitterions in water – carries positive
charge (-NH3+) and negative charge (-COO-) at
pH 7.4.

• Amino & carboxyl groups ionize in

solution:
 -COOH ↔ -COO- + H+ (donates H+)
 -NH2 + H+ ↔ -NH3+ (accepts H+)

• Amino acids linked together by peptide

bonds forming a polypeptide chain.
• Polypeptides size - a few to many thousand
monomers.

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• Each polypeptide has a unique linear sequence

of amino acids.

Levels of Protein Structure

•Functional protein - one or more polypeptides
→ twisted, folded, & coiled into a unique shape.
• Order of amino acids determines three-
dimensional conformation of the protein.
• Protein’s specific conformation determines its
function.

Primary Structure
(See Figure 5.20, Campbell, page 82)

• Refers to a protein’s unique sequence of

amino acids.
 Example: Lysozyme consists of 129 amino
acids.
 Precise primary structure determined by
inherited genetic information.

Secondary Structure
(See Figure 5.20, Campbell, page 80)

• Refers to repetitive coiling or folding of

the polypeptide backbone due to hydrogen
bond formation between peptide linkages.
• Typical secondary structures:

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 Coils (α -helix); or
 Folds (β -pleated sheets).

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Tertiary structure
(See Figure 5.20, Campbell, page 83)

• Refers to overall shape of polypeptide chain

resulting from interactions between the side
chains (R) of the various amino acids.
Interaction includes:
 Weak interactions:
1. H bonds between polar and/or charged areas
2. Ionic bonds between charged R groups
3. Hydrophobic & van der Waals interactions
among hydrophobic (non-polar) R groups.
 Strong covalent bonds - disulfide bridges.

Quaternary structure
(See Figure 5.20, Campbell, page 83)

• Refers to the overall protein structure

resulting from the aggregation of two or
more polypeptide subunits.
 Collagen: fibrous protein of three
polypeptides, super-coiled like a rope.
 Hemoglobin: globular protein.
 Four polypeptide subunits: two alpha and
two beta chains.
 Each subunit has a heme component with
an iron atom that binds oxygen.

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Fibrous and Globular Proteins

• Fibrous protein
 Elongated molecule
 Dominant structure - secondary structure (α -
helix or β -pleated sheets).
 Insoluble.
 Plays structural or supportive role in body, &
involved in movement.
 Often have regular repeating structures.
 Example: Structural proteins
i. Keratin – helix of two helices (2 pairs of α -
helices wound around one another)
consisting of 7 repeating amino acids.
ii. Silk – β -pleated sheets only (glycine-alanine-
serine repeats)

• Globular protein
 Compact and spherical proteins.
 Many are folded – hydrophobic groups on inside
of molecule &hydrophilic groups face outwards
– thus, soluble in water.
 Examples: Non-structural proteins – enzymes,
transport protein, receptor proteins, and
myoglobin.

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Factors determining protein

conformation

1) Primary structure (sequence of amino acids)

2) Physical & chemical conditions of protein’s
environment.
 Alterations in pH, salt concentration,
temperature, or other factors - denatures a
protein.
 Denaturation – process in which a protein
unravels & loses its native conformation,
thereby becoming biologically inactive.
3) Organic solvent - polypeptide chain refolds,
causing hydrophobic regions to face outward
towards solvent.
4) Heat - disrupts the weak interactions that
stabilize conformation.
(See Figure 5.22, Campbell, page 85)

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2.3.4 Nucleic Acid

• Amino acid sequence of a polypeptide is

programmed by unit of inheritance, the gene.
• Gene consists of DNA.

Role of Nucleic Acids

• Two types:
Ribonucleic acid (RNA): and
Deoxyribonucleic acid (DNA).

• DNA provides directions for its own replication.

• DNA also directs RNA synthesis and, through
RNA, controls protein synthesis.
• The flow of genetic information (The Central
Dogma):

DNA → RNA → protein

• Protein synthesis occurs on ribosomes.

