DNA Replication.

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Since most cells in the body share the same genome (excluding mature red blood cells and haploid cells) it is important that DNA is accurately replicated so both diploid cells get the identical copy.

The process of DNA Replication:

This process is not thermodynamically favorable, meaning energy must be put into the system. The process begins with a DNA/RNA primer – this binds to template strand which is expossed when the DNA is unraveled. The Deoxynucleotide triphosphates the bases (dATP, dGTP, dTTP, dCTP) contain a triphosphate group. High energy phosphodiester bonds are then formed when nucleotide monophosphate is bound to the 3’OH to grow the newly synthesized strand (growing from the 5′- 3′ region). To make this occur at the rapid rate it does in cells, DNA polymerase is required. This enzyme catalysis the release of a pyrophosphate – this breaks down into two phosphates powering the reaction.

Problem: Supercoiling. Have you ever pulled your earphones out of your pocket and found they are coiled together, then you just pull them apart and make it 10 times worse? This is known as super coiling and the same thing would happen in DNA replication if the interwound double helixes were just pulled apart.

Solution: Topoisomerase will cut the strands of DNA to pass them around the other to prevent them from getting tangled in the process (relegate). Additionally, The DNA replication process will only unwinding small sections at a time. This resolves the issue of supercoiling.

DNA is replicated through Initiation (this is what the primer is used for), chain-elongation (this is when the base sequence is added growing the newly synthesised chain), termination (this is when it stops and is released.)

As previously mentioned, Strands of bacteria DNA is circular, this is commonly refered to as dsDNA. For replication of this both strands of dsDNA, they are copied at the same time. Eventually, they meet up again and the newly produced strands are separated forming two dsDNA copies.

The DNA polymerase can only travel from the 5′ to 3′ region, however, in DNA replication both the DNA strands which run in opposite directions need to be replicated. This means two process’ of replication are required. Leading chain and lagging chain replication.

Leading strand replication:

• Helicase enzyme unzips the double stranded DNA while topoisomerase relegates the chain to prevent supercoiling.
• RNA primer initiates the construction of the new DNA strand. (initiation)
• DNA polymerase III travels from 5′ to 3′ adding bases to the growing chain as it goes. (chain propagation)

Lagging strand replication:

• Helicase enzyme unzips the double stranded DNA while topoisomerase relegates the chain to prevent supercoiling.
• RNA primers are placed in small intervals along the chain
• DNA polymerase travels from 5′ to 3′ laying down bases forming ozaki fragments, however, they are not all connected yet.
• DNA polymerase I now has to swap out the RNA primers with DNA
• DNA ligase can now connect all the ozaki fragments together to for a continous chain

Termination for both occurs once the two ends of two loops of the dsDNA join resulting in both being completed.

Memorize.

Master.

Question 1.

Define these structures and enzymes.

Ligase, Helicase, Topoisomerase, Primase, Replication fork

Question 2.

Identify two problems and the biological solution in DNA replication.

Question 3.

DNA-replication requires a number of proteins outline the functions of the major proteins in DNA-replication.

Question 4.

Contrast leading chain and lagging chain replication in DNA.

Question 5.

Outline the general mechanism for copying DNA to
DNA before cell division -replication.

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Answers.

Question 1.

Ligase: Joins together molecules

Helicase: Unzips the DNA strand

Topoisomerase: Required to prevent super coiling by cutting one of the DNA chains to pass it over the other allowing the loops to be removed from the DNA.

Primase:

Replication fork: This is the active area where DNA replication is occuring.

Question 2.

• Super coiling – toposomerase
• Lagging strand replication – DNA polymerase forms ozaki fragments, the primers are swapped out with DNA then DNA polymerase connects all the strands together.

Question 3.

Polymerase connects the strands together forming a continuous chain. Topoisomerase, prevents super coiling by cutting one of the DNA chains to pass it over the other allowing the loops to be removed from the DNA.

Question 4.

Both leading chain and lagging chain replication have the synthesis of the DNA running from the 5′ to 3′ region. However, the lagging strand runs in the opposite direction hence it requires ozarki fragments which are regions of RNA to be laid down. DNA polymerase then uses these primer regions to initiat the laying down of DNA from the 5’to 3′ of the ozaki fragments. The fragments are then swapped out with DNA and and polymerase creates a continuous chain. The loops both then meets up again resulting in termination for both.

Question 5.

Leading and lagging strand synthesis.