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CMMB 411 > Topic 9 (RNA Structure) > Flashcards

Topic 9 (RNA Structure) Flashcards

1
Q

What are the biological functions of RNA?

A
  • information flow (mRNA)
  • structural (rRNA)
  • regulatory (non-coding RNA)
  • enzymatic activity (ribozymes)
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2
Q

What is a unique feature of ssRNA structure?

A

It has rotational freedom and flexibility

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3
Q

How is RNA’s functional diversity comparable to protein’s?

A

Not as functionally diverse as proteins

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4
Q

What is the flow of genetic information (central dogma)?

A

DNA -> RNA -> protein

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5
Q

Why is RNA more vulnerable to exonucleases than DNA?

A

Doesn’t need as long as a half-life and can be transcribed again if necessary. DNA encodes RNA so it must be kept preserved

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6
Q

What are the structural features of RNA?

A

2’ OH and uses U instead of T, otherwise the same as DNA

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7
Q

Is RNA or DNA more stable at high pH? Why?

A

DNA is more stable because the 2’ OH in RNA gets deprotonated in high pH, which makes it less stable

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8
Q

What is the benefit of RNA having the 2’ OH?

A

It serves as an electron acceptor/donor, which allows RNA to fold into various structures

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9
Q

Why is T found in DNA rather than U?

A

T is more stable (DNA needs to be stable), it’s harder to synthesize (requires more energy), so it can’t be used in RNA, and U is detected by DNA repair machinery as a result of C deamination

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10
Q

Why is U used in RNA?

A

It costs less energy to synthesize, and RNA is synthesized many more times than DNA is within one cell’s life, so it’s more energetically favourable

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11
Q

Can RNA be double-stranded?

A

Yes, such as in siRNA and miRNA

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12
Q

What form of DNA can RNA fold into?

A

A-form DNA

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13
Q

What are the groove characteristics of A-form RNA?

A

Deep but narrow major groove, shallow but wide minor groove

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14
Q

What are the structures sometimes formed by dsRNA?

A

Bulges, internal loops, junctions, and hairpins

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15
Q

How does a bulge occur?

A

One base is unable to base pair on one strand, while all other adjacent bases pair regularly. For example, the top strand: 5’-GAC-3’, and the bottom strand: 3’-CG-5’, A in the top strand would bulge

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16
Q

How does an internal loop occur?

A

Incomplete complementarity between two RNA strands

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17
Q

What is the tetraloop structure? What is the associated sequence?

A

A complex structure in which RNA may take that involves non-Watson-Crick base pairing, base stacking, and hydrogen bonds with the backbone; C(UUCG)G

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18
Q

The first base in C(UUCG)G (tetraloop) base pairs with _________ and has base-stacking interactions with _________

A

Base 6 (G); base 2 (U)

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19
Q

The second base in C(UUCG)G (tetraloop) base pairs with _________ and has base-stacking interactions with _________

A

Base 5 (G); bases 1 (C) and 4 (C)

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20
Q

The third base in C(UUCG)G (tetraloop) base pairs with _________ and has base-stacking interactions with _________

A

None; the sugar of base 4 (C)

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21
Q

The fourth base in C(UUCG)G (tetraloop) base pairs/hydrogen bonds with _________ and has base-stacking interactions with _________

A

The P of base 2 (U); base 2 (U)

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22
Q

The fifth base in C(UUCG)G (tetraloop) base pairs with _________ and has base-stacking interactions with _________

A

Base 2 (U); base 6 (G)

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23
Q

The sixth base in C(UUCG)G (tetraloop) base pairs with _________ and has base-stacking interactions with _________

A

Base 1 (C); base 5 (G)

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24
Q

Where is non-Watson-Crick base pairing in the tetraloop structure?

A

Between base 2 (U) and 5 (G)

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25
Q

Where is hydrogen bonding between a base and the phosphate backbone in the tetraloop structure?

A

Base 4 (C) and phosphate of base 2 (U)

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26
Q

Where are the base-stacking interactions in the tetraloop structure?

A

Between bases 1 (C) and 2 (U), 2 (U) and 4 (C), 3 (U) and the sugar of base 4 (C), and 5 (G) and 6 (G)

27
Q

What is unique about the base stacking of base 3 (U) in the tetraloop structure?

A

It is not pi-stacking

28
Q

What is a pseudoknot?

A

An RNA structure that forms through base pairing between non-contiguous complementary sequences (unpaired bases in a hairpin may pair with other bases farther down the sequence)

29
Q

What prevents RNA from adopting a B-form helix?

A

Presence of 2’ OH

30
Q

Where do proteins interact with dsRNA? Why?

A

Minor groove; major groove is too deep and narrow, which decreases its accessibility to proteins, while the minor groove is shallow and wide, which increases its accessibility

31
Q

What do proteins recognize when binding to dsRNA?

