Nucleic Acids & Protein Synthesis

1. Nucleic Acids & the Nucleotide

Nucleic acids come in two types: DNA (deoxyribonucleic acid) and RNA (ribonucleic acid). Both are polymers — more precisely polynucleotides (poly = many; many nucleotides joined together).

Nucleotide = the monomer / basic unit of nucleic acids. DNA strand = polymer of nucleotides, RNA strand = polymer of nucleotides.

A nucleotide has 3 parts:

1. Pentose sugar (5-carbon sugar)
2. Nitrogenous base — adenine, thymine, cytosine, guanine, uracil
3. Phosphate group (at least one)

The 2 pentoses (learnt in biological molecules): deoxyribose and ribose. A DNA nucleotide and an RNA nucleotide differ in the sugar and in one base:

• Ring = O + 4 carbons (C1′–C4′). There is no room in the ring for the 5th carbon → C5′ sits outside the ring.
• Base on C1′ · phosphate on C5′ · OH on C3′ (used later to make the bond).
• de-oxy = “oxygen missing”: at C2′ only, OH becomes just H. The other OH groups keep their oxygen. (Two different pentoses — not the same sugar with an O removed from everywhere.)
• Only ONE base is attached to each nucleotide — any one of the allowed four.

	Nucleotide name	Sugar	Possible bases
in DNA	deoxyribonucleotide	deoxyribose	A · T · G · C
in RNA	ribonucleotide	ribose	A · U · G · C

Uracil replaces thymine in RNA ⇒ in total 5 bases: A, T, G, C, U.

Two ways to classify the bases

① by which nucleic acid they are in (above). ② by size = number of rings:

Exam style: if a drawn nucleotide shows a double-ringed base on C1′, it is a purine (single ring ⇒ pyrimidine). Purines are larger, pyrimidines smaller.

2. Joining Nucleotides — the Phosphodiester Bond

One nucleotide = one monomer. To get a dinucleotide and then a polynucleotide, nucleotides join by condensation (a small molecule — water — is released, just like in carbohydrates, proteins, lipids).

• Phosphodiester bond = covalent = strong (strong bonds = covalent & ionic → drawn as a solid line).
• Hydrogen bonds = weak → drawn as a dotted line. (Glue analogy: weak bond = temporary glue that peels off; strong bond = permanent adhesive.)
• Every nucleotide in the chain is held to the next by one phosphodiester bond. More than 2 units ⇒ polynucleotide (e.g. 6 nucleotides = 5 bonds).

A polynucleotide = one DNA (or RNA) strand

Why is a free phosphate seen only at ONE end? The phosphate on C5′ of every other nucleotide has lost its OH in condensation — it is busy in a phosphodiester bond. Likewise the 3′-OH of every nucleotide except the last is used up.

5′ end = the end with a free (unbonded) phosphate.

3′ end = the end with a free (unbonded) OH.

Backbone: the chain of sugar + phosphate held by phosphodiester bonds = the sugar–phosphate backbone (like our spine). We do not say “deoxyribose-phosphate” because it is the same idea for DNA & RNA.

Nitrogenous bases are NOT part of the backbone and do not join nucleotides together — they stick out and later hydrogen-bond with bases of another strand.

3. Structure of DNA

Classic statement: DNA is a double-helical structure. Break it down: double = made of 2 strands; helical = spiral (braided, like a twisted rope-ladder).

• ✱ double stranded → 2 sugar–phosphate backbones (each a polynucleotide) — held together by H-bonds between bases
• ✱ helical
• ✱ antiparallel

Antiparallel: the two strands are parallel (side by side, not crossing) but run in opposite directions — one is 5′→3′, the other 3′→5′.

Complementary base pairing (like enzyme & substrate being complementary): each base pairs with one particular base only:

Getting the other strand: write the complement of each base, then reverse the direction: 5′ AAGTCGCCTA 3′ ⇒ 3′ TTCAGCGGAT 5′ (read 5′→3′ it is TAGGCGACTT).

Chargaff's rule

Why? every A in one strand sits opposite a T in the other, every G opposite a C. So if one strand has 6 A, the partner strand has 6 T; 10 G ⇒ 10 C. Add up: the purines of the whole molecule = the pyrimidines.

Check (ma'am's random code above): purines = 3 A + 2 G = 5 in the top strand… and the pyrimidines also 5; same in the bottom strand. ✓ rule holds.

