In-Depth Notes on DNA Replication Process

Origin of Replication

• It is the specific location in the genome where DNA replication begins.

• Enzymatic action is initiated here to begin pulling apart DNA strands for replication.

DNA Replication Mechanism

• The process starts with an enzyme finding the origin and unwinding the double helix.

• This unwinding creates a replication fork, characterized by two strands of DNA becoming single-stranded to serve as templates for new strand synthesis.

Replication Fork

• Defined by two regions moving in opposite directions (counterclockwise and clockwise).

• As the replication forks continue separating, new complementary strands are synthesized on each template strand until the entire molecule is copied.

Strands of DNA

• The ends of DNA strands are crucial and are designated as 3′ and 5′.

• DNA replication proceeds in a 5′ to 3′ direction, meaning DNA polymerase must read from 3′ to 5′ on the template strand.

Supercoiling Problem

• Helical structure causes knots (supercoiling) in the DNA ahead of the replication fork as strands are pulled apart.

• Topoisomerase functions to relieve this supercoiling by cutting and rejoining DNA strands to prevent tangling and knots.

Single-Stranded DNA Binding Proteins (SSBPs)

• After unwinding, SSBPs bind to single-stranded DNA to prevent the strands from annealing back together.

• They stabilize single strands during the replication process.

Leading and Lagging Strands

• The leading strand is synthesized continuously as DNA polymerase synthesizes in the same direction as the replication fork.

• The lagging strand is synthesized discontinuously, forming short segments known as Okazaki fragments due to its opposite direction relative to the replication fork.

DNA Polymerase III

• Primarily responsible for synthesizing new DNA strands in the 5′ to 3′ direction.

• Needs a 3′ hydroxyl group to start adding nucleotides, hence the requirement for a primer to initiate replication.

Primase

• An enzyme that synthesizes a short RNA primer, providing a starting point for DNA polymerase III on the lagging strand.

• Once synthesized, DNA polymerase can then extend this primer to form the new DNA strand.

RNA Primer Removal

• The resulting product of the lagging strand has RNA primers interspersed with DNA.

• DNA Polymerase I is responsible for removing RNA primers and replacing them with DNA nucleotides.

DNA Ligase

• After DNA Polymerase I replaces RNA primers, DNA ligase seals the gaps (nicks) between adjoining DNA strands (Okazaki fragments), ensuring continuity in the DNA molecule.

Completion of Replication

• The process continues until the replication forks meet, resulting in two identical double-stranded DNA molecules, each an exact copy of the original.