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Biochemistry > DNA Replication > Flashcards

DNA Replication Flashcards

1
Q

Function of helicase:

A

unwinds DNA

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2
Q

What is the helicase located in prokaryotes?

A

DnaB helicase

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3
Q

What is the helicase located in eukaryotes?

A

MCM protein

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4
Q

Function of topoisomerases:

A

prevents excessive supercoiling of DNA as helicase unwinds DNA
- measured by linking number

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5
Q

Types of topoisomerases:

A
  • type 1: cuts DNA strand 1 time and corrects linking number by 1
  • type 2: cuts DNA strand 2 times and corrects linking number by 2 (DNA gyrase in prok)
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6
Q

Function of DNA gyrase in prok:

A

introduces negative supercoils for packaging

- type 2 topoisomerase

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7
Q

Function of primase:

A

produces RNA complement to DNA template strand de novo

- doesn’t need 3’ OH like DNA polymerase

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8
Q

Function of DNA polymerase:

A

catalyzes the joining of 2 nucleotides by phosphodiester bond

  • synthesis only occurs in 5’ to 3’ direction
  • needs 3’ OH to start
  • can only extend
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9
Q

Function of clamp:

A

binds to DNA polymerase and helps increase processivity of replication

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10
Q

Name of clamp in prok:

A

beta clamp

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11
Q

Name of clamp in euk:

A

PCNA

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12
Q

What is processivity?

A

the ability to carry out continuous DNA synthesis without dissociation

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13
Q

Function of clamp loader:

A

necessary to open and assemble clamp onto the DNA strand

- needs E

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14
Q

Function of SSB (single stranded binding proteins):

A

binds cooperatively to newly separated DNA strand once helicase unwinds DNA
- enhances template exposure

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15
Q

DNA polymerase activities:

A
  • 5’ to 3’ DNA synthesis ONLY
  • 5’ to 3’ exonuclease activity
  • 3’ to 5’ exonuclease activity
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16
Q

What is 5’ to 3’ exonuclease activity?

A
  • occurs after synthesis occurs
  • removes primers and damaged sections of DNA
  • repairs DNA
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17
Q

What is 3’ to 5’ exonuclease activity?

A
  • removes nucleotides from 3’ end
  • proofreads during synthesis
  • increases fidelity of replication
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18
Q

Activities of DNA polymerase I in prok:

A
  • 5’ to 3’ exonuclease activity
  • 3’ to 5’ exonuclease activity
  • 5’ to 3’ synthesis
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19
Q

Activities of DNA polymerase III in e. coli:

A
  • 3’ to 5’ exonuclease activity
  • 5’ to 3’ synthesis
  • has a clamp, which increases processivity
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20
Q

How does RNA primer get removed in prok?

A

5’ exonuclease activity from DNA polymerase 1

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21
Q

How does RNA primer get removed in euk?

A

RNase H1

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22
Q

What happens if there is a problem with helicase or topoisomerase?

A
  • increases the risk of chromosomal breaks, which leads to chromosomes missing or not joined together correctly
  • increases cancer risk
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23
Q

A drug targeting euk or human enzymes will lead to…

A

a decrease or stop replication in dividing cancer cells

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24
Q

A drug that targets prok enzymes will…

A

stop bacterial infections by decreases cell division in bacteria

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25
Q

What do Cipro and other quinolone antibiotics target in prok?

A

topoisomerase type 2

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26
Q

Function of telomerase:

A

adds nucleotides to 3’ end of DNA

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27
Q

How does telomerase add nucleotides?

A
  1. RNA sequence portion of the telomerase is used as template
  2. extends DNA by complementing RNA sequence of telomerase
  3. telomerase slides down to new addition of DNA and adds more compliments
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28
Q

Importance of telomerases:

A

ensures that chromosomes don’t shorten with each replication

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29
Q

Difference between euk and prok chromosomes:

A

euk: linear and in nucleus
prok: circular and in nucleoid

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30
Q

Function of histones:

A

packages DNA in a highly condensed form via wrapping

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31
Q

In order to initiate synthesis of DNA, what has to happen with histones?