Stability of DNA — 3 reasons

1. Numerous hydrogen bonds between the complementary base pairs. Each H-bond is weak, but there are so many that together they act like glue holding the 2 strands.
2. Strong phosphodiester bonds (covalent) within the sugar–phosphate backbone.
3. Helical structure → provides protection: enzymes cannot easily attack the DNA molecule.

Why does the helix protect? Imagine DNA as a plain linear molecule: enzymes could land anywhere and cut it. Twisted into a compact spring-like helix, only the ends are open to enzyme attack → the helix shields it from damage.

4. RNA and DNA vs RNA

RNA is of 3 types:

• mRNA messenger RNA · tRNA transfer RNA · rRNA ribosomal RNA (found in ribosomes)
• Single stranded — ALL RNA is. (tRNA looks double-stranded, but it is one strand folded back on itself.)
• Pentose = ribose · bases A U G C (U replaces T)
• Shape: linear (mRNA, rRNA) or clover-leaf (tRNA) · non-helical
• Shorter than DNA · short-lived · made by transcription

	DNA	RNA
Sugar	deoxyribose	ribose
Bases	A T G C	A U G C
Strands	double stranded	single stranded
Shape	helical	non-helical (linear / clover leaf)
Size	the larger molecule	the smaller molecule
Lifespan	long-lived, stable	short-lived
Produced by	DNA replication (S phase)	transcription

5. DNA Replication

Replication = making an exact copy of DNA. It happens in the S phase of interphase (before mitosis — this is how the amount of DNA is doubled).

Requirements:

1. Parental DNA — each strand acts as a template
2. Enzymes (every reaction in the body is enzyme-catalysed): DNA helicase, DNA polymerase, DNA ligase
3. Energy in the form of ATP (the energy currency of the cell)
4. Activated nucleotides

Activated nucleotide — why & what

• A normal (non-activated) nucleotide has just 1 phosphate — it has no extra energy (like a lazy person); not good enough for the job.
• Any process needs energy. A nucleotide is activated by adding 2 more phosphate groups (phosphorylated) → 3 phosphates. These phosphates come from ATP (the bonds between phosphates are energy-rich: when a phosphate breaks off, energy is released).
• Why activated? The new strand is built from DNA monomers = nucleotides; we don't chop the old DNA up — we synthesise from fresh nucleotides.

Replication is SEMI-CONSERVATIVE

Conserve = preserve unchanged. Semi = half. So semi-conservative = half of the parent molecule is conserved: each new (daughter) DNA = 1 parental strand + 1 newly synthesised strand.

1. Helicase breaks the H-bonds → the 2 strands separate; each is now a template; bases are exposed (unbonded).
2. Activated nucleotides arrive and pair by complementary base pairing (temporary H-bonds): A–T, G–C.
3. Phosphodiester bonds form between neighbouring new nucleotides → new sugar–phosphate backbone.
4. Finally the H-bonds between the template and the new strand hold the new helix together.

The mechanism — role of each enzyme

Enzyme	Role
DNA helicase	unwinds & unzips the DNA by breaking H-bonds between the two strands (weak bonds → easy to break); makes 2 template strands
DNA polymerase	attaches (at the 3′ end of the template side) and joins complementary activated nucleotides → forms the new strand's sugar–phosphate backbone (phosphodiester bonds). Always 5′ → 3′.
DNA ligase	joins the Okazaki fragments on the lagging strand into one continuous strand

Leading vs lagging: the two parental strands are antiparallel but polymerase has only one direction.

• Leading strand = continuous replication: polymerase just follows the helicase as it unzips.
• Lagging strand = discontinuous replication: here 5′→3′ means working backwards (away from the fork). Polymerase waits until helicase has opened enough, then makes a short fragment, then another … → Okazaki fragments. (It cannot change its method, so it changes its direction.)
• If both new strands were made the same way they would both be 5′→3′ and the antiparallel principle would break.
• DNA ligase then joins all the fragments → one solid strand.
• Memory: “lag” = like a lagging screen / internet — a break in continuity.

Three models of replication

• Conservative: parental DNA stays whole (both old strands together), the other daughter is wholly new.
• Semi-conservative ✓ (accepted): each daughter = 1 old + 1 new strand.
• Dispersive: each strand of both daughters is fragmented — some parts old, some new.

Meselson–Stahl experiment — how semi-conservative was proved

• Two isotopes of nitrogen: N¹⁵ = heavy, N¹⁴ = light (both non-radioactive → harmless). Nitrogen is part of DNA (in the bases!), so growth medium decides which N goes into the DNA.
• Used E. coli — it divides (one generation) every 20 minutes.
• Grew E. coli in N¹⁵ medium → all its DNA is N¹⁵ = heavy DNA = generation 0.
• Moved the bacteria to N¹⁴ medium; sampled every 20 min and checked the DNA.

