7.

1 DNA structure

Hershey and Chase

Structure of DNA

From the scattering pattern produced by a DNA molecule, discoveries could be made about it’s structure

● Composition:​ ​DNA is a double stranded molecule

● Orientation:​ ​Nitrogenous bases are closely packed together​ on the inside and phosphates form an outer
backbone
● Shape:​ The ​DNA molecule twists at regular intervals​ (every 34 Angstrom) to ​form a helix (two strands =
double helix)
DNA replication
→ a semi-conservative process that is carried out by a complex system of enzymes

DNA Gyrase

→ ​Relieves strain created by the unwinding of DNA by helicase​ and ​prevents supercoiling of the separated
strands
- Does this by relaxing positive supercoils (via negative supercoiling) that would otherwise form during the
unwinding of DNA
SSB proteins (single stranded binding

→ Bind to the DNA strands after they have been separated and prevent the strands from re-annealing
→ Help to prevent the single stranded DNA from being digested by nucleases
Nuclease - enzyme that cuts damaged DNA in two places to then be repaired by

★ SSB proteins will be dislodged from the strand when a new complementary strand is synthesised by DNA
polymerase III

DNA primase

→ ​Generates a short RNA sequence (primer) to initiate DNA synthesis

DNA polymerase III

● DNA pol III attaches to the 3’-end of the primer​ and covalently​ joins the free nucleotides​ together in a
5’ → 3’ direction
● As DNA strands are antiparallel, DNA pol III moves in opposite directions on the two strands
○ On the ​leading strand,​ DNA pol III is moving towards the replication fork and can synthesise
continuously
○ On the ​lagging strand,​ DNA pol III is moving away from the replication fork and synthesises in
pieces (Okazaki fragments)

DNA polymerase I

● As the ​lagging strand is synthesised in a series of short fragments​, it has ​multiple RNA primers along its
length​ (thing that attach to sides of dna nucleotides)
● DNA pol I removes the RNA primers​ from the lagging strand and ​replaces them with DNA nucleotides

DNA ligase

→ Joins the Okazaki fragments together to form a continuous strand with phosphodiester bond

DNA sequencing
→ The process of determining the sequence of nucleotides

Dideoxynucleotides
- They lack the 3’-hydroxyl group necessary for forming a phosphodiester bond
- prevent further elongation of a nucleotide chain and ends replication
Nucleosomes
→ ​In eukaryotic organisms​, the ​DNA is packaged with histone proteins to create a compacted structure called a
nucleosome

● Nucleosomes help to supercoil the DNA, meaning it will make a ​very compacted structure​ that allows for
more efficient storage
● Supercoiling helps to protect the DNA from damage​ and also a​llows chromosomes to be mobile during
mitosis and meiosis

7.2 Transcription (see 2.7)

Sections of a gene

Promoter

→ The non-coding sequence responsible for the initiation of transcription

● After RNA polymerase has bound to the promoter, it causes the DNA strands to unwind and separate

Coding Sequence

● The region of DNA that is transcribed by RNA polymerase is called the coding sequence

Terminator

● RNA polymerase will continue to

transcribe the DNA until it reaches a
terminator sequence
Antisense vs Sense
→ ​A gene (DNA)​ consists of ​two polynucleotide strands​, but ​only one​ is t​ranscribed into RNA

● The ​antisense strand​ is ​the strand that ​is​ transcribed into RNA
○ Also referred as the template strand
● The ​sense strand​ is the strand that is ​not​ transcribed into RNA
○ Also referred as the coding strand because it is a DNA copy of the RNA sequence

mRNA

Capping: ​the addition of a methyl group to the 5’-end of the transcribed RNA

Polyadenylation: ​the ​addition of a long chain of adenine nucleotides​ (a ​poly-A tail)​ to the 3’-end of the transcript
→​ Poly-A tail​ ​improves the stability of the RNA transcript​ and ​facilitates its export from the nucleus

Splicing: ​The process by which introns are removed

Epigenetics
Eukaryotic DNA is wrapped around histone proteins to form compact nucleosomes

Histones:
Histone tails have a positive charge​ and hence ​associate tightly with​ the ​negatively charged DNA

★ Adding an ​acetyl group​ to the tail​ (​acetylation)​ ​neutralises the charge​, making ​DNA less tightly coiled
and ​increasing transcription

★ Adding a ​methyl group​ to the tail​ (​methylation)​ ​maintains the positive charge​, making ​DNA more coiled
and ​reducing transcription
7.3 Translation

Ribosomes and tRNA

Ribosomes
● The ​small​ subunit contains an ​mRNA​ binding site
● The ​large​ subunit contains three ​tRNA​ binding sites

tRNA

➔ The ​acceptor stem​ carries an amino acid

➔ The ​anticodon​ associates with the mRNA codon (via complementary
base pairing)
➔ The ​T arm​ associates with the ribosome
➔ The ​D arm​ associates with the tRNA activating enzyme

tRNA activation

→ Each tRNA molecule binds with a specific amino acid in the cytoplasm in a reaction catalysed by a
tRNA-activating enzyme
➢ Each amino acid is recognised by a ​specific​ enzyme
Translation

1. Initiation
a. MR CAT APP

2. Elongation
a. A second tRNA molecule pairs with the next codon in the ribosomal ​A​ site
b. Amino acid in the P site is covalently attached via a peptide bond (condensation reaction) to the
amino acid in the A site
c. The ​tRNA​ in the P site is now ​deacylated​ (​no amino acid​), while the tRNA in the A site carries the
peptide chain

3. Translocation
a. The ribosome moves along the mRNA strand by one codon position ( 5’ → 3’ direction)
b. Deacylated tRNA moves in the E site and is released,
i. The tRNA carrying the peptide chain moves to the P site
c. Another tRNA molecules attaches to the next codon in the now unoccupied A site and the process
is repeated

4. Termination
a. Elongation and translocation continue in a repeating cycle until the ribosome reaches a stop codon
b. The polypeptide is released and the ribosome disassembles back into its two independent subunits
Prokaryotic translation

→ Prokaryotes lack a nucleus therefore transcription and translation don’t need to be separated

● Ribosomes may begin translating the mRNA molecule while it is still being transcribed from the DNA
template

Polysome / Polyribosome

→ Group of 2 or more ribosomes translating an mRNA strand simultaneously

Protein destination

● If protein is targeted for​ intracellular use​ within the cytosol → the ribosome remains ​free​ and unattached
● If protein is targeted for ​secretion,​ m
​ embrane fixation​ or use in ​lysosomes​ → the ribosome becomes
bound​ to the ER (endoplasmic reticulum)