A

they have to be removed

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32
Q

What is chromatin?

A

DNA-binding proteins

  • histones
  • non-histones
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33
Q

How do histones wrap back onto DNA?

A

chromatin assembly factors re-package DNA after synthesis is done

34
Q

What is the end replication problem?

A

DNA gets shortened after every replication cycle

- only for linear chromosomes

35
Q

Why does the end replication problem exist?

A
  • RNA primers removed via RNase H1 = only primers on 5’ side of original strand can be replaced by using 3’ OH end of DNA fragment to extend it
  • primer on 5’ end of new strand doesn’t have any DNA in front of it so once it leaves, there’s no 3’ OH to extend it
36
Q

What are telomeres?

A

DNA sequences that are repeated on the end of linear chromosomes
- gets shortened due to end replication problem and eventually only coding sequences are left

37
Q

Cells with rapid division will have higher levels of…

A

telomerase, which will continually extend the telomere

38
Q

What would the complement strand of this following sequence have?

5’ ATG TGC ACG CAA 3’

A

3’ TAC ACG TGC GTT 5’

39
Q

What is the corresponding RNA transcript sequence of the following sequence?

3’ TAC ACG TGC GTT 5’ (anticoding)

A

5’ AUG UGC ACG CAA 3’

40
Q

What is the coding strand given the RNA transcript below?

5’ AUG UGC ACG CAA 3’

A

5’ ATG TGC ACG CAA 3’

- same, just that T and U are different

41
Q

What is a silent mutation?

A
  • different codon created

- same amino acid

42
Q

What is a missense mutation?

A
  • different codon created

- incorrect amino acid

43
Q

What is a nonsense mutation?

A
  • different codon created

- becomes a stop codon

44
Q

What are some proofreading mechanisms?

A
  • DNA replication proofreading: 3’ exonuclease activity
  • transcription proofreading: back tracking
  • translation proofreading: EF-Tu protein
45
Q

What are some DNA errors?

A
  • insertion/deletion
  • depurination
  • deamination
  • tautomeric
  • transitions
  • transversions
46
Q

Definition of insertion/deletion DNA errors:

A

addition/subtraction of one or more nucleotides during copying
- usually in high repeating sequences

47
Q

Definition of depurination:

A

removal of purine group by hydrolysis

48
Q

Definition of deamination:

A

removal of amine group by hydrolysis

49
Q

Definition of tautomeric errors:

A

nitrogenous base has an alternate protonation state, which alters H-bonding between base pairs

50
Q

Definition of transition errors:

A

result in mismatch, but still has one purine and one pyrimidine

51
Q

Definition of transversion errors:

A

result in mismatch with 2 purines or 2 pyrimidines

52
Q

During viral replication, the virus has…

A

long terminal repeats (LTR)

53
Q

LTRs allow:

A
  • primer reuse and recognition

- cyclization of genome into circular chromosomes

54
Q

Enzymes and structures involved in viral replication:

A
  • viral replicase
  • GP120
  • GP41
  • viral reverse transcriptase
  • viral integrase
  • viral protease
  • host DNA polymerase
  • host RNA polymerase
  • host ribosomes
55
Q

What is viral replicase?

A

a RNA dependent RNA synthesis

56
Q

What is GP120?

A

glycoprotein involved in binding to the CD4 receptor during HIV

57
Q

What is GP41?

A

glycoprotein involved in insertion into membrane and mediate fusion with host cell membrane

58
Q

How does viral reverse transcriptase work during a viral infection?

A
  1. RNA dependent DNA synthesis makes a DNA copy of RNA viral genome
  2. DNA dependent DNA synthesis makes a DNA copy of DNA copied viral genome
  3. RNase activity will degrade RNA once DNA is copied
59
Q

How does viral integrase work during a viral infection?