• After 20 min (generation 1): ALL DNA is hybrid — one N¹⁵ strand + one N¹⁴ strand → 50 % N¹⁵ : 50 % N¹⁴.
• After 40 min (generation 2): hybrid DNA and completely light DNA. Count strands: N¹⁴ : N¹⁵ = 3 : 1 (75 % : 25 %).
• Why does N¹⁴ rise? The only N in the medium is N¹⁴, so every new strand is N¹⁴; the old N¹⁵ strands are conserved but get diluted.

6. DNA & the Genetic Code

History: proteins were discovered before nucleic acids. Enzymes (proteins) control every life process, so scientists first assumed proteins = genetic information. Then Watson & Crick discovered DNA, and it became clear that DNA holds the genetic information; proteins carry out metabolism. So: how does DNA decide which protein to make?

Primary structure of a protein = the sequence of amino acids in a polypeptide (e.g. Met – Glu – Ala). Each protein has a unique amino-acid sequence ⇒ unique structure ⇒ unique function.

(Every body cell has the same DNA as the zygote it came from. Identical twins share the same genotype — but their gene expression/phenotype can still differ.)

What exactly is a gene?

• The short form is acceptable for marks, but a gene is NOT the whole DNA — DNA is made of many genes.
• One gene = one complete polypeptide (insulin gene ⇒ enough nucleotides for all of insulin's amino acids).
• Gene expression = protein synthesis = when the gene successfully converts into a protein. A gene may sit dormant for years (e.g. a hormone gene switched on only at a certain age) — it was always there, just not expressed.

Note: ma'am quoted “416 amino acids” for an insulin chain but wasn't sure — real human insulin has only 51 (A chain 21 + B chain 30). The idea is what matters.

Understanding the code — 5 features

There are 20 amino acids to be coded for, but DNA has only 4 bases. Which code length works?

bases read at a time	possible codes	enough for 20 amino acids?
1 (singlet)	4	✗ only 4 amino acids
2 (duplet)	4² = 4 × 4 = 16	✗ “cannot code for 20 amino acids”
3 (triplet)	4³ = 64 codons	✓ 64 ≥ 20

(Ma'am's lists: 1 base: A, C, G, T · duplets: AA, AC, AG, AT, CA, CC, CG, CT … → 16 · triplets: AAA, ACC, AGG, ATT, ACA, AGA, ATA … → 64. Letters X, Y, Z… stood for amino acids.)

① Genetic code is a TRIPLET code

The order of triplets decides the order of amino acids in the chain (triplets X, Y, A, Z ⇒ chain X – Y – A – Z). A gene of 45 bases = 15 triplets = 15 codons.

② Genetic code is DEGENERATE (redundant)

64 combinations is more than needed for 20 amino acids ⇒ 44 extra triplets ⇒ each amino acid is coded by more than one triplet.

More than one code for the same amino acid is fine; a codon for a different amino acid, or a nonsense code, would not “open the account”.

③ Genetic code is PUNCTUATED

A sentence with no full stop never ends! The ribosome needs to know where to start and stop: stop codons don't code for any amino acid — they are the “full stop”.

STOP codons (mRNA)	mnemonic
UAA	“U are away”
UAG	“U are gone”
UGA	“U go away”

④ Genetic code is NON-OVERLAPPING

Adjacent bases / codons do not overlap: each base belongs to only one triplet; each triplet makes its own amino acid without interfering with its neighbour.

⑤ Genetic code is UNIVERSAL

All living organisms use similar triplet codons for the same amino acids: AUG codes for methionine in humans, rats, monkeys…. Say “similar”, not “the same” (it is true for most organisms).

7. Structure of ATP

ATP = adenosine triphosphate (A from adenosine, T = “tri”, P = phosphate).

• Adenosine = ribose (pentose) + adenine (base). Triphosphate = 3 phosphate groups (mono = 1, di = 2, tri = 3).
• So ATP = a modified nucleotide = activated nucleotide = phosphorylated nucleotide (phosphorylation = adding phosphate groups): a ribonucleotide (A) with 2 extra P groups.
• AMP (adenosine mono-phosphate, 1 P) = just a simple nucleotide; ADP = 2 P; ATP = 3 P.
• The bonds between the phosphate groups are energy-rich (energy is stored there; released when a phosphate breaks off).

