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Genes to Proteins > Transcription and Control of the Gene Expression - Genes > Flashcards

Transcription and Control of the Gene Expression - Genes Flashcards

1
Q

What is the “Central Dogma”?

A

DNA –> RNA –> Proteins

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2
Q

What do eukaryotic cells have that prokaryotic cells do not?

A

Nuclei - a location in which the genetic data is stored

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3
Q

Define Transcription and Translation

A

Transcription = DNA is copied into RNA
Translation = RNA is used to synthesize amino acids (Proteins)

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4
Q

Does translation occur more quickly in eukaryotic or prokaryotic cells, why?

A

In prokaryotic cells the DNA and the ribosomes are found in the same location and so translation can occur immediately after transcription. In eukaryotic cells RNA has to move from the nucleus to the cytoplasm before it can be translated.

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5
Q

What are the 3 types of bacterial RNA?

A

1) Ribosomal RNA (rRNA)
2) Transfer RNA (tRNA)
3) Messenger RNA (mRNA)

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6
Q

Which enzyme synthesizes RNA?

A

RNA polymerase (RNApol)

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7
Q

In which direction does RNApol transcribe? (compared to the coding/sense strand)

A

5’ –> 3’

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8
Q

What are the 3 phases of transcription?

A

1) Initiation
2) Elongation
3) Termination

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9
Q

What are the two strands in DNA called?

A

The coding/sense strand and the template strand

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10
Q

Is the coding/sense strand or the template strand identical to mRNA?

A

The coding/sense strand is identical to RNA and the template strand is complementary.

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11
Q

What does the complementary mean?

A

A base will bond with its complementary base. (They work well together :D). It does NOT mean identical

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12
Q

What is the complementary pair of a U base? (Uracil)

A

A - Adenine

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13
Q

What does the σ (sigma) subunit do?

A

Finding a promoter and therefore initiating transcription

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14
Q

What happens to the σ (sigma) subunit once transcription is initiated?

A

It dissociates and so the holoenzyme becomes the core enzyme.

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15
Q

What do the β, β’, α and ω subunits do?

A

β and β’ - form the catalytic centre
α and ω - involved in assembly of the enzyme

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16
Q

How does S1 nuclease mapping help scientist define the transcription start sites and what are the steps involved?

A

1) In vitro transcription to make mRNA
2) Denatured heating of DNA to make one strand with a radioactive phosphate
3) Hybridisation of DNA and RNA
4) Removal of DNA that isnt bound to RNA by S1 nuclease
5) Addition of NaOH to remove RNA
Leaves us with a single stranded DNA that is radioactive and the length of RNA. The position of the promoter can be inferred

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17
Q

Define in vitro and in vivo

A

In vitro - “in glass” - outside of cells
In vivo - “in the living” - inside of cells

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18
Q

What is the distance in base pairs from the two promotor sites to the transcription site?

A

-35 and -10 base pairs upstream of the transcription start site

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19
Q

How many base pairs are the promoter sites and how far away from each other are they?

A

They are hexameric - 6 base pairs and are usually 16-19 base pairs apart.

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20
Q

What is the purpose of DNase Footprinting and what is the enzyme used?

A

To find where an enzyme binds to the promoter - the enzyme is DNase 1 which cleaves double stranded DNA

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21
Q

RNApol protects are region of ~40bp from -20 to +20 but cannot bind to that region only. What does this tell us about -35 region of DNA?

A

The -35 region is required for binding of RNApol to DNA but not used during initiation of mRNA synthesis

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22
Q

What are the steps in initiation for transcription?

A

1) RNApol binds to promoter and forms RPc (RP closed)
2) DNA melts and opens up to form RPo (RP open)
3) rNTPs enter and attempt to start initiation (this occurs ~6 times)
4) The promoter is cleared and mRNA synthesis begins

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23
Q

How does termination work?

A

A hairpin loop is formed in the RNA due to inverted repeats in the DNA

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24
Q

What is rho-independent termination?

A

A G:C rich stem with a run of U upstream. The hairpin loop causes RNApol to pause and the weak A:U bp causes RNA to dissociated from DNA, mRNA to dissociate from RNApol and the DNA strands anneal as the transcription bubble closes.

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25
Q

What is rho-dependent termination?

A

The hairpin loop is not followed by a run of U bases. The rho protein tracks along the mRNA behind RNApol and separates DNA from mRNA. When RNApol pauses at a hairpin the protein “catches up” and unwinds the hybrid in the transcription bubble - stopping elongation

26
Q

What are constitutively expressed genes?

A

“House keeping” genes whos functions are needed under all circumstances

27
Q

What determines the level of transcription for a constitutively expressed gene?

A

The strength of the interaction between RNApol and the promoter (K)

28
Q

What type of sugar is lactose?

A

β-galactoside

29
Q

What is lacI?

A

Lac repressor

30
Q

Are genes or proteins italicised?

A

Genes are italicised and proteins have capital letters

31
Q

What is the opposite of a constitutively expressed gene?

A

An inducible gene

32
Q

What are the functions of LacY and LacZ?