A
  1. cuts into host genome
  2. inserts DNA copy of viral genome
  3. seals into host genome
60
Q

What happens with viral proteases during a viral infection?

A
  • final processing of viral proteins once host cell begins producing enveloped viruses
  • involved in the maturation of virus
61
Q

What happens with host DNA polymerase during a viral infection?

A

copies host genome with newly integrated viral genome

62
Q

What happens with host RNA polymerase during a viral infection?

A

transcribes host gene along with newly integrated viral genes

63
Q

What happens with host ribosomes during a viral infection?

A

translates host proteins and viral proteins

64
Q

6 classes of antiretroviral agents:

A
  • nucleoside reverse transcriptase inhibitors (NRTIs)
  • non-nucleoside reverse transcriptase inhibitors (NNRTIs)
  • protease inhibitors (PIs)
  • integrase inhibitors (INSTIs)
  • fusion inhibitors (FIs)
  • chemokine receptor antagonists (CCR5)
65
Q

Function of NRTIs:

A

block reverse transcriptase activity by competing with active site for the substrate

66
Q

Function of NNRTIs:

A

interfere with substrate binding to reverse transcriptase by distorting active site

67
Q

Function of PIs:

A

interfere with maturation of virus

68
Q

Function of INSTIs:

A

interfere with cutting of host sequence and insertion of the DNA copy of viral genome into host genome

69
Q

Function of FIs:

A

block insertion of glycoprotein into the membrane

70
Q

Function of CCR5 antagonists:

A

block binding of virus to host cell receptors

71
Q

For transcription, what happens if there is a mutation?

A

most likely will change the efficiency of the promoter

72
Q

What happens during initiation of transcription?

A

rate determining step

  1. sigma subunit recognizes promoter sequence at TATA box in promoter
  2. RNA polymerase binds to sigma and forms closed complex
  3. RNA polymerase forms open complex (transcription bubble)
73
Q

What happens during elongation of transcription?

A
  1. short sequence of nucleotides are created (7-10)
  2. sigma subunit dissociates to increase stability of transcription complex
  3. elongation continues
74
Q

Types of transcription termination:

A
  • factor dependent: needs protein to knock RNA polymerase off (rho protein)
  • factor independent: needs formation of an RNA hairpin in an area that blocks RNA polymerase
75
Q

What happens during transcript processing, which occurs after transcription but before translation?

A
  • addition of poly A tail and cap to start of sequence

- splicing

76
Q

What happens during splicing?

A
  • spliceosome mediates it
  • introns removed (non-coding) and loops to form a lariat structure before exons are linked
  • exons joined (coding)
77
Q

The lac operon regulates transcription by…

A

turning on transcription when repressor protein is prevented from binding to operon
- presence of inducer = repressor can’t bind

78
Q

The Trp operon regulates transcription by…

A

allows cell to express Trp synthesis enzyme by turning on when Trp levels are low

79
Q

What happens to the Trp operon when Trp levels are high?

A
  • enough Trp charged tRNA so ribosomes don’t pause when reaching the first Trp codon
  • doesn’t allow 2-3 hairpin to form
  • 3-4 hairpin forms and acts as terminator
80
Q

What happens to the Trp operon when Trp levels are low?

A
  • ribosomes reach Trp codons and waits for Trp charged tRNA
  • allows formation of 2-3 hairpin (anti-terminator)
  • Trp synthesis enzymes created
81
Q

Steps of translation:

A
  1. 30S subunit binds the mRNA and alignment is ensured by initiation factors (IF) and Shine-Dalgarno sequence
  2. initiator tRNA binds to P site
  3. 30S binds with 50S subunit
82
Q

Shine-Dalgarno sequence is utilized by what structure?

A

ribosomes

Biochemistry (9 decks)

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