Biological Molecules

DNA Extraction (Practical)

Steps:

1. Grind sample in mortar & pestle → breaks cell walls.

2. Mix with detergent → breaks membranes, releases cell contents.

3. Add salt → breaks hydrogen bonds between DNA and water.

4. Add protease enzyme → digests proteins associated with DNA.

5. Add alcohol (ethanol/ice cold) → precipitates DNA.

6. DNA appears as white strands → can be spooled onto glass rod.

Collagen

Collagen (tendons, ligaments, skin, bones, cartilage):

Each molecule is three polypeptide chains wound into a triple helix.

Glycine every third residue (small side chain) allows tight packing

Hydrogen bonds stabilize the triple helix.

Molecules align staggered and cross-link to form fibrils, which bundle into fibres → very high tensile strength.

Elastin

Elastin (skin, lungs, blood vessel walls):

Made by linking many tropoelastin molecules.

Tropoelastin has alternating hydrophobic regions and lysine-rich areas

Molecules are cross-linked by covalent bonds between lysines, creating an elastic network that stretches and recoils without breaking → elasticity.

Keratin

Keratin (hair, skin, nails): rich in cysteine → many disulfide (S–S) bonds.

More S–S bonds ⇒ harder

Less flexible (nails)

Fewer ⇒ more flexible (hair).

Burning hair/skin smells due to sulfur.

Fibrous Protein

Built from a limited range of amino acids with many hydrophobic R-groups in the primary sequence → repetitive, highly ordered structures

insoluble

mechanical strength rather than metabolic roles.

Insulin

Insulin: globular hormone for blood glucose regulation.

Must be soluble for transport in plasma and have a precise shape to fit receptors on cell membranes.

Conjugated Protein: Catalase

Contain a non-protein prosthetic group (may be lipid → lipoprotein, carbohydrate → glycoprotein, metal ions or vitamin-derived groups).

Catalase: enzyme that decomposes H₂O₂ (harmful metabolic by-product). Quaternary protein with haem groups (Fe) that let it interact with hydrogen peroxide and rapidly break it down so it doesn’t damage cells.

Conjugated Protein: Haemoglobin

Contain a non-protein prosthetic group (may be lipid → lipoprotein, carbohydrate → glycoprotein, metal ions or vitamin-derived groups).

Red oxygen-carrier in RBCs. Quaternary protein of 4 polypeptide subunits (2α + 2β), each with a haem prosthetic group containing Fe²⁺. Iron binds O₂ reversibly, enabling pick-up in lungs and release in tissues.

Globular Proteins

Form when the chain folds so hydrophobic R-groups tuck inside and hydrophilic R-groups face out → soluble in aqueous environments (blood, cytosol).

Vital in regulation and rapid processes: enzymes, hormones, antibodies, transport.

Tertiary Structure

3D folding due to R-group interactions:

• Hydrophobic/hydrophilic interactions.

• Hydrogen bonds( formed by oppositely charge groups).

• Ionic bonds.

• Disulfide bonds (covalent, between sulfur-containing R groups).

  • Cysteine has sulfur. When two cysteines are close together, a covalent bond can form.

Produces complex 3D shape → functional protein.

Order of strength:

• Disulfide bonds (covalent) – strongest, permanent cross-links between cysteine residues.

• Ionic bonds (salt bridges) – strong electrostatic interactions between oppositely charged R groups.

• Hydrogen bonds – weaker than ionic but very numerous, important for shape and stability.

• Hydrophobic interactions / van der Waals forces – weakest individually, but collectively contribute a lot.

Primary & Secondary Structure

1. Primary Structure

Sequence of amino acids in polypeptide chain.

Determines final shape and function.

2. Secondary Structure

Forms immediately after being formed in ribosome.

No R Group interactions are present.

Regular structure formed by H-bonds between amino acids (amine & carbonyl groups) causing folding or coiling:

Note: Carbonyl is C=O group

• α-helix (coil).