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Structure of Nucleic Acids

(See Figure 5.26, Campbell, page 87)

• Nucleic acids are polymers called

polynucleotides.
•Polynucleotide is made of nucleotide
monomers.
• Each nucleotide consists of a
(i.) Nitrogenous base
(ii.) Pentose sugar
(iii.) Phosphate group
 Pentose sugar +nitrogenous base =
nucleoside .
 Nucleoside + phosphate = nucleotide
(nucleoside monophsphate)
• Nitrogen bases - rings of carbon and nitrogen.
Two types:
(i.) Pyrimidines
 Have single six-membered ring.
 Three types: cytosine (C),
thymine (T), and uracil (U).
(ii.) Purines
 Have a six-membered ring joined to a five-
membered ring.
 Two types: adenine (A) and guanine (G).
(See Figure 5.26, Campbell. Page 87)

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• In RNA, pentose sugar is ribose; in DNA it is

deoxyribose.
 Difference: Deoxyribose
lacks oxygen atom on
carbon 2.
(See Figure 5.26 (c), Campbell, page 87)

Synthesis of polynucleotides
• Adjacent nucleotides
joined by covalent bonds
called phosphodiester
linkages.
 Formed between —OH group
on 3’ C of one nucleotide & phosphate on 5’ C
of the next.
 This forms repeating
backbone of sugar-
phosphate units, with
appendages consisting of
nitrogenous bases =
polynucleotide chain.
 One end has a phosphate
attached to a 5’ C = 5’ end.
 The other end has a
hydroxyl group on a 3’ C= 3’
end.
(See Figure 5.27, Campbell, page 88)
• Sequence of bases along a DNA or mRNA
polymer is unique for each gene.

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 Specifies order of amino acids (primary

structure) of a protein, which in turn
determines its 3-D conformation & function.

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The DNA Double Helix

(See Figure 5.27, Campbell, page 88)

• DNA - two polynucleotide strands →double

helix.
 Double helix structure first
proposed in 1953 by James
Watson and Francis Crick.
• Sugar-phosphate backbones on outside of
helix.
 Backbones run in opposite 5’ -> 3’
directions from each other =
antiparallel.
• Pairs of nitrogenous bases, one from each
strand, connect polynucleotide chains with
hydrogen bonds.
 Chargaff’s Rule of Complementary
base pairing:
 Adenine (A) with Thymine (T)
 Guanine (G) with Cytosine (C).
 2 H bonds between A-T; 3 H bonds
between C-G.
• Strands are complementary.

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RNA
(Structure of mRNA & tRNA, refer Campbell, page
320 -321)
• Single polynucleotide chain.
•Nitrogenous base: Adenine (A), Uracil (U),
Guanine (G), & Cytosine.
• Pentose sugar : ribose

mRNA – messenger RNA:

 Delivers information encoded in
genes from DNA to ribosomes, where
information is decoded into protein.

tRNA – transfer RNA:

 Serves as adapter molecule in
protein synthesis.
 Translates mRNA codons into
amino acids.

rRNA – ribosomal RNA

 Structural component of ribosomes.
 Forms extensive secondary
structures.
 Plays active role in recognizing
conserved portions of mRNAs and tRNAs.
 Assist in protein synthesis.

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2.4 Techniques of Analysis

2.4.1 Chromatography

• Physical method to separate & to analyze

complex mixture.
• Involves a sample (or sample extract) being
dissolved in a mobile phase (which may be a gas,
liquid, or a supercritical fluid).

• Mobile phase is then forced through an

immobile, immiscible stationary phase.
• Phases chosen so that components of the
same sample have differing solubilities in each
phase.
• Component which is quite soluble in stationary
phase will take longer to travel through it than a
component which is not very soluble in
stationary phase but very soluble in mobile
phase.
• Due to differences in mobilities, sample
components will become separated from each
other as they travel through the stationary
phase.

Paper Chromatography (PC)

• Stationary phase is liquid soaked into a sheet

or strip of paper.