A

Secondary structures (hairpins, stem-loops, bulges, etc)

32
Q

Describe how L. monocytogenes works at low temperatures

A

PfrA (TF) is unable to turn on the virulence gene because the ribosome binding site for its transcript is inaccessible due to the secondary RNA structure

33
Q

Describe how L. monocytogenes works at high temperatures

A

The secondary structure of the RNA is melted, forming a bubble where the ribosome binding site is. The ribosome can bind and translates the PrfA protein to turn on the virulence gene

34
Q

Describe regulation of translation using temperature

A

L. monocytogenes can turn on/off gene expression depending on high/low temperature, respectively

35
Q

When is a U:A:U base triple formed?

A

Usually in ssRNA, which helps the folding of tertiary structures (like tRNA)

36
Q

What is the structure of the MLV transcript?

A

5’-Gag-UAG-pseudoknot-Pol-3’

37
Q

How does MLV control translational readthrough of its mRNA?

A

Through the use of a pseudoknot

38
Q

Gag is translated _____% of the time in the MLV transcript

A

90-95%

39
Q

Pol is translated _____% of the time in the MLV transcript

A

5-10%

40
Q

Describe how the pseudoknot works in MLV

A

Deprotonated: 90-95% of the time. Adenine forms a pseudoknot structure in which it crosslinks with another part of the transcript. This terminates translation at the UAG codon (stop codon). Gag is translated

Protonated: 5-10% of the time. Adenine is protonated and does not form a pseudoknot, allowing the ribosome to read through the UAG codon and translate Gag-Pol

41
Q

What is a riboswitch?

A

Another name for the function of the pseudoknot in MLV

42
Q

What kind of protein is Gag?

A

A structural protein

43
Q

What kind of protein is Pol?

A

A reverse transcriptase

44
Q

What are the advantages of having 2 genes encoding structural and enzymatic proteins in tandem?

A

Only need one transcript for two genes that can be regulated independently

45
Q

What is an aptamer?

A

An oligonucleotide/peptide molecule that binds to a specific targeting molecule

46
Q

True/False? Aptamers are man-made

A

True

47
Q

How does an aptamer work? What is an application of this?

A

Binds to a specific biomarker on the surface of the target cell; can be used to trigger apoptosis in cancer cells

48
Q

What does SELEX stand for?

A

Systematic evolution of ligands by exponential enrichment

49
Q

What is SELEX used for?

A

To create aptamers that can bind to specific cells

50
Q

Describe how SELEX works

A
  1. Synthetic synthesis of RNA molecules
  2. Selection of RNAs that bind to a specific ligand (selected based on affinity for their ligand)
  3. Amplification by PCR and mutagenesis (progressively enrich the aptamers with a higher affinity by introducing mutations)
  4. Repeat steps 2-3
  5. Recovery of RNAs with desired affinity
51
Q

SELEXS works similarly to _______ in immunity

A

B-cell antibody synthesis

52
Q

What is GFP?

A

Green fluorescent protein used as a reporter gene in C. elegans

53
Q

Describe how an RNA-fluorophore complex works

A
  1. A metabolite-sensing RNA binds a metabolite
  2. Metabolite binding reinforces RNA structure
  3. Fluorophore binds RNA complex due to reinforced structure
  4. Complex glows, mimics GFPs but allows for a greater range in colours
54
Q

Can RNA be an enzyme?

A

Yes. For examples, ribozymes are able to catalyze reactions like protein enzymes can

55
Q

What are the five processes an enzyme must be able to do?

A
  • bind a substrate
  • carry out a chemical reaction
  • release the product
  • remain unchanged by the reaction it catalyzes
  • repeat the process many times
56
Q

Describe the steps of alkaline hydrolysis

A
  1. 2’ OH becomes deprotonated at high pH
  2. Oxyanion attacks 3’ phosphate
  3. RNA chain breaks, producing a 2’ 3’ cyclic phosphate and an RNA strand with a 5’ OH
57
Q

Alkaline hydrolysis is an example of:

A

RNA self-splicing/degradation as a result from acting as a ribozyme

58
Q

What three things are ribozymes composed of?

A
  • an active site
  • a binding site for the substrate
  • a binding site for the co-factor
59
Q

What is RNase P?

A

The first ribozyme discovered that generates tRNAs from large precursor RNAs through endonuclease activity

60
Q

What are RNase P’s two moieties?

A

RNA: catalyzes cleavage of tRNA precursor, contains co-factor (metal ions)
Protein: facilitates binding to its RNA substrates

61
Q

What does the shape of RNase P resemble?

A

A hammer

62
Q

What is the function of the co-factors in RNase P?

A

Metal ions allow RNase to have more active function to cleave tRNA precursor

63
Q

What does RNase P cleave off of the tRNA precursor?

A

The 5’ end is cleaved shorter than the 3’ end, which allows the 3’ end to bind to amino acids

CMMB 411 (12 decks)

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