• β-pleated sheet (zig-zag sheet).

Thin Layer Chromatography (TLC) – Separating Amino Acids

- Technique for separating/identifying amino acids.

- Components:

- Stationary phase = thin layer of silica gel on sheet.

- Mobile phase = solvent.

- Method:

1. Draw pencil line 2 cm from bottom of plate; mark spots.

2. Add amino acid solution.

3. Place plate in solvent (no more than 1 cm deep).

4. Solvent moves up plate; amino acids separate by solubility + interactions.

5. Remove plate when solvent front ~2 cm from top.

6. Spray with ninhydrin → amino acids visible as purple/brown spots.

7. Measure distance travelled.

- Retention factor (Rf):Rf=distance travelled by solvent/distance travelled by component

- Each amino acid has characteristic Rf value (under identical conditions).

Emulsion Test

1. Mix sample with ethanol.

2. White emulsion = lipid present.

3. Clear = negative.

4. Mix with water + shake.

5. Should form two layers of liquid.

Cholesterol

Manufactured in liver & intestines.

Role:

• Positioned between phospholipids.

• Adds stability to membranes

  • cholesterol has hydrophobic and hydrophilic ends that interact with phospholipids (1), acting to make cell membranes more stable/rigid (1)

• Prevents membranes becoming too fluid at high temps or too rigid at low temps.

• Used to make: Vitamin D, steroid hormones, bile.

Sterols

• Known as steroid alcohols.

• Not fats/oils, but complex alcohol-based molecules.

• Structure: 4 carbon rings + polar –OH group at one end + hydrophobic rest of molecule

Phospholipid behaviour in Water

Form surface films/monolayers (heads in water, tails out).

Form bilayers (hydrophobic tails inside, heads outside).

→ Essential for membranes.

Called surfactants (reduce surface tension) e.g. in lungs

Phospholipids

- Modified triglycerides: contain P, C, H, O.

- One fatty acid replaced by phosphate group (PO₄³⁻).

- Phosphate group = negatively charged → soluble in water.

Structure:

- Hydrophilic head (phosphate group).

- Hydrophobic tails (fatty acid chains).

- → Molecule is amphipathic (dual nature).

Triglycerides

- Made by combining 1 glycerol with 3 fatty acids.

- Glycerol: part of alcohols group (contains –OH).

- Fatty acids: carboxylic acids (–COOH + hydrocarbon chain).

- Reaction: condensation/esterification → forms 3 ester bonds (type of covalent bond) + 3 water molecules.

Functions of Lipids

• Protection of vital organs

• To prevent evaporation or for waterproofing in plants & animals

• To insulate the body

• They form the myelin sheath around some neurones

• As a water source (water is released as lipids are respired)

• As a component of cell membranes

• Hormone production

• Buoyancy for aquatic animals

Benedict's for Reducing & Non Reducing

Reducing Sugars:

Procedure:

1. Place sample in boiling tube (if solid, grind or blend with water).

2. Add equal volume of Benedict’s reagent (alkaline copper(II) sulfate).

3. Heat gently in a boiling water bath for 5 minutes.

Reaction:

- Reducing sugar donates electrons to Cu²⁺ (blue) → reduced to Cu⁺ (brick-red Cu₂O precipitate).

Non-reducing sugars (e.g. sucrose) do not react with Benedict’s and remain blue (negative result).

To test them:

1. Boil sample with dilute HCl (hydrolyses sucrose → glucose + fructose, both reducing).

2. Neutralise with alkali.

3. Repeat Benedict’s test → positive result if non-reducing sugar was present.

Colorimetry

Benedict’s test colour intensity depends on sugar concentration. A colorimeter measures absorbance or transmission of light by the solution. More concentrated solution = more light absorbed, less transmitted.