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• Mobile phase is a liquid solvent.

• Some components spend more time in the
stationary phase others.
• Components appear as separate spots spread
out on paper after drying or developing.

Retention Factor, Rf

• Rf - quantitative indication of how far a

particular compound travels in a particular
solvent.
• Rf value - a good indicator of whether an
unknown compound and a known compound are
similar, if not identical.
• If Rf value for unknown compound is
close/same as a known compound, then both
compounds are most likely similar or identical.

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• Rf = Distance the solute moves (D1)

Distance traveled by solvent front (D2).

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2.4.2 Electrophoresis

• Movement of an electrically charged

substance under the influence of an electric
field.
• Gel electrophoresis separates
macromolecules (nucleic acids or proteins) on
basis of their rate of movement through a gel in
an electrical field.
 Rate of movement depends on size, electrical
charge, and other physical properties of
macromolecules.

(See Figure 20.8, Campbell, page 393)

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2.4.3 X-ray diffraction

• X-ray diffraction - scattering of X-rays by atoms

of a crystal.
o Diffraction pattern shows structure of the
crystal.
• Can determine 3-D structure of a molecule.
o By determining distances between atoms
of molecules arranged in regular, repeating
crystalline structure.

• Technique: X-ray crystallography

o An X-ray beam of light is directed at
crystallized protein.
o Atoms of crystal diffract (deflect) X-rays
into an orderly array.
o Diffracted X-ray exposes photographic
film, producing a pattern of spots, known as an
X-ray diffraction pattern.

• Result:
o Using data from X-ray diffraction patterns,
such as a detailed mathematical analysis of
measurements of the spots, as well as amino
acid sequence determined by chemical
methods, a 3-D computer model of the protein
is built.

(See Figure 5.24, Campbell, page 86)

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2.4.4 Centrifugation

• Cell fractionation - to separate major

organelles of cells so their individual functions
can be studied.
 Uses an ultracentrifuge - can spin at up to
130,000 revolutions per minute and apply
forces of more than 1 million times gravity
(1,000,000 g).
• Fractionation begins with homogenization,
gently disrupting the cell.
• Homogenate is spun in a centrifuge to
separate heavier pieces into the pellet while
lighter particles remain in the supernatant.
° As process is repeated at higher speeds and
for longer durations, smaller and smaller
organelles can be collected in subsequent
pellets.
• Cell fractionation prepares isolates of specific
cell components.
• Enables functions of organelles to be
determined, especially by reactions or processes
catalyzed by their proteins.

(See Figure 6.5, Campbell, page 97)

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2.4.5 Microscopy

(a) Light Microscope (LM)

• In LM, visible light passes through specimen

and then through glass lenses.
° Lenses refract light, &image is magnified into
eye or onto a video screen.
• Vary in magnification and resolving power.
• Minimum resolution of a light microscope ≈
200 nanometers (nm) = small bacterium.
• Can magnify effectively to about 1,000 times
the size of actual specimen.
• Various techniques have enhanced contrast
and enabled particular cell components to be
stained or labelled.
• Can resolve individual cells, but not internal
anatomy, especially organelles.
(See Figure 6.3, Campbell, page 96)

(b) Electron Microscope (EM)

• Can resolve smaller structures.

• EM focuses a beam of electrons through the
specimen or onto its surface.
° Have finer resolution.
° Resolution could reach 0.002 nanometer (nm),
but practical limit is closer to about 2 nm.

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• Two types:
(i) Transmission electron microscopes
(TEMs)
 To study the
internal ultrastructure of cells.
° Electron beam is aimed through a thin section
of specimen.
° Image is focused and magnified by
electromagnets.
° To enhance contrast, the thin sections are
stained with atoms of heavy metals.
(ii) Scanning electron microscopes
(SEMs)
 To study surface structures.
° Sample surface covered with a thin film of
gold.
° Beam excites electrons on sample’s surface.
° These secondary electrons are collected and
focused on a screen.
° Result: 3-D image of specimen.
• EM reveals organelles impossible to resolve
with the LM.
° However, EM can only be used on dead cells.
• LM do not have as high a resolution, but can be
used to study live cells.

