Flashcards in DNA replication Deck (22)
following replication, each newly replicated DNA double helix contains one intact parental strand and one newly synthesized daughter strand.
replication begins at a site named replication origin and simultaneously moves out in both directions from this origin. Eukaryotes have multiple origins of replication
The lagging strand of DNA is formed by discontinuous copying of the parental strand that runs 3´- to-5´ away from the replication fork. As more of the helix is unwound, synthesis of the lagging strand begins from another primer. The short fragments (100-200 bp in eukaryotes and 1000-2000 bp in prokaryotes) formed by this process are known as Okazaki fragments.
Region where DNA replication begins. Contain unique short repeats. AT rich. Eukaryotes have multiple, bacteria one
Forks are the sites at which DNA synthesis is occurring.
Origin recognition complex (ORC)
consists of 6 protein subunits and binds to the origin of replication. Once recruits Cdc6 and Cdt1, becomes pre-RC. Before replication can initiate, it must be activated by kinases and then it recruits DNA helicase.
Unwinds the parental strands.
Single-strand binding proteins
bind to single strand of DNA and hold in single-stranded conformation. Also protects strand from nuclease attack.
lays down 10-20 bp RNA primer which DNA polymerase can bind to
DNA polymerase alpha
Contains DNA polymerase and primase activity. Does not have proof-reading activity. DNA Pol α synthesizes the first ~20 deoxyribonucleotides after the RNA primer, and is then swapped for Pol δ and Pol ε for lagging and leading strand synthesis, respectively.
DNA polymerase delta
DNA polymerase that synthesizes okazaki fragments
DNA polymerase epsilon
DNA polymerase that synthesizes leading strand
catalyzes formation of phosphodiester bonds between 3´-hydorxyl group and 5´-phosphate group. Fuses okazaki fragments.
Sliding clamp (PCNA)
Associated w/ eukarytoic DNA polymerase delta. Keeps DNA polymerase tightly associated with parent increasing its processivity.
prevents supercoiling of the DNA at the fork. Perform by breaking and rejoining DNA strands.
a reverse transcriptase that uses an RNA template to copy DNA repeats onto the ends of telomeres.
RNA dependant DNA polymerase. Creates cDNA from an RNA template. Mainly associated with retroviruses which then integrate their genome into a host w/ integrase.
Describe the order of events that occur during, the differences between, and coordination of, DNA synthesis on the leading strand and the lagging strand. (7)
ORC (origin recognition complex) binds to the origin of replication. Recruits Cdc6 and Cdt1, become pre-RC (replication complex). Kinase phosphorylates the complex and it recruits DNA helicase.
DNA helicase melts the parent strands bidirectionally
Single strand binding proteins (RPA euk) bind to the parent strands to protect from nuclease attack and hold it in a single-stranded conformation.
DNA polymerase alpha synthesizes the first 20ish nucleotides. It is than swapped by polymerase delta (synthesizes lagging strand) and polymerase epsilon (synthesizes leading strand)
Replisome coordinates synthesis of the leading and lagging strand by rotating the lagging strand 180 degrees.
DNA polymerase (not sure which one) removes RNA primers w/ exonuclease activity and adds dNTP’s in a 5’-3’ direction.
DNA ligase seals nicks left from the fragments.
Describe the "end replication problem" and the activity of telomerase. (2)
The lagging strand cannot be synthesized to the very end because there is nothing for the last primer to bind to (in creation of okazaki fragments). Because of this, the telomeres shorten with replication until the lack of genome initiates cell death.
Telomerase is an RNA dependent DNA polymerase that maintains telomeres by copying the telomeric repeat sequence from an RNA template.
Caused by RecQ helicase mutations
Werner’s syndrome: WRN helicase mutations
Bloom syndrome: BLM helicase mutations
Caused by Pre-RC mutations
partial loss of function in any of the 6 subunit proteins of ORC