Procedure:

1. Filter sample.

2. Calibrate colorimeter with distilled water.

3. Perform Benedict’s test on range of glucose concentrations.

4. Filter solutions to remove precipitate.

5. Measure % transmission of each solution with colorimeter.

6. Plot calibration curve of % transmission vs concentration.

7. Use curve to find concentration of unknown solution.

Starch & Glycogen Location

Starch- grains in storage organs (tubers or seeds or in chloroplast)

Glycogen- granules stored in the cytoplasm of liver & muscle cells.

Cellulose Fibres

Cellulose chains form hydrogen bonds with each other → microfibrils.

Microfibrils bundle into macrofibrils, which form fibres

Fibres = strong, insoluble, used to make cell walls.

Cellulose = dietary fibre/roughage:

- Hard to digest (humans lack cellulase enzyme).

- Provides fibre essential for healthy digestive system.

- High tensile strength

Cellulose

• Made from β-glucose (not α-glucose).

• β-glucose molecules cannot join unless alternate molecules are rotated 180°.

• This allows 1–4 glycosidic bonds to form.

• Results in a straight, unbranched chain

Key shared properties of amylopectin & glycogen:

• Insoluble.

• Branched.

• Compact.

• Adapted to efficient storage and rapid glucose release.

Glycogen

• Main energy storage polysaccharide in animals and fungi (functionally equivalent to starch in plants).

• Highly branched due to more alpha 1-6 bonds than amylopectin.

• Compact, requires less storage space, more insoluble structure.

• Branching = many free ends → glucose can be rapidly added/removed.

• Important for mobility in animals, ensuring a constant energy supply.

Amylopectin

• 1–4 glycosidic bonds between α-glucose molecules.

• –6 glycosidic bonds, creating branching points.

• Branches occur ~ every 25 glucose subunits.

• Branched structure = more compact & less soluble than glucose

• Efficient storage.

Amylose

• Long chain of α-glucose monomers

• 1–4 glycosidic bonds.

• Helix, (hydrogen bonding)

• Helical structure -> compact & less soluble than glucose.

• Efficient for storage

Monosaccharides of the following:

1. Maltose

2. Sucrose

3. Lactose

1. Glucose & Glucose

2. Fructose & Glucose

3. Galactose & Glucose

Water Properties Essential for Life

1. Solvent:

- Polar molecules (e.g., amino acids, nucleic acids, salts) dissolve in water.

- Important for cytosol of prokaryotes and eukaryotes, and for transport of ions/molecules.

- Salt= ionic= polar = can interact with water (dissolve)= hydrophilic (water loving)

- BUT Oil= non polar= does not dissolve= hydrophobic (or polysaccharides)

2. Transport medium:

- Cohesion allows water to move as one mass.

- Capillary action = adhesion + cohesion → enables water to rise in narrow tubes (important in plants).

3. Temperature stabiliser:

- Acts as a coolant.

- Absorbs large amounts of energy to break hydrogen bonds → buffers organisms from rapid temperature changes.

- Helps maintain stable enzyme function within the narrow temperature range required.

4. Habitat:

- Aquatic organisms depend on stable water environments.

- Ice floats → insulates water beneath, preventing freezing solid.

- Surface tension allows some organisms to live on water (e.g., pond skaters).

Characteristics of Water

Unusually high boiling point for a small molecule:

- Caused by hydrogen bonding, which requires lots of energy to break.

- Explains why water is liquid at room temperature, unlike CO₂ or O₂.

Freezing behaviour:

- Water expands when frozen.

- At 4°C and below, hydrogen bonds fix molecules in a rigid, open, tetrahedral lattice → ice less dense than liquid water → floats.

Cohesion & Adhesion:

- Cohesion: molecules stick to each other → enables water transport in plants (e.g., drawn up xylem).

- Adhesion: molecules attracted to other surfaces → e.g., water wets skin but doesn’t run off.

Surface tension:

- Cohesion creates a ‘skin’ at the surface.

- Strong enough to support small organisms (e.g., pond skaters).