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Biochem & Cell bio > Unit 9.1 > Flashcards

Unit 9.1 Flashcards

1
Q

Fill in the blank: DNA is the only molecule that_______ b/c it is the database for information

A

Gets repaired

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2
Q

What are the 3 R’s of DNA metabolism

A
  • Replication
  • Repair
  • Recombination
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3
Q

what is replication, repair and recombination

A

replication: the duplication of genetic information
repair: maintenance of information
recombination: cleaving and rejoining DNA for repair and shuffling information

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4
Q

What form of transmission is DNA passed to the next generation? Horizontal or vertical?

A

Vertical transmission- passed to the next generation
not horizontal transmission b/c that involves passing to another cell

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5
Q

What does DNA replication require

A

Fidelity
- faithful copying of each strand
- requires proofreading

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6
Q

What happens if defects in DNA replication are left unrepaired?

A

they lead to mutations and are passed on to the next generation.

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7
Q

describe DNA replication

A
  • uses 1 strand as template
  • Semi conservative mechanism
  • DNA polymerases (actually carry out the synthesis) and others responsible for replication
  • need at least 20 proteins to carry out replication
  • bidirectional replication (2 replication forks, 1 origin)
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8
Q

Meselson-stahl experiment

A
  • uses sedimentation equilibrium centrifugation
  • semiconservative mechanism
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9
Q

describe replication forks

A
  • where proteins assemble for replication
  • assembly points (aka replisome)
  • y shaped structures
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10
Q

what are the required steps for replication?

A

1st. melt/denature the double helix.
- this exposes the bases for selection
2nd. need primer
- a short piece of nucleic acid that hybridizes with the template
- provides the free -OH group on the 3’ end
3rd. add new nucleotides to the 3’ OH end
- hydrogen bonds between template and new nucleotide
- phosphodiester bonds to create a new backbone in the newly synthesized strand
4th. release torsional stress
- topoisomerases

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11
Q

Describe DNA polymerases

A
  • catalyze the addition of complementary nucleotide to the 3’ end of the DNA strand
  • requires: DNA template, Primer (DNA or RNA), and deoxynucleotides (can be dNTP)
  • need Mg2+ b/c to orient the molecules the proper distance for binding
  • synthesis is 5’ to 3’
  • template is read 3’ to 5’
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12
Q

What happens during the DNA polymerase rxn

A
  • Mg2+ is needed to position the nucleotide in the right orientation for binding
  • ## pyrophosphatase cleaves the pyrophosphate
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13
Q

describe proofreading activity

A
  • works backwards 3’ to 5’
  • intrinisic 3’ to 5’ exonuclease activity (cuts at the end of the strand.
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14
Q

What determines fidelity?

A
  • proofreading activity (exonuclease, endonuclease)
  • polymerase activity ( template-directed enzymes, priming produces more accuracy which is why DNA polymerase is more accurate than RNA polymerase)
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15
Q

What is processive in terms of polymerases?

A

polymerases synthesize several bonds at the same time before releasing the substrate (strand)

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16
Q

What triggers the proofreading function?

A
  • incorrect base does not fit properly in active site stalling the polymerase and inducing a conformational change
17
Q

What happens if the polymerase proofreading activity is not triggered?

A
  • replication mismatch repair can switch out the base
18
Q

Who has more DNA polymerases prokaryotes or eukaryotes?

A
  • Eukaryotes
19
Q

describe the DNA polymerases in prokaryotes

A

DNA Polymerase I - primer removal, fills gaps in lagging strand
DNA Polymerase II- DNA repair
DNA Polymerase III- major replicative polymerase

20
Q

describe the DNA polymerases in eukaryotes

A
  • Polymerase for initiating replication (contains primase)
  • Polymerase for DNA repair
  • Polymerase for major replicative polymerase (lagging strand)
  • Polymerase for major replicative polymerase (leading strand)
21
Q

Describe helicase

A
  • catalyzes double strand separation of partially melted dsDNA
  • helicase requires ATP hydrolysis for conformational changes to bind to single strand
  • Binds to ssDNA. initial denaturation is done by primosome
  • moves along and separates the double helix
22
Q

What do single-strand DNA binding proteins do?

A
  • stabilize ssDNA by preventing secondary structure from forming
  • protect ssDNA from degrading
  • keep bases accessible for polymerases
  • aka helix destabilizing proteins
23
Q

describe primase

A

responsible for primer synthesis
- it is RNA polymerase in prokaryotes
- it is RNA-DNA primer in eukaryotes (Primase- Pola complex)
it interacts with helicase
lagging strand requires multiple priming
-primers are replaced by DNA. RNAse removes it (RNA degrading enzyme)

24
Q

describe DNA ligase

A
  • seals ssDNA breaks in backbone during replication and repair
  • forms phosphodiester bond between 3’ OH and 5’ Phosphate end
  • used in dsDNA
  • requires ATP
25
Q

describe the lagging strand synthesis process

A
  • new RNA primer synthesis by DNA primase
  • DNA polymerase adds to new RNA primer to start a new Okazaki fragment
  • DNA polymerase finishes DNA fragment
  • old RNA primer is erased and replaced by DNA
  • sealing by DNA ligase joins new Okazaki fragment to the growing chain
26
Q

describe polymerase sliding clamp

A
  • clamps the DNA polymerase to keep it from sliding off the template
  • enhances polymerase processivity by preventing dissociation of polymerase
    is initially loaded to the template by a clamp loader enzyme
27
Q

describe the sliding clamp loader

A
  • displaces primase after primer synthesis
  • key organizer of replisome
  • ATP binding to clamp loader opens sliding clamp
  • open clamp moves on to DNA
  • ATP hydrolysis locks sliding clamp around DNA and releases clamp loader
  • DNA polymerase binds to sliding clamp
28
Q

describe E.coli DNA polymerase holoenzyme

A
  • it’s 2 DNA Pol core domains linked through central clamp loading complex
  • interacts with helicase through subunits
29
Q

describe initiation of replication

A
  • occurs at special DNA sequences (origins)
  • is a single site in prokaryotic chromosomes
  • is multiple sites in eukaryotic chromosomes
  • act as assembly sites for initiator proteins that initiate synthesis
  • tightly regulated. the only regulated site in DNA synthesis
  • where you would act if you wanted to stop replication
  • region is AT rich because melting is easier since there are only two H bonds
30
Q

What is the primosome in eukaryotes?

A

Helicase and primase complex

Biochem & Cell bio (20 decks)

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