DNA Replication
Objective
• At the end of this lecture, student will be able to
– Explain steps of DNA Replication
Replication of DNA
• Replication is a process in which DNA copies itself to produce identical daughter molecules of DNA
• Process by which new identical DNA is formed from existing DNA as template
• Replication of DNA occurs in 5′ to 3′ direction, simultaneously on both the strands of DNA
• On the leading strand-the DNA synthesis is continuous
• On the lagging strand-the synthesis of DNA is discontinuous
• Short pieces of DNA (15-250 nucleotides) are produced on the lagging strand
Copying DNA
• Replication of DNA
– base pairing allows each strand to serve as a template for a new strand
– new strand is 1/2 parent template & 1/2 new DNA
Semiconservative model:
• Parent DNA has two strands complementary to each other
• Both the strands undergo simultaneous replication to produce two daughter molecules
• Each one of the newly synthesized DNA has one-half of the parental DNA (one strand from original) and one-half of new DNA
• This type of replication is known as semiconservative since half of the original DNA is conserved in the daughter DNA
• Synthesis of a new DNA molecule is a complex process involving a series of steps
1. Replication fork formation
2. Primer binding
3. Elongation
4. Termination
Step 1-Replication fork formation
• Double stranded DNA is separated into 2 single strands
• To unwind DNA, interactions between base pairs must be broken & this is performed by DNA helicase
• DNA helicase disrupts H bonding to separate strands into a Y shape – replication fork at replication origin
• Single-stranded binding proteins (SSB) work with helicase to keep the parental DNA helix unwound
DNA Replication
• Large team of enzymes coordinates replication
Replication: 1st step
• Unwind DNA
– helicase enzyme
• unwinds part of DNA helix
• stabilized by single-stranded binding proteins
Starting DNA Synthesis: RNA Primers
RNA primer built by primase serves as starter sequence for DNA polymerase III
Step 2-Primer binding
• Leading strand is the simplest to replicate
• As the DNA strands have been separated, a short piece of RNA called a primer binds to 5’end of the strand
• Primer always binds at the starting point for replication
• Primers are generated by the enzyme DNA primase
Step 3-Elongation
• DNA polymerases are responsible for creating the new strand by a process called elongation
• 5 different types of DNA polymerases are known in bacteria and human cells
• DNA polymerase III is the main replication enzyme involved in leading stand synthesis
• DNA polymerase I is involved in lagging strand synthesis
• Other enzymes are involved in error checking and repair
• Lagging strand begins replication by binding with multiple primers and each primer is only several bases apart
• DNA polymerase then adds pieces of DNA called Okazaki fragments to the strand between primers
• This process of replication is discontinuous as the newly created fragments are disjointed
Leading & Lagging strands
Lagging strand
Okazaki fragments
Joined by ligase “spot welder” enzyme
Leading strand
Continuous synthesis
Step 4-Termination
• After the formation of continuous and discontinuous strands, exonuclease enzyme removes all RNA primers from the original strands
• Primers are then replaced with appropriate bases
• DNA ligase enzyme joins Okazaki fragments together forming a single continuous strand
• Ends of parent strands consists of repeated DNA sequences called telomeres
• Telomeres act as protective cap to prevent nearby chromosome from fusing
• Telomerase enzyme catalyses synthesis of telomere sequences
• Parent strand and complementary strand then coil into helical shape
Replacing RNA Primers with DNA
DNA polymerase I removes sections of RNA primer and
replaces with DNA nucleotides
But DNA polymerase I still can only build onto 3 end of an
existing DNA strand
Telomeres
Protective cap at the end of chromosome to prevent nearby
chromosome from fusing
Telomerase
Enzyme extends telomeres
Replication fork
Summary
• DNA replication
– Involves 2 main steps
• Unwinding
• Replication
– Two strands
• leading strands
• lagging strands
– Replication only in 3I-5I direction