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33 Cards in this Set
- Front
- Back
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DNA Replication is conservative or semiconservative? What's this mean?
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Semiconservative: means one of chains in "new DNA" helix was the original template
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What is the basic enzyme involved in DNA replication? What are 3 of its important characteristics?
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DNA Polymerase.
1. unable to unwind helix 2. unable to initiate transcription on new strand; it can only add nucleotides to preexisting chain 3. Directs chain growth only in 5-3 direction |
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What are the 3 basic "stages" of Replication
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1. Initiation
2. Elongation 3.Polymerase Editing |
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In E.coli, how many origins are there? What are/is their name(s)?
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1 origin; a segment of DNA that is 245 nucleotides long and called Ori-C (from Origin E. Coli)
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Within the Ori-C sequence, there are two important sequences. What are they?
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1. 9-mers: nine base pair sequence tt is repeated 4 times
2. 13-mers: three repeats of 13 base pair sequence. Ts AT rich region is easy to unzip |
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What are the 5 main genes and protein involved in initiation phase of transcription?
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1. Dna A (protein)
2. Dna B (protein) 3. Dna C (protein) 4. Ssb (protein) 5. Dna G (Gene) |
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What is the function of Dna A protein?
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It forms a multimeric complex tt binds to the 9-mers. Ts complex uses ATP to distort the DNA chain, melting the helix primarily at the weak 13-mer region
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What is the function of Dna B and Dna C?
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Dna C is a carrier protein tt delivers Dna B to the origin region. Dna B is a helicase. It moves along the helix using ATP to separate the two strands
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What is the function of Dna G?
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It produces Primase enzymes that synthesizes RNA primer. In E.coli, the helicase and primase (Dna B & G) bind tgtr forming the "Primosome". It's used in both initiation and elongation.
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What is the function of Ssb?
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Multiple copies of a "single strand binding protein (Ssb) keep the unwound DNA in an unpaired configuration to be read and dupilicated.
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Describe the transcription of the lagging strand
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-As the helicase moves forward it exposes new regions of DNA.
- The primase tn lays down a new primer -DNA polymerase tn elongates the primer 5'-3' until it reaches the previous primer. -Polymerase I replaces the RNA primer and DNA Ligase bonds the segments tgtr. |
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What are the 3 DNA Polymerases in E. coli?
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1. Pol I
2. Pol II 3. Pol III |
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What does Pol I do?
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Two functions;
a. 5'-3' exonuclease b. 5'-3' polymerase This allows it to remove and replace the RNA primer nucleotides on the lagging strand |
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What does Pol II do?
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It's not used in replication. It is used in repair of damaged DNA.
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What is the primary replication enzyme (specifically)?
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Pol III; functioning on both leading and lagging strand.
*If mutated, cell will die. |
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What is the primary function of Pol III?
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Synthesizes new DNA complementary to template strands while simu7ltaneously displacing the bound Ssb proteins.
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What is the "Core Polymerase" made up of?
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The α (polymerase proper),
ε (3'-5' exonuclease [editor]), and θ subunits constitute the "core polymerase" |
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WHat dimerizes the core polymerase?
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τ subunit
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What does the ß subunit do?
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provides a clamp mechanism tt holds the core polymerase on the DNA
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What is the "γ complex" and what does it do?
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It is γσσ'χψ subunits function together to form the "γ complex". It loads the ß clamp and trfor the core polymerase onto the DNA
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What does the core polymerase do?
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It compares a free nucleotide to the next requied base and forms the phosphodiester bond wn the right base pair is recognized.
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What does the ß clamp do?
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Holds the core polymerase onto the DNA strand because the core tends to fall off after 10-15 linkages hv been made
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What happens when the core polymerase stalls?
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The ß clamp releases the core fm the DNA because the core came upon the previous primer. This allows the core to recycle to a new primer
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What is the basic error rate for the synthetic rxn? How can this be reduced and wt's the reduced #?
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1 base for every 10^3-10^4 polymerized. This is a high rate but Pol III has both proofreading and exonuclease activity and can "backup" over improper bases. This makes the error only 1 in 10^6.
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In eukaryotes, what is the equivilant to each of the following: Dna A, Dna B, Ssb, Pol III, Combo of pimase, Pol III and Pol I?
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-Dna A = ORC (origin recognition complex)
-Dna B = MCM -Ssb = RPA (replication protein A) -Pol III ~ Pol σ: it however works almost exclusively on leading strand -Combo of pimase, Pol III and Pol I= Pol α combines w/eukaryotic primase to work on lagging strand |
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What is the solution in eukaryotes to the problem of shortening DNA strands after replication?
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Telomeres: an enzyme called Telomerase acts as a reverse transcriptase, elongating the 3' end of the template DNA in the 5'-3' direction, directing short segments of RNA to be added to the end. The telomere segment can act as a template to replicate the final segment of the lagging strand and then fold over to protect DNA fm degredation damage
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Eukaryotic chromosomal DNA has multiple origins of replication. What is the name for each segment of DNA replicated fm an individual origin?
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Replicon
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What is a Replication Unit?
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Those replicons w/sufficiently similar regulatory sequences tt ty r replicated together
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What is the pattern to the sequence of activation of replication units?
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1. Replication units containing "housekeeping" genes are replicated early in S-Phase
2. Active euchromatin regions are replicated next 3. Inactive euchromatin is tn replicated 4. Heterochromatin regions of DNA are replicated *This increases # of most important genes early in cell cycle |
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During DNA repair, how can the mismatch proofreading system distiguish between the old and new DNA strand?
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By the degree of methylation: newly synthesized DNA are free of methyl groups
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What follows behind the replication fork to insure proper base pair matching? What degree of fidelity does it have?
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The Mismatch Proofreading system. The removal and replacement of misadded bases can raise the overall fidelity of the replication/proofreading process to only 1 in 10^9-10^10 bases.
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What are the 3 steps involved in the repair mechanism of damaged DNA?
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1. Recognition and removal of damaged nucleotides by a specific "DNA Repair Nuclease" leaving a gap in the effected strand of the DNA helix
2. A DNA repair Polymerase binds the free 3OH and fills the gap 3. DNA Ligase closes the remaining "nick" |
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What are the main DNA repair nucleases?
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1. AP nucleases for depurination damage
2. DNA glycolases for base excision repair 3. A combo of 3' and 5' nucleases and helicases for nucleotide excision repair |
