DNA Structure and Replication

What are nucleic acids responsible for?

Storage, expression, and transmission of genetic information

What are the 2 classes of nucleic acid?

• Deoxyribonucleic acid (DNA), which stores genetic information coded in sequences of genes

• Ribonucleic acid (RNA), which is involved in protein synthesis and controls gene expression

Bases in DNA

Purines (double ring): Adenine and guanine

Pyrimidines (single ring): Cytosine and thymine

A-T pairing

G-C pairing

What are DNA and RNA made up of?

Nucleotides

What are nucleotides made up of?

Phosphate (backbone)

5-carbon sugar

Ring of C and N atoms, which is the base (genetic information

Primary structure of Nucleic Acids

Polymerisation occurs via condensation reactions. What is polymerisation in nucleic acids?

• The formation of phosphodiester bonds between C3 of one sugar, to C5 of the next sugar.

• It forms the primary structure

• Gives 5’ to 3’ directionality to single stranded nucleic acids

5’ to 3’ directionality. What is the 5’ end and what is the 3’ end?

5’ end = phosphate

3’ end = terminal sugar

What is the secondary structure of DNA?

There are 2 antiparallel chains of nucleotides in a double helix.

• Phosphate backbone

• Hydrogen bonds

• 2 H bonds between A-T

• 3 H bonds between G-C

Nucleus of the cell contains….

…DNA

What is the genetic code?

• The genetic code is the set of rules by which the sequence of DNA nucleotides is translated into a sequence of amino acids.

• Three nucleotides form a codon (triplet code), with each codon specifying a particular amino acid or a stop signal during protein synthesis.

Explain the hierarchical organisation of genetic information in eukaryotic cells.

DNA → Sequence of nucleotides → Triplet code → Amino acids → Polypeptides → Genes → Genome

How is genetic information organised within the nucleus?

• The nucleus of a cell contains DNA

• DNA contains all genetic information encoded in the sequence of nucleotides

• 3 DNA nucleotides code for a specific amino acid called the triplet code (genetic code)

• Many triplet codes will form a specific polypeptide chain called a gene

• A single molecule of DNA contains many genes

• The total genetic information encoded in the DNA is known as the genome

What is the tertiary structure of DNA?

DNA in eukaryotes in the nucleus are compacted into chromosomes

• DNA is bound to histone proteins which form DNA protein particles called nucleosomes

• Nucleosomes associate with each other, forming more compact chromatin fibres

• Chromatin fibres are supercoiled into loops and condensed into a chromosome

How many separate chromosomes are there in all human cells (except egg and sperm) that encode the whole of the genome?

46

DNA replication results in the exact duplication of DNA, producing 2 complete double helices. What are the 3 reasons as to why DNA replication required?

• Reproduction

• Maintenance of cells and tissues

• Growth of cells and tissues

When does DNA replication occur in the cell cycle?

During the S phase in interphase, before the cell divides

Where does DNA replication occur in a eukaryotic cell?

In the nucleus

What is required for DNA replication to occur?

Complementary base pairing and a series of enzymes.

What type of DNA replication is in eukaryotic cells?

Semi-conservative

What is semi conservative replication?

The two strands of parent DNA separate and both strands acts as a template for a new strand.

There is one daughter (new) strand and one parent (old) strand in each new DNA molecule

Name the stages of DNA replication

1. Initiation

2. Elongation

3. Termination

1. Initiation

DNA is very stable and tightly coiled together and compacted into chromosomes

2. Initiation

Initiator proteins are needed to pry open the 2 DNA strands

3. Initiation

Initiator proteins are needed to pry open the 2 DNA strands

4. Initiation

This occurs at the replication origin

5. Initiation

• Replication bubbles are formed

• Prokaryotes have a single bubble, while eukaryotes have many bubbles.

• In these bubbles, there are 2 Y-shaped replication forks

Name the initiator proteins involved DNA replication

• Helicase

• Topoisomerase

• Single strand binding proteins

• Primase

What does helicase do in DNA replication?

Unwinds and separates the two DNA strands by breaking the weak H-bonds using ATP energy

What does topoisomerase do in DNA replication?

Uncoils the supercoiled DNA and relieves stress of the DNA molecule as it separates

What do single-strand binding proteins do in DNA replication?

Attaches to each strand and keep them separated and untwisted

What do primase do in DNA replication?

• Primase acts as a starting point for DNA polymerase

• Primase is an RNA polymerase that synthesised RNA using DNA as a template

• It does not synthesise DNA, but maes short length of RNA using the DNA strand as a template reading in 5’ to 3’ direction

• Serves as an RNA primer for DNA synthesis, hence the name primase

6. Elongation

• DNA polymerase binds to the RNA primer and moves along the replication fork and catalyses the addition of nucleotides. It works from 3’ end (hydroxyl end) of the DNA, using the parent strand as a template

• Free nucleotides in the nucleus bind via complementary based pairing and join via phosphodiester bonds

7. Elongation

• DNA polymerase reads in the 3’ to 5’ direction which means the daughter strand grows in the 5’ to 3’ direction.

• However, the strands in the DNA double helix run antiparallel and DNA polymerase will ONLY add new subunits to the 3’ end.

8. Elongation

• The DNA strand being read from 3’ to 5’ is called the leading strand.

• The other DNA strand is the lagging strand.

9. Elongation

• The other DNA strand is formed via discontinuous ‘backstitching’ → DNA polymerase moves along the template to find the 3’ hydroxyl groups then working back towards the origin or replication.

• These small DNA pieces are called Okazaki fragments.

10. Elongation

• The leading strand will only need one RNA primer at the origin of replication.

• The lagging strand will need several RNA primers to keep polymerisation going.

11. Elongation

• These RNA primers are eventually removed by exonuclease.

• Gaps are filled by DNA polymerase

12. Elongation

• Fragments are then joined by DNA ligase to form a complete strand.

DNA polymerase is self correcting. What does that mean?

DNA polymerase carefully monitors the base-pairing; proofreading.

• Before adding a new nucleotide in the 5’ to 3’ direction, it checks the previously added nucleotide.

• If correct base-pairing it continues.

• If incorrect, it cuts the phosphodiester bond, releasing the incorrect nucleotide and tries again!

Image depicting Initiation and Elongation:

13. Termination

The leading strand can be replicated right up to the end of the strand. The lagging strand cannot.

14. Termination

Over time the lagging strand would get shorter and shorter, losing genetic information.

So instead, long repetitive nucleotide sequences called telomeres are added to the ends of every chromosome, providing “extra” DNA to complete the lagging strand.

15. Termination

Telomeres are attached to these sites by telomerase and they form a ‘cap’ at the end of chromosomes

What is this image showing?

Telomeres shortening with age