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MBB164 - Molecular Biology > The Mechanisms of Translation and Transcription > Flashcards

The Mechanisms of Translation and Transcription Flashcards

1
Q

What carriers out transcription?

A

DNA-dependent RNA polymerases (RNAPs)

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2
Q

Between which sites does transcription occur?

A

specific “start” and “stop” sites

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3
Q

At what part of the polynucleotide chain are nucleotides added?

A

the 3’ end of the polynucleotide chain

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4
Q

What synthesises RNA?

A

DNA-dependent RNA polymerases (RNAPs)

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5
Q

In which strand of the DNA is the genetic information transcribed into RNA?

A

the coding/sense strand

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6
Q

What does NTP stand for?

A

nucleotide triphosphate

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7
Q

How are NTPs selected for?

A

by base-pairing with the template strand

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8
Q

At which end of the extending RNA strand are NTPs added?

A

at the 3’ end

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9
Q

What does the RNA Polymerase active site contain?

A

a short RNA/DNA heteroduplex

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10
Q

What is transcription?

A

The process by which genetic sequence information that is stored in double-stranded DNA is copied into single-stranded RNA molecules

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11
Q

What do RNA polymerases do?

A
  • join nucleotides together in a specific order

- determined by the sequence of the DNA

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12
Q

What is the process of transcription intrinsically dependent upon?

A

The DNA template

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13
Q

Where is transcription initiated?

A

specific transcription start sites

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14
Q

Where is transcription terminated?

A

specific transcription termination sites

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15
Q

In what direction is the RNA polynucleotide generated?

A

5’ to 3’

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16
Q

What must occur for complementary base-pairing to happen between DNA and RNA nucleotides?

A

a region of double-stranded DNA found within the active site of the DNA-dependent RNA polymerase is melted into constitutive single strands

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17
Q

How long is the transcription bubble?

A

approximately 10 nucleotides long

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18
Q

What does transcription produce?

A

an RNA transcript with the same genetic information sequence as the coding or sense strand of DNA

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19
Q

What type of chemical reaction occurs via the addition of a nucleotide to the RNA transcript?

A

a nucleophilic substitution reaction

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20
Q

What is the basic mechanism of this nucleophilic substitution reaction?

A
  • 3’ OH group of the RNA attacks the alpha-phosphate group of the incoming NTP
  • the oxygen atom connecting the alpha and the beta phosphate groups is displaced
  • pyrophosphate (diphosphate ester) is released
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21
Q

How is Pyrophosphate/diphosphate ester abbreviated?

A

PPi

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22
Q

How does the 3’ OH group act as a nucleophile?

A
  • has a lone pair of electrons

- giving it a partially negative charge

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23
Q

How does the alpha-phosphate of the NTP act as an electrophile?

A
  • has a partially positive charge

- due to neighbouring electronegative oxygen atoms

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24
Q

In what direction are nucleotide sequences always written?

