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Biochemistry Final Prep > Chapter 36&37: RNA Synthesis & Regulation in Prokaryotes > Flashcards

Chapter 36&37: RNA Synthesis & Regulation in Prokaryotes Flashcards

1
Q

Similarities between replication and transcription

A
  • Both use DNA as template
  • Phosphodiester bonds formed in both cases
  • Both synthesis directions are from 5’ to 3’
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2
Q

Differences between replication and transcription

A

Replication: Transcription:
template: double stands single strand
substrate: dNTP NTP
primer: yes no
enzyme: DNA polymerase RNA polymerase
product: dsDNA ssRNA
base pair: A-T, G-C A-U, G-C

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3
Q

What are operons?

A

Clusters of genes encoding for enzymes of metabolic pathways (on the chromosome)

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4
Q

What do operons allow?

A

Coordinated regulation and gene expresssion

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5
Q

What is an operator?

A

A regulatory sequence adjacent to an operon that determines whether it is transcribed

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6
Q

___________ interacts with operators to control transcription of the genes.

A

Regulatory proteins

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7
Q

What is the difference between polycistronic and monocistronic operons?

A

Polycistronic can make/transcribe several genes because it has multiple genes controlled by 1 operon.
Monocistronic can only make 1 gene (operon only controls 1 gene).

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8
Q

What enzyme carries out transcription?

A

RNA polymerase

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9
Q

What does RNA polymerase need to carry out transcription?

A
  1. DNA template
  2. Activated precursors in the form of the 4 nucleotides (ATP, GTP, UTP, CTP)
  3. Divalent metal ions- Mg2+ or Mn2+
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10
Q

What are the two strands of DNA called and what is the difference between them?

A

Coding strand and template strand- coding strand looks like the mRNA, template strand used to make mRNA (complementary to mRNA)

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11
Q

How do nucleotides get added to the growing nucleotide chain? (direction and what attacks)

A

Grows in the 5’ to 3’ direction.

3’ OH of the growing chain attacks the inner most phosphoryl group of the incoming nucleotide.

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12
Q

In prokaryotes, ___ RNA polymerase synthesizes the 3 major classes of RNA: ______, _______, ______.

A

In prokaryotes, 1 RNA polymerase synthesizes the 3 major classes of RNA: messenger RNA (mRNA), ribosomal RNA (rRNA), and transfer RNA (tRNA).

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13
Q

What is the structure of RNA polymerase?

A

Multisubunit protein= 1 omega, 2 beta, 2 alpha, 2 beta’, 1 sigma
*everything but sigma is core enzyme, adding sigma makes holoenzyme

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14
Q

What are the functions of the subunits of RNA polymerase?

A

alpha- assembly, activation of enzyme by regulatory proteins
beta- binds NTPs, interacts w/ sigma, forms catalytic site with beta’
beta’- binds nonspecifically to DNA, forms catalytic site with beta
sigma- recognizes promoter in DNA, decreases affinity for nonpromoter regions

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15
Q

What directs RNA to proper initiation site?

A

Promoters- specific DNA sequences

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16
Q

What are the 2 DNA sequences that act as a promoter in E. coli?

A
  1. -10 sequence (Pribnow sequence)

2. -35 sequence

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17
Q

What is the difference between strong and weak promoters?

A

Strong promoters- closely match the consensus sequence
Weak promoters- have multiple substitutions at consensus sequences
**other sequences upstream of promoter and action of transcription factors can enhance promoter effectiveness

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18
Q

The 3 phases of transcription of prokaryotes are:

A
  1. initiation- RNA-pol. recognizes promoter and starts transcription
  2. elongation- RNA strand is continuously growing
  3. termination- RNA-pol. stops synthesis and RNA is separated from DNA template
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19
Q

First step in initiation.

A

RNA-pol recognizes -35 sequence, slides to -10 region, then opens DNA duplex.

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20
Q

How long is the unwound region during initiation?

A

about 17 +/- 1 base pair

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21
Q

What has to happen to unwind the DNA during initiation?

A
  • RNA polymerase searches for/binds promoter.
  • RNA pol. and promoter undergo conformational change from closed complex (DNA double stranded) to open complex (unwound)
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22
Q

What is the rate-limiting step of initiation/transcription?

A

Conformational change allowing DNA unwinding

23
Q

After DNA is unwound, __________ can take place.

A

elongation can take place

24
Q

The first nucleotide on the RNA transcript is always a __________.

