DNA Replication in Prokaryotes

Submitted by:Fatima Parvez

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DNA ReplicationA process in which daughter DNAs are

synthesized using the parental DNAs as template.

Transferring the genetic information to the descendant generation.

Prokaryotic DNA Replication

•DNA replication is semi conservativeEach strand of template DNA is being copied.

•DNA replication is semi discontinuousThe leading strand copies continuouslyThe lagging strand copies in segments

(Okazaki fragments) which must be joined•DNA replication is bidirectional

Bidirectional replication involves the formation of replication forks, which move in opposite directions

DNA ReplicationDNA replication includes:

• Initiation – replication begins at an origin of replication

• Elongation – new strands of DNA are synthesized by DNA polymerase

• Termination – replication is terminated differently in prokaryotes and eukaryotes

Prokaryotic DNA Replication•The chromosome of a prokaryote is a circular molecule of DNA.•Replication begins at one origin of replication and proceeds in both directions around the chromosome.

Bidirectional replication of circular DNA molecules

Initiation•The enzymes involved in DNA replication are closely associated in one large complex called, the replisome.•The replisome consists of

• the primosome - composed of primase and helicase

• DNA polymerase III molecules• DNA gyrase, topoisomerase, SSB, ligase,

initiator proteins•The replication fork moves in opposite direction, synthesizing both strands simultaneously.

DNA Replication in Bacteria•To begin DNA replication, unwinding enzymes called DNA helicases cause the two parent DNA strands to unwind and separate from one another at the origin of replication to form two "Y"-shaped replication forks.

•These replication forks are the actual site of DNA copying.

DNA Replication in Bacteria•Helix destabilizing proteins bind to the single-stranded regions so the two strands do not rejoin.•Enzymes called topoisimerases produce breaks in the DNA and then rejoin them in order to relieve the stress in the helical molecule during replication.

Model for the formation of a replication bubble at a replication origin in E. coli and the initiation of the new DNA strand

Model for the “replication machine,” or replisome, the complex of key replication proteins, with the DNA at the replication fork

• Primase binds to the first priming sequence on the leading strand template and synthesizes a short RNA primer that is complementary to the DNA template.DNA Polymerase III uses the primer to initiate DNA synthesis by adding deoxyribonucleotides to its 3’ end. The leading strand requires only one priming event, because DNA synthesis is continuous thereafter, in the 5’ 3’direction.

Elongation

DNA Replication in Bacteria•As the strands continue to unwind in both directions around the entire DNA molecule, new complementary strands are produced by the hydrogen bonding of free DNA nucleotides with those on each parent strand

•As the new nucleotides line up opposite each parent strand by hydrogen bonding, enzymes called DNA polymerases join the nucleotides by way of phosphodiester bonds.

DNA Replication by Complementary Base Pairing

DNA Replication in Bacteria• DNA polymerase enzymes are only able to join the phosphate group at the 5' carbon of a new nucleotide to the hydroxyl (OH) group of the 3' carbon of a nucleotide already in the chain.

•As a result, DNA can only be synthesized in a 5' to 3' direction while copying a parent strand running in a 3' to 5' direction.

DNA Replication in BacteriaDNA polymerase enzymes cannot begin a new DNA chain from scratch.

• It can only attach new nucleotides onto 3' OH group of a nucleotide in a preexisting strand.

• To start the synthesis of the leading strand and each DNA fragment of the lagging strand, an RNA polymerase complex called a primosome or primase is required.

• The primase is capable of joining RNA nucleotides without requiring a preexisting strand of nucleic acid - forms what is called an RNA primer .

RNA primer

DNA Replication in Bacteria

•After a few nucleotides are added, primase is replaced by DNA polymerase.•DNA polymerase can now add nucleotides to the 3' end of the short RNA primer.

DNA Replication in Bacteria•The two strands are antiparallel –

• one parent strand - the one running 3' to 5' is called the leading strand can be copied directly down its entire length .

• the other parent strand - the one running 5' to 3' is called the lagging strand must be copied discontinuously in short fragments called as Okazaki fragments.

• Okazaki fragments are of around 100-1000 nucleotides each as the DNA unwinds.

•For the lagging strand, DNA synthesis is discontinuous and requires a series of RNA primers. DNA is synthesised at the 3’ end of each primer, generating an okazaki fragment that grows untill it meets the adjacent fragments. The RNA primer is then removed by the 5’ to 3’ exonuclease activity of DNA polymerase I and replaced with DNA by the polymerase activity of the same enzyme.

DNA ligase links together adjacent Okazaki fragments with covalent, phosphodiester bonds.

It joins the 5’ phosphate of one DNA molecule to the 3’ OH of another, using energy in the form of NAD (prokaryotes) or ATP (eukaryotes). It prefers substrates that are double-stranded, with only one strand needing ligation, and lacking gaps.