A

in the 5’ - > 3’ direction

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25
What does the coding strand encode for?
encodes for the functional protein product of the gene
26
Are the template and coding strands gene-specific?
- yes | - it is only specific to that particular transcription "unit"
27
In what direction(s) can two RNA polymerases move along DNA?
- convergently (the RNA polymerases come closer together) | - divergently (RNA polymerases move away from each other)
28
Which sequence do we refer to when talking about the DNA sequence of a gene?
the coding strand's genetic sequence
29
How does transcription termination occur in prokaryotic cells?
via destabilising the RNA/DNA heteroduplex
30
Which enzymes in prokaryotes can synthesise RNA in the absence of DNA?
- polynucleotide phosphorylase - nucleotidyltransferase - poly(A) polymerase
31
What is the E. coli RNA polymerase core enzyme?
- a protein complex containing 5 subunits | - a,a,ß,ß',w
32
What do the beta subunits do in prokaryotic RNA polymerase?
- ß and ß' have a catalytic function - groove between the subunits allows for the binding of the double-stranded DNA - where RNA synthesis takes place
33
What do the alpha subunits do in prokaryotic RNA polymerase?
- a and a bind transcription factors | - which regulates the enzymes activity
34
what does the omega subunit do in the prokaryotic RNA polymerase?
w is involved in the assembly and stability of the prokaryotic RNA polymerase
35
How are prokaryotic RNA polymerases targeted to transcription sites?
- via specific DNA-binding proteins | - sigma factors
36
What does the enzyme polynucleotide phosphorylase (PNPase) do?
- synthesise RNA - requires nucleoside diphosphate (DNPs) - functions equally well in the reverse reaction to degrade RNA
37
What do tRNA nucleotidyltranserases do?
- add a small number of specific nucleotides onto the 3' end of tRNAs - without a DNA template - ensure tRNA molecules have the correct trinucleotide sequence at their 3' end
38
What does poly(A) polymerase (PAP) do?
- extended 3' ends of mRNA molecules with a stretch of A residues - in the absence of a DNA template sequence
39
What is a general feature of gene expression in both prokaryotic and eukaryotic organisms?
the ability of particular proteins to recognise and specifically interact with defined nucleotide sequences within DNA or RNA
40
What does the protein sigma factor do?
- specifically recognises the sequence of NDA at the 5' end of the gene - so that RNA polymerase is correctly positioned to initiate transcription at the correct site
41
What is the region of DNA which RNA binds to to start transcription called?
the promoter region
42
How does the polymerase/sigma factor complex initially bind to the DNA?
- weakly and non-specifically | - so it can scan along the length of the DNA until it interacts with a gene promoter sequence
43
What happens when the RNA polymerase binds to the promoter gene?
- the RNA polymerase adopts an open and catalytically active conformation
44
What forms/happens once the RNA polymerase has switched to an open conformation?
- transcription bubble in the DNA binding site via the double-stranded DNA melting - the enzyme synthesises a short RNA primer
45
When is the sigma factor released?
once the RNA/DNA heteroduplex is of sufficient length
46
What happens once the sigma factor is released?
- RNA polymerase moves away from the promoter | - RNA polymerase engages fully int he process of transcription
47
How is RNA polymerase different from DNA polymerase?
RNA polymerase has inherent DNA unwinding and primate activity
48
What is the stable interaction between the RNAP and DNA dependent on?
the DNA/RNA heteroduplex within the enzymes active site
49
What does destabilisation of the heteroduplex result in?
- the release of RNA polymerase | - transcription termination
50
What would continued transcription result in?
- block transcription events of other genes - generate unwanted RNA sequences that would titrate out RNA binding proteins - wasteful of important nucleotide resources
51
What is the least stable heteroduplex sequence?
A residues in DNA and U residues in RNA
52
What will a continuous stretch of A residues in the DNA template strand result in?
- a continuous stand of U residues in RNA - this continuously unstable heteroduplex will destabilise the DNA/RNA interaction - cause release of the RNA polymerase
53
What methods can be used in prokaryotes to terminate transcription?
- A-rich template strand DNA sequence | - Rho factor helices activity
54
What is Rho?
- a termination factor | - an RNA helices
55
How does Rho work?
- triggers the release of RNA polymerase | - uses the energy released through ATP hydrolysis to drive dissociation of the polymerase from the DNA
56
What are the 3 nuclear RNA polymerases present in eukaryotic cells?
- RNA polymerase I - RNA polymerase II - RNA polymerase III
57
What does RNA polymerase I synthesise?
- rRNA | - single transcript which is later process into 3 of the 4 rRNAs
58
What does RNA polymerase II synthesise?
- mRNA | - noncoding RNAs
59
What does RNA polymerase III synthesise?
- tRNA | - 5S rRNA
60
What do the 3 eukaryotic RNA polymerases have in common?
- common subunits | - a conserved structure which is homologous to prokaryotic RNA polymerase
61
What do the 3 eukaryotic RNA polymerases not have in common?
- unique subunits - all 3 are visually different - all 3 interact with distinct sets of genes
62
How are the genes within mitochondria and chloroplasts transcribed?
by additional RNA polymerases found within these organelles
63
How are eukaryotic RNA polymerases assembled at the promoter?
by protein-complexes / general transcription factors
64
How is eukaryotic RNA polymerase II assembly at the promoter initiated?
- transcription factor IID (TFIID) binds to an A/T rich sequence in the gene promoter region - known as the TATA box - through the TFIID subunit called TATA-box binding protein (TBP)
65
How does the TATA-box binding protein interact with the DNA?
- binds to the minor groove of the double-stranded DNA
66
What does the binding of the TATA-box binding protein to the DNA cause?
- the DNA to bend | - allowing the recruitment of further transcription factors
67
After more transcription factors are recruited to the TATA-box binding protein is bound what happens?
- preinitiation complex (PIC) is assembled at the promoter | - involving RNAPII and a number of general transcription factor complexes (gTFs)
68
Where does RNAPIII termination occur?
- at dA tracts in the coding strand | - resulting in transcripts having oligo (U) sequences at their 3' end
69
How does termination of transcription by RNAPII occur?
- destabilisation of the polymerase/DNA complex - through the action of exoribonucleases the cleavage and polyadenylation complex endonucleolytically cuts the transcript and adds a poly(A) tail
70
What does the poly(A) tail of the RNA transcript allow for?
- binding of specific nuclear proteins which protect the released fragment from degradation
71
What happens to the 3' fragment generated by cleavage?
- it contains an accessible 5' end - targeted by exoribonucleases which degrade downstream - until the RNA polymerase is reached - this displaces it from the DNA
72
In prokaryotes is translation and transcription couples?
- yes - mRNA is translated into protein whilst it is being made by the RNAP - allows for rapid gene expression
73
In eukaryotes is translation and transcription couples?
- no - transcription occurs in the nucleus - translation occurs in the cytoplasm - allows for regulation at different steps in the gene expression pathway

MBB164 - Molecular Biology (10 decks)

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