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My Packs: (UWO) (BIOCHEM 2280) > (Dr. Heinemann) (Unit C) Topic 18 > Flashcards

(Dr. Heinemann) (Unit C) Topic 18 Flashcards

1
Q

Central Dogma

A

DNA to RNA to Protein

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2
Q

Why is there an RNA step in gene expression?

A

Not required, present for more points for regulation

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3
Q

Reasons for an RNA step in gene expression

A
  1. Provides an amplification which contributes to differential gene expression
  2. RNA can be degraded, expression of a gene can be stopped quickly
  3. Provides additional opportunities to regulate gene expression (especially in eukaryotes)
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4
Q

Consensus sequence

A

The most frequent base at each position in a group of functionally related DNA elements

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5
Q

Promoter

A

DNA sequence required to initiate transcription of a gene or operon

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6
Q

Terminator

A

DNA sequence required to stop transcription

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7
Q

Operon

A

Set of bacterial genes trancribed from a single promoter and thus expressed from a common RNA

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8
Q

In bacteria, what is the transcriptional start site called?

A

+1 site

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9
Q

What are two consensus sequences in the promoter of bacteria?

A

-35 and -10 regions

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10
Q

How do we know that -35 is important?

A

Making mutations and seeing if it has any effect on transcription

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11
Q

Bacterial RNA Polymerase

A

RNAP
* Enzyme that makes RNA transcripts using DNA as a template and nucleoside triphosphates (NTPs) as substrates

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12
Q

Descibe:

RNAP core enzyme

A

Comprised of 4 subunits that never separate

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13
Q

True or False:

RNAP core enzyme can recognize promoters

A

False, they can only make RNA

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14
Q

What is promoter specificity of RNA polymerase determined by?

A

Sigma subunit

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15
Q

What is the combined polymerase core and sigma subunit known as?

A

RNA polymerase holoenzyme

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16
Q

What sequences does sigma make contact with?

A

-10 and -35 sequences

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17
Q

Initiation of Transcription:

Step 1

A

RNAP holoenzyme binds the promoter
* Forms closed complex

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18
Q

Initiation of Transcription:

Step 2

A

RNA polymerase unwinds the DNA strands around the start site
* Forms open complex

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19
Q

Open vs. Closed complex

A

Open complex occurs with unwinded DNA

20
Q

Initiation of Transcription:

Step 3

A

First NTP is brought to the template base pairing with the base at +1

21
Q

Are primers required for transcription?

A

No

22
Q

Initiation of Transcription:

Step 4

A

Using ATP, GTP, CTP, and UTP as substrates
* Chain elongation proceeds in a 5’ - 3’ direction, following base pairing rules

23
Q

What is released when phosphodiester bonds are formed?

A

Pyrophosphate

24
Q

Initiation of Transcription:

Step 5

A

After addition of 5-10 nucleotides, sigma falls off the holoenzyme

25
Q

Initiation of Transcription:

Step 6

A

Transcription bubble moves downstream (5’-3’) with the template DNA re-annealing behind

26
Q

Initiation of Transcription:

Step 7

A

RNA synthesis proceeds until a terminator is reached and RNA polymerase falls off

27
Q

Initiation of Transcription:

Step 8

A

Sigma rebinds RNAP and the cycle is repeated

28
Q

In total, how many steps are there in initiation and replication in transcription?

A

8 steps

29
Q

What are the 2 mechanisms for transcirption termination in prokaryotes?

A
  1. Rho-independent
  2. Rho-dependent
30
Q

Rho-independent termination

A

Also known as instrinsic termination
* Terminator sequences can form loop on it self
* Pushes transcription bubble off and ends transcription

31
Q

Rho-dependent transcription termination

A

Rho protein is attached to RNA
* Transcription slows at terminator
* Rho catches up and pushes polymerase off of DNA

32
Q

What is Rho-independent transcription termination dependent on?

A

Sequence and Secondary structure

33
Q

What is Rho-dependent transcription termination dependent on?

A

ATP

34
Q

How are different promoters transcribed at different levels?

A
  1. Some genes have better -10 and -35 sequences
  2. There is more than one sigma factor (each recognizes different promoter sequences)
  3. Gene specific regulatory proteins
35
Q

What are -10 and -35 sequences known as?

A

Cis element

36
Q

What are gene specific regulatory proteins known as?

A

Trans elements
* Dynamic

37
Q

Negative vs. Positive regulation

A

Negative
* Factors repress transcription

Positive
* Factors activate transcription

38
Q

Protein - DNA interactions

A

One alpha helix fits nicely into the major groove of the DNA
* Sequence specific interactions
* Interactiosn with the DNA backbone

39
Q

What is the preferred carbon source for E. coli?

A

Glucose

40
Q

What happens in E. coli when glucose and lactose is present?

A

Cell metabolizes glucose and represses use of lactose

41
Q

What is the purpose of glucose repression or catabolite repression?

A

Prevents wasteful duplication of energy-producing enzyme systems by repressing transcription of lac & other operons

42
Q

How is transcription obscurred in lac operon?

A

Repressor protein blocks the promoter, preventing RNA polymerase from attaching and initiating transcription

43
Q

Lac operon
* Negative control
* Positive control

A
  • Negative: Lac repressor (LacI)
  • Positive: Catabolite activator protein (CAP)
44
Q

Binding sites of:
* Lac repressor
* CAP

A
  • Lac operator: DNA element that binds Lac repressor
  • CAP binding site: DNA element that binds CAP
45
Q

How does Lac repressor not bind to DNA?

A

Allolactose - a metabolite of lactose - binds to lac repressor
* The allolactose-Lac repressor complex does not bind DNA

46
Q

What allows CAP to bind to DNA?

A
  1. cAMP - A metabolite of glucose metabolism
  2. Low glucose conditions

My Packs: (UWO) (BIOCHEM 2280) (23 decks)

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