A

LacY - lactose permease - carries lactose into the cell
LacZ - β-galactosidase - breaks down lactose and also forms allolactose

33
Q

How does allolactose cause the breakdown of lactose?

A

Allolactose binds to LacI causing it to change shape and no longer bind to the Lac operator. This allows RNApol to bind to the promoter and begin translation of lacY and lacZ and results in the eventual formation of LacY and LacZ which breakdown lactose

34
Q

Where does the helix-turn-helix motif bind?

A

At the major groove - H bonding that isn’t used in base pairing

35
Q

Why is Lac repressor a tetramer if it is recognising a dimer?

A

If DNA were to undergo looping (deformation of the structure) then LacI can bind to multiple points along the operator that are similar so that repression can still occur

36
Q

What is the purpose of palindromic DNA sequences?

A

They act as a recognition site for enzymes that are frequently involved in regulatory genes

37
Q

What is the synthetic equivalent of allolactose?

A

IPTG

38
Q

In the absence of glucose, what happens to crp?

A

In the absence of glucose cAMP levels are high and therefor crp binds to DNA and promotes the binding of RNApol to the Lac operon

39
Q

When is the Trp Operon required?

A

When tryptophan isn’t available

40
Q

What does tryptophan do to the Trp repressor?

A

The Trp repressor binds to tryptophan which changes the shape of the repressor, allowing it to bind to the Trp operator (TrpO)

41
Q

What is an attenuator?

A

It is a conditional terminator - only causes termination in the Trp operon if high tryptophan is present

42
Q

How does the structure leader peptide result in termination in high concentrations of tryptophan?

A

The leader peptide requires tryptophan to form - in high concentrations RNApol does not pause resulting in the formation of a hairpin loop followed by a run of U. This results in no biosynthesis of tryptophan

43
Q

How does the structure leader peptide result in attenuation in low concentrations of tryptophan?

A

The leader peptide requires tryptophan to form - in low concentrations RNApol pauses as it attempts to form the leader peptide. This allows a hairpin structure to form but not followed by a run of U. This results in no termination and so the biosynthesis of tryptophan occurs

44
Q

What do alternate σ-factors do?

A

They bind to core enzymes to allow alternative binding sites for RNApol by recognising different promoter sequences

45
Q

What is the role of σ32?

A

An alterante σ-factor for heat shock genes - to protect the bacteria above 37 degrees

46
Q

What is bacteriophage λ?

A

A bacterial virus that infects E. coli

47
Q

What does lysis and lysogeny mean?

A

Lysis - virus replicates and causes cells burst open
Lysogeny - dormant phase of the virus

48
Q

What occurs during cohesion and attachment?

A

Cohesion - the linear DNA circularises
Attachment - the phage DNA attaches itself to the bacterial DNA (used in lysogeny)

49
Q

What happens in the very early stage of bacteriophage?

A

Transcription of N in the left direction and cro in the right direction

50
Q

What is the function of N in the early stage of bacteriophage?

A

An antiterminator for both further genes in both direction expresses Q and CII genes

51
Q

What happens during the late stage of lysis?

A

Q acts as an antiterminator at the PR’ promoter and allows expression of lysis, tail and head genes

52
Q

What happens during the late stage of lysogeny?

A

CII is a gene activator protein, allowing expression of recombination and CI (λ repressor protein)

53
Q

Which of these genes are active during lysis which during lysogeny and which during both:
- Cro
- CI
- N
- CII
- Q
- CIII

A

Cro - lysis
CI - lysogeny
N - both
CII - lysogeny
Q - lysis
CIII - lysogeny

54
Q

CII is sensitive to bacterial proteases, what is a consequence of this?

A

This determines divergence for lysis or lysogeny. In rich medium, bacterial proteases are active and CII is degraded resulting in expression for lysis genes.
In starvation conditions, bacterial proteases are inactive and CII is stable and lysogeny genes are expressed

55
Q

What does CI (λ repressor) do?

A

A repressor at PL and PR and an activator of PRM - It turns of early genes and activates its own gene expression. Can also repress its own gene when [CI] is high

56
Q

What does Cro do?

A

A repressor at PRM. As [Cro] rises;
- first represses PL
- and then PR

57
Q

What operator sites does Cro bind to as its concentration increases and what does it result in?

A

Cro binds first to OR3 then 2 and 1. Binding to OR3 stops expression of the PRM promoter so no CI is made

58
Q

What operator sites does CI bind to as its concentration increases and what does it result in?

A

CI binds first to OR1 then OR2. These block the PR promoter so no Cro is made. If the [CI] is too high then binding at OR3 occurs and the PRM promoter is block and CI is no longer made

59
Q

What happens to CI affinity for OR2 when CI is bound to OR1?

A

Cooperative behaviour is seen between CI proteins at OR1 and OR2 so affinity for CI at OR2 increases

60
Q

Why do bacteriophages leave the stable lysogeny stage?

A

UV radiation damages DNA causing RecA protein to become an activated protease which cleaves CI between the C and N domain resulting in a huge loss in [CI] and Cro is made and binds to OR3 and lysis begins

Genes to Proteins (4 decks)

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