DNA structure & synthesis

Question 1

Describe the main structural features of the DNA helix

Answer 1

• Double helix structure made up of two long strands of nucleotides twisted around each other
• Nucleotide: phosphate group, a deoxyribose sugar, and a nitrogenous base, which can be AT (2H bonds) GC (3H bonds)
• Antiparallel orientation, with one running from the 5′ to 3′ direction and the other from 3′ to 5′.
• The sides of the DNA “ladder” form a sugar-phosphate backbone, which provides stability and structure.
• The twisting of the helix creates major and minor grooves that allow proteins to bind to specific DNA sequences, playing a role in gene regulation.*

Helix measures about 2 nanometers in diameter, and one complete turn of the helix spans 3.4 nanometers, containing approximately 10 base pairs.

Question 2

Explain how the sequence of bases in DNA carries the genetic blueprint of life?

Answer 2

• Every three bases, called a codon, correspond to a specific amino acid, the building blocks of proteins.
• The sequence of codons in a gene determines the exact order in which amino acids are joined together to form a particular protein.
• Since proteins carry out most of the body’s functions—such as catalysing reactions, forming structures, and regulating processes—the information encoded in DNA determines an organism’s traits and how its cells function.

Question 3

Describe what is meant by ‘higher order structure of DNA’, how it is maintained in bacteria through gyrase, and through chromatin formation in eukaryotes?

Answer 3

• The higher-order structure of DNA refers to the way DNA is further folded, coiled, and compacted beyond the simple double helix so that it can fit inside a cell while still remaining accessible for processes like replication and transcription.
• In bacteria (prokaryotes), the DNA is typically circular and located in the nucleoid.
• To compact their DNA, bacteria use enzymes such as DNA gyrase, a type of topoisomerase.
• DNA gyrase introduces negative supercoils into the DNA molecule, which helps to relieve tension caused by unwinding during replication and transcription, and allows the DNA to coil tightly into a smaller, more manageable form.
• This supercoiling is essential for maintaining the compact, organised structure of bacterial DNA
• In eukaryotes, the higher-order structure is maintained through chromatin formation.
• The DNA wraps around groups of proteins called histones, forming bead-like units known as nucleosomes.
• These nucleosomes coil and fold further to form chromatin fibres, which can then condense even more tightly to form chromosomes during cell division.
• This hierarchical packaging not only allows the large eukaryotic genome to fit inside the nucleus but also plays a key role in regulating gene expression, as tightly packed DNA is generally less accessible for transcription.

Question 4

Describe how chemicals and other environmental insults can damage DNA and how cells respond through their DNA repair systems

Answer 4

• Spontaneous Damage: DNA can undergo chemical changes without any external cause.
  E.g. include: depurination – spontaneous loss of a purine base (adenine or guanine) from DNA
  deamination – hydrolysis of cytosine to uracil, causing C→T transition mutations if not repaired.
• Chemical and Metabolic Damage
• Reactive oxygen species (ROS) generated during normal metabolism can oxidise DNA bases, alter their structure, or cause strand breaks.
• Chemicals and drugs can also modify DNA: Alkylating agents (e.g., cyclophosphamide, an anticancer drug) add alkyl groups to DNA bases, causing mispairing or strand breaks.
• Intercalating agents (e.g., doxorubicin) insert between base pairs, distorting the DNA helix and corrupting DNA replication and transcription.
• These drugs exploit DNA damage to kill rapidly dividing cancer cells.
• Radiation —- Ultraviolet (UV) light causes the formation of thymine dimers (not fibres), where two adjacent thymine bases become covalently bonded. This distorts the DNA helix and blocks replication and transcription. Ionising radiation such as X-rays and gamma rays produces free radicals that can cause single- and double-strand breaks in DNA. These chromosome breaks can lead to mutations, chromosomal rearrangements, and diseases such as leukaemia

Question 5

Explain how DNA synthesis starts at replication origins and is co-ordinated with the cell cycle?

Answer 5

• DNA replication begins at specific sites called replication origins, which are recognised by an initiation complex.
• The DNA at these origins unwinds to form replication bubbles, allowing the replication machinery to access and copy the strands.
• This process occurs only once per cell cycle, during the S phase, when the parental DNA is fully unwound and duplicated to ensure accurate genome replication

Question 6

Describe how the properties of DNA polymerase ensure DNA synthesis occurs with high fidelity (accurate copying of DNA, with very few mistakes).