(See Figure 6.4, Campbell, page 96)

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2.5 Enzymes

• Catalyst - chemical agent that speeds up rate

of a reaction without being consumed by the
reaction.
• Enzyme – a catalytic protein speeds up rate of
reaction without being consumed by the
reaction.
• Endergonic reaction – chemical reaction
that consume energy.
o Products formed contain more energy than
reactants.
• Exergonic reaction – chemical reaction that
releases energy.
o Products formed contain less energy than
reactants.
(See Figure 8.6, Campbell, page 147)

2.5.1 Properties of Enzymes

1. Reaction catalyzed is specific.

Specificity results from its 3-D shape.
2. Not destroyed by reactions they catalyzed and
are reusable
3. Sensitive to high temperature and pH.
4. Catalyze a reaction in either direction
(reversible).
5. Can be inhibited.

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2.5.2 Catalysis and Activation Energy

• Chemical reaction involves bond breaking &

bond forming.
• For this to occur, reactant must absorb energy
from their surroundings.
• Formation of new bonds in product
accompanied by release of energy as heat.
 Products will have stable shapes with
lower energy.

• The initial energy that reactants must absorb

before a chemical reaction will start = free
energy of activation or activation energy
(EA).
• Activation energy necessary to enable
reactants overcome the energy barrier so that
reaction can proceed.
(See Figure 8.14, Campbell, page 151)

° At the summit, molecules are in an unstable

condition, the transition state.
° Reactant may absorb activation energy in the
form of heat from surroundings.
° When reactant have absorbed enough energy,
their bonds will break and they become
unstable and, thus, more reactive.

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° As the products are formed with new, stable

bonding, energy is released to surroundings.
• For some processes, EA is not high:
○ Thermal energy from room temperature is
enough for reactants to reach transition state.
• In many cases, EA is high:
○ Transition state is seldom reached and
reaction hardly proceeds at all.
○ Reaction will only occur if reactants are
heated.
• Proteins, DNA, and other complex organic
molecules are rich in free energy and can
overcome energy barrier.
° However, there is not enough energy in a
typical cell.
° Most organic molecules cannot make use of
this energy to overcome the barrier.
° They may absorb heat, but it would denature
proteins and kill cells.
• Enzymes speed reactions by lowering EA.
° Transition state can then be reached even at
moderate temperatures.
• Enzymes do not change the free energy (∆ G).
° They speed up reactions that would occur
eventually.
(See Figure 8.15, Campbell, page 152)

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2.5.3 Mechanism of Action and Kinetics

• Substrate - reactant that an enzyme acts on.

• Enzyme binds to substrate or substrates,
forming enzyme-substrate complex.
• Catalytic action of enzyme converts substrate
to product or products.
• Only a specific region of enzyme binds to
substrate by weak chemical bonds- the active
site.
° Active site is usually formed by only a few
amino acids.
(See Figure 8.16, Campbell, page 153)

• As substrate enters active site, interactions

between its chemical groups and amino acids of
enzyme cause an induced fit – change in shape
of active site of enzyme so that it binds more
closely to substrate.
• Substrates are held in active site by weak
interactions - hydrogen & ionic bonds.
° R groups of a few amino acids on active site
catalyze conversion of substrate to product.
° Product then leaves the active site.
(See Figure 8.17, Campbell, page 153)

• A single enzyme molecule can catalyze

thousands of reactions a second.

