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Molecular Bio > DNA Replication > Flashcards

DNA Replication Flashcards

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Biology 101 flashcards
1
Q

Direction(s) of replication

A

Bidirectional

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2
Q

Origins of replication in eukaryotes vs. prokaryotes

A

Prokaryotes: 1 origin of replication
Eukaryotes: multiple origins of replication

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3
Q

Differences between Okazaki fragment processing in eukaryotes vs. prokaryotes

A

Prokaryotes: Okazaki fragments get chewed up by DNA pol I
Eukaryotes: Okazaki fragments come off as flaps as synthesis continues past them- flaps get cleaved by endonucleases

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4
Q

Activity of all DNA replication endonucleases

A

3’ -> 5’ activity (like a delete key)

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5
Q

Replisome

A

Complex of DNA replication proteins

Proteins recruit, activate, and inhibit one another

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6
Q

Replicator

A

Sequence containing origin of replication

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7
Q

Initiator

A

Protein that binds to replicator, initiating replication

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8
Q

Gel electrophoresis mobility shift assay (GEMSA)

A

Method of determining protein-DNA interaction
DNA is run on gel by itself and with protein bound to it: DNA by itself migrates down the gel faster than DNA and protein together

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9
Q

Initiation of replication steps in E. coli

A
  1. Initiators bind to replicator
  2. Localized unwinding
  3. Recruitment of helicase and helicase loading protein
  4. Helicase recruits primase and kicks off initiators
  5. DNA pol III recognizes DNA and synthesizes off of primer
  6. Continues until 2 forks run into each other
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10
Q

How many times replicator is used per cell cycle in eukaryotes

A

Once: makes sure that DNA is only replicated once and is replicated completely

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11
Q

Telomerase

A

Enzyme that extends the 3’ ends of DNA

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12
Q

2 parts of telomerase

A 
Telomerase RNA (TER): attaches to 3' end of ssDNA and acts as template
Telomerase reverse transcriptase (TERT): synthesizes DNA off of RNA template
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13
Q

Inhibition of telomerase

A

Telomere becomes longer -> more telomere binding proteins accumulate -> telomerase is inhibited

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14
Q

Protein associated with yeast telomeres

A

Cdc 13: recruits telomerase

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15
Q

Protein associated with human telomeres

A

POT 1: inhibits telomerase

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16
Q

t-loop

A

Masks ends of telomeres (ends are folded into middle of dsDNA)
Protects telomeres from random fusion events and acts as a method of telomere length control

17
Q

Major themes of initiation control in eukaryotes

A
  1. Helicase can only load onto the origin replication complex during G1 phase
  2. Licensing factors (kinases that control the start of replication) can only bind to helicase during S phase
  3. After use, licensing factors are degraded and can only be remade during G1 phase
  4. Once helicase splits, it can’t come back together
18
Q

How elongation works

A

Hydroxyl group of growing strand attacks alpha-phosphate of incoming nucleotide
Incoming nucleotide binds to complementary nucleotide

19
Q

Fidelity

A

Accuracy of polymerase

20
Q

Kinetic selectivity of polymerase

A

Polymerase can accept the wrong base, but reaction runs faster when binding is correct

21
Q

How polymerase is selective

A

Enzyme contains selectivity pocket: base pairing needs to be correct for ribose sugar to be correctly positioned in active site

22
Q

Discriminator amino acids

A

Can discriminate between dNTP (DNA) and rNTP (RNA) to prevent DNA-RNA hybrids from being formed
Contained in polymerase

23
Q

Polymerase structure

A

Situated like a hand
Fingers: alpha helices
Thumb: alpha helices
Palm: beta-pleated sheets

24
Q

I helix

A

Thumb of polymerase: maintains correct positioning of template (specifically 3’ end) at active site

25
Q

O helix

A

Finger of polymerase
Coordinates active site (makes sure that dNTPs are properly oriented)
Acts as a hinge that opens and closes active site
Contains amino acids (Lys, Arg) that interact with incoming -PO3
Tyr stabilizes base through stacking
Has Mg+2 that primes -OH for nucleophilic attack

26
Q

What happens when a mismatched nucleotide is added to the growing strand

A

Polymerase slows down and DNA unwinds
Incorrect nucleotide is flipped into the exonuclease site
Exonuclease clips DNA 1 mistake at a time
DNA synthesis resumes

27
Q

Processivity

A

How long polymerase can synthesize strand before it falls off of the template

28
Q

Clamp protein

A

hPCNA (human proliferating cell nuclear antigen)

Holds polymerase onto template

Molecular Bio (10 decks)

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