Answer 6

• Cells contain several DNA polymerases, which are enzymes responsible for synthesising new DNA strands.
• DNA polymerase works in the 5′ to 3′ direction, adding nucleotides to the 3′-OH end of a growing strand using A–T and C–G base pairing for accurate copying of the template strand.
• It requires a DNA template, a primer (DNA or RNA), the four dNTP building blocks, and Mg²⁺ ions for activity.
• Because DNA polymerase can only extend in the 5′→3′ direction, one strand (the leading strand) is made continuously, while the other (the lagging strand) is made in short pieces called Okazaki fragments.
• High fidelity in DNA replication is achieved through complementary base pairing, the proofreading (3′→5′ exonuclease) activity of DNA polymerase that removes incorrectly paired nucleotides, and the mismatch repair system that corrects any remaining errors after replication

Question 7

Base pairing in DNA, proof-reading by DNA polymerase, and mismatch repair system ensure DNA replication proceeds with high fidelity:

Human cells must replicate 3 x 10º basepairs of DNA

Error rate in DNA replication is ~1 in 101°. Why so low?

Answer 7

• DNA polymerase- error rate ~ 1 in 108- low due to base pairing and proof-reading/editing function of the enzyme
• Mismatch repair system- corrects most of the polymerase errors.
• Multi-enzyme system highly conserved across species

Question 8

Classify the various enzymes required for DNA replication.

Answer 8

• Imitation enzymes: Helicase – Unwinds the DNA double helix at the replication fork
• Single-Stranded Binding Proteins (SSBs) – Stabilise the unwound DNA strands and prevent them from rejoining
• Primase – Synthesises short RNA primers to provide a starting point for DNA polymerase.
• Elongation enzymes: DNA Polymerase – Adds nucleotides to the growing DNA strand in the 5′→3′ direction using the template strand
• Sliding Clamp (Clamp Protein) – Holds DNA polymerase onto the DNA, increasing its efficiency and Clamp Loader – Helps attach the sliding clamp to DNA.
• Strand separation and tension relief enzyme: Topoisomerase (DNA gyrase in bacteria) – Relieves the torsional strain (supercoiling) created by unwinding the DNA.
• Accessory proteins: Origin Recognition Complex (ORC) – Identifies replication origins and helps initiate replication.

Question 9

Describe how failure to correct DNA synthesis errors leads to cancer.

Answer 9

• During DNA replication, errors can sometimes occur when the wrong nucleotide is added.
• Normally, these mistakes are corrected by the proofreading activity of DNA polymerase and by the mismatch repair system after replication.
• If these correction mechanisms fail, the mutations (permanent changes in DNA sequence) remain in the genome.
• When such mutations occur in important genes that control cell growth and division—like proto-oncogenes, tumour suppressor genes, or DNA repair genes—they can disrupt normal cell cycle control.
• This may cause cells to divide uncontrollably, avoid normal death signals, and accumulate further genetic damage.
• Over time, these changes can lead to the formation of a tumour and eventually result in cancer.

Question 10

Recognise that DNA synthesis is an important target for chemotherapy

Answer 10

• DNA synthesis is a key process required for cell division, so rapidly dividing cells—like cancer cells—depend heavily on it.
• Because of this, DNA synthesis is an important target for chemotherapy.
• Many anticancer drugs work by inhibiting enzymes or blocking pathways involved in DNA replication, preventing cancer cells from making new DNA and dividing.
• Examples include Antimetabolites (e.g. methotrexate, 5-fluorouracil): interfere with nucleotide synthesis and DNA polymerase inhibitors and topoisomerase inhibitors (e.g. doxorubicin, etoposide): block DNA replication or cause DNA breaks.
• While these drugs mainly target fast-growing cancer cells, they can also affect normal rapidly dividing cells (like bone marrow and hair follicle cells), leading to side effects.

Question 11

Which pathways are used to repair specific types of damage?

Answer 11

• DNA repair: maintains genome stability, crucial to DNA structure and function. Nucleotide excision repair removes bulky lesions such as UV-induced thymine dimers. Patients with xeroderma pigementosum have a defect in Univision repair that deals with UV damage to DNA = very prone to skin cancer. Other cancer prone families have DNA repair defects
• Base Excision Repair (BER) fixes small base changes
• Nucleotide Excision Repair (NER) removes bulky lesions like UV-induced thymine dimers
• Mismatch Repair (MMR) corrects replication errors.**
• Double-strand breaks are repaired by Homologous Recombination (HR) or NHEJ, while Direct Repair reverses certain chemical modifications.

Together, these systems prevent mutations, maintain genetic integrity, and protect against diseases such as cancer