8. Protein Synthesis — Overview

Protein synthesis is an active process = energy-consuming → needs ATP (ATP is the cell's energy currency — like only Pakistani rupees work in Pakistan, only ATP energy works in the cell).

Two stages (each followed by a modification):

• (i) TRANSCRIPTION — in the nucleus: synthesis of mRNA using a gene; then post-transcriptional modification (RNA processing).
• (ii) TRANSLATION — at the ribosomes: synthesis of a polypeptide using mRNA; then post-translational modification (= protein folding).

Why do we need mRNA?

• All genetic information is in the DNA, which cannot leave the nucleus; but proteins cannot be made inside the nucleus (too little space) → the information must be carried out to the cytoplasm.
• mRNA = the translator / messenger between the language of DNA and the language of protein (like an interpreter between an Urdu speaker and an English speaker).
• Only the gene needed at that moment is unzipped and expressed. If the whole DNA were transcribed, hundreds of unneeded polypeptides would be made together → protein levels rise → harmful, the cell could burst.

9. Transcription

• Site: nucleus · result: synthesis of mRNA using a gene.
• Why only one DNA strand? RNA is single stranded, so one template is enough.
• Template = a pre-existing design (like a Canva template): the strand that gets copied = template strand = transcribed strand.

Requirements:

1. Activated ribonucleotides — ribo, not deoxyribo (deoxyribonucleotides would build DNA, we want RNA!); activated by ATP phosphates.
2. Template strand of the gene (of the DNA) — only the gene needed, not the whole DNA.
3. RNA polymerase — joins the monomers (poly-mer-ase!) = the ribonucleotides complementary to the template.
4. Energy in the form of ATP

Gene structure & the 3 stages

• Promoter = start point of a gene; the site to which RNA polymerase binds for transcription.
• Terminator (transcription terminator sequence) = marks the end of transcription.
• Exons = coding part (carry a message). Introns = non-coding (no code, no message — like the appendix: present, but no role).

• Before the phosphodiester backbone is made, the mRNA is held to the template by H-bonds. These are temporary: when mRNA is complete they break — mRNA must leave the nucleus and attach to a ribosome.
• The product is the primary transcript (immature mRNA): a complementary copy of the whole gene including intron sequences — not yet able to bind a ribosome.

Which strand was transcribed? The mRNA is complementary to the template: AUG… ⇒ template TAC… (the “TAC strand”). The ATG-strand is the non-transcribed one. AUG = start, UAG = one of the 3 stop codons.

10. Post-transcriptional Modification (RNA processing)

Site: nucleus. The primary transcript is modified so that it can survive in the cytoplasm and attach to a ribosome after leaving the nucleus. Also called RNA processing. 3 steps:

(1) Splicing

• Removal of introns (non-coding) and joining of exons (coding).
• Enzyme: spliceosome — a “genetic scissor”.
• Non-coding ≠ no bases. Introns are copied into the primary transcript; their base sequence just doesn't code for any amino acid.

(2) 5′ capping   (3) Polyadenylation at 3′

• 5′ capping = addition of a modified guanine at the 5′ end (like putting a cap on a toothpaste tube — protects the contents).
• Polyadenylation at 3′ = poly (many) + adenylation (adenine nucleotides added): about 150–200 adenine nucleotides at the 3′ end (the poly-A tail).
• Cap + tail protect both ends from intracellular enzymes that would attack the mRNA.

11. tRNA and Ribosomes

tRNA — the amino-acid carrier

• Single stranded · bases A U G C (common to all RNA).
• Clover-leaf shaped — looks double-stranded but is ONE strand folded (5′ end and 3′ end are both on top).
• Intramolecular complementary base pairing with H-bonds between the bases — within the same molecule (A–U, G–C), which holds the fold.
• Anticodon: each tRNA has a specific anticodon, and is specific for carrying one particular amino acid. The anticodon determines which amino acid attaches to the tRNA = its “badge / ID card”. One tRNA carries one amino acid (alanine-tRNA → alanine, glycine-tRNA → glycine …).
• The amino acid attaches at the 3′ end (opposite the anticodon). The 3′ end always ends with CCA (read 5′→3′; drawn top-down as A-C-C).

Board slip: the notes write the anticodon of UCA as “A G A” — the complement is A G U.

Drawing tRNA quickly: draw a cross; the long line is the 3′ end and the anticodon sits opposite to it.

tRNA + its amino acid = charged tRNA = aminoacyl-tRNA complex. The attachment is reversible (an enzyme is involved — details not needed): what attaches can detach.

