DNA Replication
DNA Replication Characteristics Accurate • A single chromatin continually replicates itself for the entire duration of the cell’s life Semi-conservative • Parental (original) DNA is not lost • But the parental paired strands are no longer together Organised and Fast • Large number of base pairs, particularly for humans • Multiple origins of replication in chromosomes allowing an organised and quick process
Directional • Replication occurs in both directions using different enzymes (DNA Replication catalysts) and processes o Due to antiparallel structure of paired strands o 3'C must have an OH– attached to it to allow polymerase in the correct direction o Leading Strand = 3’C à 5’C § DNA replication synthesised in 5’C à 3’C direction § Simple and continuous o Lagging Strand = 5’C à 3’C § DNA replication is complex and broken up into segments § Direction of DNA synthesis remains in 5’C à 3’C direction Nucleotides • A = T and C = G • Phosphate group = 3 phosphate ions = nucleoside triphosphate o Phosphate ions hold potential chemical energy which is used in DNA replication in the polymerisation process o Similar to ATP hydrolysis releases ATP
Simple Process:
1. Parental DNA molecule is made up of a specific base pairs 2. DNA separates itself into 2 strands 3. The parental strand’s serves as a "template" for a certain pattern of nucleotides to be added to the original parental strand. o Creating a complementary strand to be added to the original strand. 4. The parental strand and complementary strand connect through reforming a sugar-phosphate backbone. o 2 DNA molecules created
DNA Replication in depth Catalyses the addition of new nucleotides only in one direction (5'C to 3'C) for the replication of DNA
Initiation
1. Helicases o Starts at the origin of replication o Double helix unwinds o It is ahead of the DNA Polymerase so that it can do its job on the single strand o Single-stranded DNA templates are provided 2. Single-strand binding proteins o Behind the helicase but in front of DNA Polymerase o Prevents the helix from reforming again
Synthesis of Leading Strand
The leading strand is the single strand of unwound parental DNA that is in the 3’C à 5’C direction
3. Primase o Join RNA nucleotides to lay down a RNA primer which does not require OH– – • The end of the RNA primer as an OH which then allows further synthesis of DNA replication • Later on this has to be replaced o Once completed RNA Primer synthesis primase falls off the DNA strand 4. DNA Polymerase (III) o Polymerise specific complementary nucleotide triphosphates to the 3’C OH– • H2O and 2 phosphate ions are released • 2Pi provide energy for the polymerisation reaction o Starting at the RNA Primer end • The leading strand is continually elongated in the 5'C –> 3'C direction as the "unzipping" action of the parental DNA occurs o "Sliding Clamp" enzyme is behind the DNA Polymerase ensuring that it does not fall off the single strand of DNA 5. DNA Polymerase (I) o Replaces the RNA primer at the origin of replication with DNA
Synthesis of Lagging Strand
The lagging strand is the single strand of unwound DNA which is antiparallel to the leading strand in that it follows a 5’C à – 3’C direction. Thus DNA replication must be segmented as there is no OH for nucleotides to polymerise onto.
