Lecture 9 - DNA Replication and Okazaki Fragments Overview

Short segments of DNA synthesized on the lagging strand during DNA replication.

DNA synthesis always proceeds in this direction.

The strand synthesized continuously towards the replication fork.

The strand synthesized discontinuously away from the replication fork.

The enzyme responsible for synthesizing new DNA strands.

DNA replication in a cell begins with the synthesis of a short RNA primer by the enzyme DNA primase.

DNA primase is an enzyme that synthesizes a short RNA primer, which is essential for initiating DNA replication.

The lagging strand of DNA is replicated discontinuously in short fragments called Okazaki fragments. These fragments are initiated by RNA primers.

Rifampicin, an antibiotic that inhibits bacterial RNA polymerase, also inhibits DNA primase, demonstrating the RNA polymerase nature of this enzyme.

Okazaki fragments are not completely destroyed by DNase because they contain short RNA primers that are resistant to the enzyme's activity.

DNA ligase joins Okazaki fragments by using ATP as an energy source, releasing pyrophosphate and attaching AMP to the 5' phosphate of the downstream fragment. This forms a phosphodiester bond between the 3'-OH of the upstream fragment and the 5' phosphate of the downstream fragment.

The two DNA polymerase III molecules that synthesize the leading and lagging strands are held together by a clamp holder protein, allowing them to move in opposite directions.

The lagging strand is synthesized discontinuously, with Okazaki fragments being joined together by DNA ligase.

During DNA replication, the two DNA polymerase III molecules move in the same direction, but the lagging strand is looped around the clamp holder protein.

DNA ligase forms a phosphodiester bond between the 3'-OH of the upstream Okazaki fragment and the 5' phosphate of the downstream fragment by joining the ends of the fragments.

The 3'-OH end of the upstream Okazaki fragment is joined to the 5' phosphate end of the downstream fragment, creating a continuous DNA strand.

DNA ligase requires ATP as an energy source to join Okazaki fragments.

DNA ligase is used to seal the gaps between Okazaki fragments during lagging strand synthesis.

The DNA polymerase used in DNA replication, it has a high processivity when clamped.

A ring-shaped protein complex that encircles DNA and keeps DNA polymerase associated with the DNA during replication.

An enzyme that synthesizes RNA primers necessary for DNA replication. These primers provide a starting point for DNA polymerase.

The site where DNA replication occurs.

The unwinding of the DNA double helix to expose the DNA template for replication. This process is catalyzed by an enzyme called helicase.

Type II topoisomerases, like gyrase in bacteria, convert overwound, positively supercoiled DNA into underwound, negatively supercoiled DNA. This involves nicking both DNA strands, passing a loop through the break, and then re-ligating the strands.

Type I topoisomerase relaxes negatively supercoiled DNA by nicking one DNA strand, unwinding the DNA, and then re-ligating the strand.

Bacterial DNA replication is bi-directional, meaning it proceeds in both directions from an origin of replication. Two DNA polymerase III complexes enter the DNA at the origin and move in opposite directions.

When the two replication forks meet, Topoisomerase IV separates the catenated daughter chromosomes. It does this by making a double-stranded break and then re-ligating the DNA.

DNA polymerase III (Pol III) has a lower processivity, meaning it releases the new Okazaki fragment easily, which is crucial for the efficient synthesis of the lagging strand.

The lagging strand is synthesized in short fragments called Okazaki fragments, primed by DNA primase. DNA polymerase I (Pol I) fills the gaps between these fragments and DNA ligase joins them together to form a continuous strand.

DNA Pol I replaces RNA primers with DNA and joins adjacent fragments together to create a continuous strand.

Eukaryotic DNA polymerases synthesize DNA at a slower rate than bacterial DNA polymerases. This can be problematic as it takes much longer to replicate eukaryotic chromosomes.

The trombone model describes the mechanism of lagging strand synthesis, where the DNA template loops out to accommodate the synthesis of Okazaki fragments. This loop extends as the DNA polymerase moves along the template and retracts as the Okazaki fragments are joined together.

DNA Pol I has lower processivity compared to Pol III, meaning it releases the template more easily. This allows for the efficient removal of RNA primers and the joining of Okazaki fragments.

The polymerase is moving away from the replication fork on the lagging strand.