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2.5.4 Factors Affecting Enzyme Activity

(1) Substrate Concentration

• Rate of enzyme-catalyzed reaction increases

as substrate concentration increases until
reaction reaches a maximum rate.
• At high substrate concentrations, active sites
on all enzymes are filled.
° Enzyme is saturated.
•To increase productivity at this point - add
more enzyme.
Rate of reaction

High enzyme concentration

Low enzyme concentration

Substrate concentration

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 HMM1414/Chapter 2

(2) Enzyme
Concentration

• Rate of reaction increases as enzyme

concentration increases, provided no other
factors are limiting.
• Relationship only holds when pH, pressure, and
temperature are constant, and substrates are
in excess.
• Under these conditions, the more active sites
there are available, the more substrates can
be converted to products.
Rate of reaction

Enzyme concentration

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 HMM1414/Chapter 2

(3) Tempera
ture

• Rate of reaction increases with temperature up

to a maximum, the optimum temperature.
• As temperature increases, collisions between
substrates and active sites occur more
frequently as molecules move more rapidly.
• As temperature increases further, weak bonds
are disrupted, and enzyme denatures.
• Each enzyme has an optimal temperature:
 Most human enzymes: 35 – 40°C.

 Bacteria in hot springs contain enzymes with

optimal temperatures of 70°C or above.

(See Figure 8.18 (b), Campbell, page 155)

(4) pH

• Most enzymes active at narrow pH range:

° Optimal pH: 6 - 8 for most
enzymes.
• Many enzymes become inactive, and denatured,
if medium is very acidic or very basic.
 Substrates no longer fit into active

sites.

(See Figure 8.18 (b), Campbell, page 155)

(5) Cofactors
(6) Inhibitors

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Michaelis-Menten Constant, KM

 Constant represents substrate

concentration required to make reaction go at
half its maximum rate.
 Value always same for a particular
enzyme, but varies from one enzyme to another.
 Measures affinity of enzyme for its
substrate.

° Low KM: high affinity between

enzyme & substrate.
° High KM: low affinity – substrates
react less readily with enzyme.

Maximum rate
Rate of reaction

½ maximum rate

Substrate concentration

Michaelis-Menten constant

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2.5.5 Inhibitors

• Inhibitors – substances which interfere,

reduce, or destroy enzyme activity.
• Binding of inhibitors to enzymes prevents the
enzymes from catalyzing reactions.

Competitive Inhibitors

• Inhibitors that resemble substrates and

compete for binding to the active sites.
° Binding of inhibitor to active site prevents
enzyme from combining with substrate – no
products are generated.
° Competitive inhibition overcome by increasing
concentration of substrate.
° Reversible.

(See Figure 8.19 (b), Campbell, page 144)

Noncompetitive Inhibitors

• Inhibitors that retard enzymatic reactions by

binding to another part of the enzyme.
° Binding causes enzyme to change shape,
rendering active site less effective at
catalyzing reaction.
° Irreversible.
• Example: Toxins and poisons.

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Allosteric Regulation

• Refers to binding of a regulatory molecule to a

protein (enzyme) at a site (allosteric site)
that affects function of protein at a different
site (active site).
• Regulatory molecules - inhibitor or activator.
° Binding by these molecules can either inhibit
or stimulate (activate) enzyme activity.
• The binding of an activator stabilizes the
conformation of the enzyme so that the active
sites can function or function more effectively.
• The binding of an inhibitor stabilizes the
conformation of the enzyme so that the active
sites become inactive or have reduced
catalytic activity.

(See Figure 8.20 (a), Campbell, page 156)

° For example, ATP functions as inhibitor to

catabolic enzymes, inhibiting their activity (so
that less ATP are produced).
° ADP functions as an activator of the same
enzymes (so that more ATP are produced)

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Feedback Inhibition

• In feedback inhibition, an early step in a

metabolic pathway is switched off by
pathway’s final product.
° Product acts as an inhibitor of enzyme in that
step.
• Feedback inhibition prevents a cell from
wasting chemical resources by synthesizing
more product than is needed.
(See Figure 8.21, Campbell, page 157)

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2.5.6 Cofactors

• Non-protein enzyme helpers.

° Bind permanently or reversibly to the enzyme.