3. Primase o Join RNA nucleotides to lay down a RNA primer which does not require OH– • Later on this has to be replaced o Once completed RNA Primer synthesis primase falls off the DNA strand o Many RNA primers are placed onto the lagging strand 4. DNA Polymerase (III) o Polymerise the addition of nucleotides starting at the end of the RNA Primer and then continually add nucleotides onto the 3'C OH– until it reaches the next RNA primer where it detaches • This forms an Okazaki Fragment 1 o "Sliding Clamp" enzyme is behind the DNA Polymerase ensuring that it does not fall off the strand of DNA 5. DNA Polymerase (III) continued o Step 4 is repeated along every RNA primer, synthesising the DNA in a 5'C –> 3'C direction o Lagging strand thus creating an incremental increase of 1 for each "Okazaki Fragment"
6. DNA Polymerase (I) o Replaces the RNA primer at the origin of replication with DNA • But the DNA is still not connected to the Okazaki fragment to the left of it • Example: Replacing the Okazaki fragment 2's RNA Primer has still left Okazaki fragment 1detatched from the rest of the replicated DNA
7. Ligase o Forms a bond between the new DNA formed from DNA Polymerase (I) and the Okazaki fragment next to it which is detached o Completes the synthesis of the DNA lagging strand
Directional • Replication occurs in both directions using different enzymes (DNA Replication catalysts) and processes o Due to antiparallel structure of paired strands o 3'C must have an OH– attached to it to allow polymerase in the correct direction o Leading Strand = 3’C à 5’C § DNA replication synthesised in 5’C à 3’C direction § Simple and continuous o Lagging Strand = 5’C à 3’C § DNA replication is complex and broken up into segments § Direction of DNA synthesis remains in 5’C à 3’C direction Nucleotides • A = T and C = G • Phosphate group = 3 phosphate ions = nucleoside triphosphate o Phosphate ions hold potential chemical energy which is used in DNA replication in the polymerisation process o Similar to ATP hydrolysis releases ATP
Simple Process:
1. Parental DNA molecule is made up of a specific base pairs 2. DNA separates itself into 2 strands 3. The parental strand’s serves as a "template" for a certain pattern of nucleotides to be added to the original parental strand. o Creating a complementary strand to be added to the original strand. 4. The parental strand and complementary strand connect through reforming a sugar-phosphate backbone. o 2 DNA molecules created
DNA Replication in depth Catalyses the addition of new nucleotides only in one direction (5'C to 3'C) for the replication of DNA
Initiation
1. Helicases o Starts at the origin of replication o Double helix unwinds o It is ahead of the DNA Polymerase so that it can do its job on the single strand o Single-stranded DNA templates are provided 2. Single-strand binding proteins o Behind the helicase but in front of DNA Polymerase o Prevents the helix from reforming again
Synthesis of Leading Strand
The leading strand is the single strand of unwound parental DNA that is in the 3’C à 5’C direction
3. Primase o Join RNA nucleotides to lay down a RNA primer which does not require OH– – • The end of the RNA primer as an OH which then allows further synthesis of DNA replication • Later on this has to be replaced o Once completed RNA Primer synthesis primase falls off the DNA strand 4. DNA Polymerase (III) o Polymerise specific complementary nucleotide triphosphates to the 3’C OH– • H2O and 2 phosphate ions are released • 2Pi provide energy for the polymerisation reaction o Starting at the RNA Primer end • The leading strand is continually elongated in the 5'C –> 3'C direction as the "unzipping" action of the parental DNA occurs o "Sliding Clamp" enzyme is behind the DNA Polymerase ensuring that it does not fall off the single strand of DNA 5. DNA Polymerase (I) o Replaces the RNA primer at the origin of replication with DNA
Synthesis of Lagging Strand
The lagging strand is the single strand of unwound DNA which is antiparallel to the leading strand in that it follows a 5’C à – 3’C direction. Thus DNA replication must be segmented as there is no OH for nucleotides to polymerise onto.
3. Primase o Join RNA nucleotides to lay down a RNA primer which does not require OH– • Later on this has to be replaced o Once completed RNA Primer synthesis primase falls off the DNA strand o Many RNA primers are placed onto the lagging strand 4. DNA Polymerase (III) o Polymerise the addition of nucleotides starting at the end of the RNA Primer and then continually add nucleotides onto the 3'C OH– until it reaches the next RNA primer where it detaches • This forms an Okazaki Fragment 1 o "Sliding Clamp" enzyme is behind the DNA Polymerase ensuring that it does not fall off the strand of DNA 5. DNA Polymerase (III) continued o Step 4 is repeated along every RNA primer, synthesising the DNA in a 5'C –> 3'C direction o Lagging strand thus creating an incremental increase of 1 for each "Okazaki Fragment"
6. DNA Polymerase (I) o Replaces the RNA primer at the origin of replication with DNA • But the DNA is still not connected to the Okazaki fragment to the left of it • Example: Replacing the Okazaki fragment 2's RNA Primer has still left Okazaki fragment 1detatched from the rest of the replicated DNA
7. Ligase o Forms a bond between the new DNA formed from DNA Polymerase (I) and the Okazaki fragment next to it which is detached o Completes the synthesis of the DNA lagging strand