A

Purine triphosphate

*GTP more often than ATP

25
Q

Does RNA polymerase need a primer to start RNA synthesis?

A

No, can start de novo.

26
Q

What begins the process of elongation?

A

The release of the sigma subunit.

27
Q

Why does releasing the sigma subunit allow elongation to start?

A

Release causes a conformational change of core enzyme, so it can slide on DNA template toward 3’ end.
*During elongation, free NTPs added sequentially to 3’OH of nascent RNA strand.

28
Q

What is the transcription bubble?

A

Region containing RNA polymerase, DNA, and RNA product (40ntds).
*transcription bubble moves along DNA as DNA is unwound/rewound; RNA product extrudes from complex.

29
Q

What is an intermediate in RNA synthesis?

A

A DNA-RNA hybrid helix of approximately 8 nucleotides.

30
Q

What happens during termination?

A

RNA-pol. stops moving on DNA template.

RNA transcript falls off from transcription complex.

31
Q

Termination occurs in either _______ or ________ manner.

A

Either rho-dependent or rho-independent manner.

*Both types involve signals on newly synthesized RNA

32
Q

How is termination triggered at rho-dependent termination sites?

A

Rho binds to ssRNA chain, destabilizing RNA-DNA hybrid and terminating transcription.
*has helicase activity and uses ATP

33
Q

How does termination occur in rho-independent termination?

A

The termination signal on the RNA transcript forms stem-loop structures that causes polymerase to pause after the hairpin, terminating the transcription.

34
Q

What is the termination signal?

A

A stretch of 30-40 nucleotides on the RNA transcript.

Made of many GCs followed by series of U.

35
Q

What are the weakest hydrogen bonding basepairs?

A

rU-dA

36
Q

What are 2 antibiotics that inhibit prokaryotic transcription and how do they do so?

A
  1. Rifamycin- binds pol. preventing RNA from exiting.

2. Actinomycin- intercalates btw bases of the DNA, preventing DNA from being used as a template

37
Q

When genes are organized into operons, _____ can be transcribed from the same ______ and at the same time.

A

multiple genes can be transcribed from the same promoter

38
Q

What is transcription controlled by?

A

The binding of regulatory factors.

Regulatory proteins help in activating or repressing gene transcription.

39
Q

What is the benefit of organizing the genome into operons?

A

More efficient.

40
Q

What are the 2 types of transcription factors?

A
  1. repressors- DNA binding proteins that decrease the efficiency of transcription at the promoter
  2. activators- DNA binding proteins that increase the efficiency of transcription at the promoter
41
Q

What is an example of an inducible operon?

A

Lac operon- genes code for enzymes that break down lactose

42
Q

What does it mean to be an inducible operon?

A

Operon is usually off but an be stimulated/activated.

43
Q

When is transcription on and off in the lac operon?

A

Off in the absence of digestable lactose

On when there’s lactose that needs to be digested

44
Q

What happens when the lac operon is off?

A
  1. lacI (repressor) transcribed making active repressor.
  2. Repressor binds to operator, blocks polymerase.
  3. No transcription.
45
Q

How does the lac operon get turned on?

A
  1. Lactose binds lacI repressor, inhibiting its binding to operator.
  2. Polymerase can bind, so genes needed to digest lactose are transcribed.
46
Q

When is lactose used?

A

Only used when glucose is low.

47
Q

What happens when glucose is low?

A

Levels of cAMP build up.

48
Q

What does cAMP bind to?

A

CAP- regulatory protein that is only active when cAMP binds to it.

49
Q

What happens after CAP and cAMP bind?

A

CAP binds to promoter and stimulates RNA Polymerase to bind- speeds up transcription.

50
Q

What happens when glucose levels rise?

A

cAMP levels drop so no longer binds to CAP.

CAP can’t bind to promoter so transcription slows down.

51
Q

How is the lac operon regulated?

A

Positive gene regulation (2 levels):

  1. presence of lactose determines if transcription can occur
  2. CAP in active form determines how fast transcription occurs.
    * * See table in notes for summary
52
Q

What is an example of a repressible operon? (Controlled by negative gene regulation)

A

Trp operon- ON most of the time

53
Q

How does negative gene regulation occur?

A 
TrpR gene (repressor gene) ON all the time- produces inactive repressor protein
When Trp present it binds to repressor, repressor becomes active and can block transcription

Biochemistry Final Prep (6 decks)

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