° Two types:

(i) Inorganic cofactors (activators):

2+ 2+ 3+
• Example: zinc (Zn ), iron (Fe or Fe ), and
copper (Cu+ or Cu2+).
• Helps bind together enzyme & substrate or
serve as catalytic centre of enzyme.
(ii) Organic cofactors (coenzymes)
 Example: Biotin, NAD, FAD, ATP.

 Transfers chemical group or atom from

active site of one enzyme to active site of

another enzyme.
 Sometimes, function of coenzyme carried by

a non-protein group of atoms attached to

enzyme, called prosthetic group.
 Example: Heme, flavin, and retinal

 Transfers chemical groups/atoms from

active site of enzyme to other substance.

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2.5.7 Nomenclature of Enzymes (IUB)

• Produced by the Nomenclature Committee of

the International Union of Biochemistry and
Molecular Biology (NC-IUBMB) in consultation
with the IUPAC-IUB Joint Commission on
Biochemical Nomenclature (JCBN).
• Enzyme nomenclature is a resource providing
general information on enzyme nomenclature.
• Six groups of enzyme:

Group Reaction Examples

Catalyzed
1. Oxidation-reduction Alcohol
Oxidoreductase reactions. dehydrogenase &
cytochrome oxidase
2. Transferase Transfer of a functional Transaminase &
group from a donor phosphorylases
molecule to an
acceptor molecule.
3. Hydrolases Hydrolytic reaction. Lipase, amylases,
Addition of water to, or peptidase.
removal of water from
substrates.
4. Isomerases Conversion of a Phosphoglucomutase
molecule from one isomerase.
isomeric form to
another – transfer
atoms from one part of
a molecule to another.
5. Ligases Reactions in which two Aminoacyl tRNA
molecules join in a synthetases
process coupled to the
hydrolysis of ATP.
6. Lyase Reactions in which Pyruvate
double bonds form or carboxylase & RuBP
break, by means other carboxylase
than hydrolysis.

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2.5.8 Enzyme Technology

Immobilized Enzyme

 Enzyme that is physically attached to a solid

support over which a substrate is passed and
converted to product.
• Advantages:
i. Multiple or repetitive use of a
single batch of enzymes.
ii. Ability to stop reaction rapidly by
removing enzyme from reaction solution (or
vice versa)
iii. Enzymes are usually stabilized by
bounding.
iv. Product is not contaminated with the
enzyme (especially useful in food and
pharmaceutical industries)

• Application: example, lactose hydrolysis

 Purpose of using immobilized enzymes is to
convert lactose via hydrolysis into glucose
and galactose.
 Lactose occurs naturally in both human and
cow's milk. Widely used in baking and
commercial infant-milk formulas.
 Problem: Many people are lactose intolerant
-their body is incapable of digesting lactose.
 So it must be hydrolyzed into its
monosaccharide components, allowing

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digestion, which is the purpose of products

today such as LACTAID.
Biosensor

 Device that uses specific biochemical reactions

mediated by isolated enzymes, immunosystems,
tissues, organelles, or whole cells to detect
chemical compounds, usually by electrical,
thermal or optical signals.
• It is an analytical device which converts a
biological response into an electrical signal
• The term 'biosensor' is often used to cover
sensor devices used in order to determine the
concentration of substances and other
parameters of biological interest even where
they do not utilise a biological system directly.
• A successful biosensor must possess at least
some of the following beneficial features:
i. The biocatalyst must be highly specific for the

purpose of the analyses, be stable under

normal storage conditions.
ii. Reaction should be as independent of such

physical parameters as stirring, pH and

temperature as is manageable. This would
allow the analysis of samples with minimal
pre-treatment.
iii. The response should be accurate, precise,

reproducible and linear over the useful

analytical range, without dilution or
concentration.

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iv. The complete biosensor should be cheap,

small, portable and capable of being used by
semi-skilled operators.

Schematic diagram showing the main components of a

biosensor

• The biocatalyst (a) converts the substrate to product.

• This reaction is determined by the transducer (b)
which converts it to an electrical signal.
• The output from the transducer is amplified (c),
processed (d) and displayed (e).

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