Ribosome

• Two sub-units: a large and a small sub-unit, made of protein + rRNA (ribosomal RNA).
• The large sub-unit contains the enzyme peptidyl transferase (makes peptide bonds).
• The two sub-units assemble (ribosome assembly — possibly helped by Mg²⁺ ions) when needed and disassemble afterwards.

12. Translation

Requirements: ① ATP ② ribosomes (the site) ③ mRNA (the molecule being translated; it left the nucleus through a nuclear pore) ④ amino acids ⑤ tRNA (carrying specific amino acids).

Divided (like transcription) into initiation → elongation → termination. The mRNA of the example has start AUG, 4 triplets in between, and a stop codon (UAG): AUG GCG GGA CUC UGC UAG.

Post-translational modification = protein folding

Takes place in the rough ER: the polypeptide enters the RER sacs. Primary structure (from translation) → secondary (α-helices, β-pleated sheets) → tertiary (R-group interactions: H-bonds, hydrophobic interactions, ionic bonds, disulfide links) → quaternary (when > 1 polypeptide chain interact).

	Transcription	Translation
Where	nucleus	ribosomes (free / RER)
Makes	mRNA (using a gene)	polypeptide (using mRNA)
Enzyme	RNA polymerase	peptidyl transferase
Bond made	phosphodiester	peptide
Start / end	promoter / terminator	AUG / stop codon
Then	RNA processing	protein folding (RER)

13. Gene Mutation

Mutation = a change in the genetic information (DNA = our hereditary information bank). Two kinds:

3 types of gene mutation: base substitution · base addition (insertion) · base deletion.

Base substitution — a base is replaced by a different base

Type	Definition	Why (analogy)
Silent	no overall change in the polypeptide chain despite gene mutation / base substitution	degenerate code: new codon codes the same amino acid
Missense	a mutation that changes an amino acid in a polypeptide chain	miscommunication; complete chain, same length, but wrong amino acid
Nonsense	a mutation that changes a codon to a stop codon, so protein synthesis (the polypeptide chain) ends abruptly	makes no sense; chain incomplete

Base addition (insertion) & base deletion

Addition / removal of one or more nucleotides. This may lead to a FRAMESHIFT mutation. (Think photo frame: add or remove something and the frame shifts.)

Board example (codons): UAC UGG CUA GUA CAU = Tyr–Trp–Leu–Val–His. Insert one C → UAC UCG GCU AGU ACA U = Tyr–Ser–Ala–Ser–Thr: one C, and everything after it is read wrongly.

Board slip: its first line is printed “TAT TGG…” — it should be TAC TGG… for the inserted C to give the second line.

Not every insertion/deletion shifts the frame: adding or removing a multiple of 3 nucleotides (a whole triplet) keeps the frame — the code is a triplet code. Leftover 1 or 2 bases = the problem.

14. Sickle-Cell Anaemia

• An inherited (genetic) disorder caused by base substitution — a classic missense mutation.
• Homozygous recessive: the child inherits the recessive allele from both parents.
• Affects the RBCs — which are mostly haemoglobin (Hb). Hb = 4 polypeptide chains: 2 α-globin + 2 β-globin. The disorder is due to the β-globin chain (made by one gene).

Genotype	Allele	Result
HbA HbA	HbA = normal / dominant	normal
HbA HbS	HbS = abnormal / recessive	carrier (sickle-cell trait) — no disease
HbS HbS	both abnormal	sickle-cell DISEASE

What goes wrong?

• Normally the 6th triplet of the β-globin gene is CTC → glutamic acid (hydrophilic = soluble). After substitution: CAC → valine (hydrophobic = insoluble). T is replaced by A.
• The β-globin chain, so haemoglobin, becomes less soluble → precipitates (small lumps — like lumps in a flour/besan mix) inside the RBC. Lumps polymerise into long fibres which pull the RBC wall inward → sickle (crescent) shape, smaller capacity.
• RBCs have no nucleus/mitochondria so as to carry the max. of Hb and O₂ — so any loss of capacity matters a lot.

Symptoms & treatment

reduced O₂-carrying capacity of the blood ⇒

1. Fatigue — less O₂ ⇒ less aerobic respiration ⇒ less ATP ⇒ tires quickly
2. Swelling / pain in hands & feet
3. Reduced number of RBCs (anaemia — RBCs are smaller, wear out faster)
4. Frequent infections
5. Stroke / heart attack — rigid sickle cells block blood vessels (obstruction → blood cannot reach tissues)

15. Full Chapter Cheat-Sheet