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MBB164 - Molecular Biology > Dr Mitchell > Flashcards

Dr Mitchell Flashcards

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1
Q

What is RNA bound to protein in?

A

Ribonucleoproteins (RNP)

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2
Q

What are the differences between DNA and RNA helices?

A

The major groove of RNA is deeper and narrower than that of B-form DNA

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3
Q

What are the unusual base interactions within RNA?

A
  • Non-canonical base-pairs
    - Base-triples
    - Tetraloops RNAs
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4
Q

What are the tertiary structure motifs in RNA?

A
  • A pseudoknot structure

- The A-minor motif

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5
Q

What are the percentages of mRNA, rRNA and tRNA in the cellular RNA profile?

A

5%, 75% and 10%

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6
Q

How does rRNA differ from its primary transcripts?

A

The sections are separated by extra genetic information which is removed, leaving separate shorter mature RNAs

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7
Q

How does tRNA differ from its primary transcripts?

A

Its transcript has extra genetic information either side which is removed to live a single shorter mature RNA

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8
Q

How does mRNA differ from its primary transcripts?

A

The sections are separated by extra genetic information which is removed and polyadenylation occurs, leaving a single shorter mature RNA

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9
Q

To which end of the RNA are new NTPs added?

A

The 3’ end

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10
Q

What region of DNA is the Pribnow box?

A

-10

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11
Q

What are the 5 subunits in the core RNA polymerase of E.coli?

A

a, a, B, B’ and w

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12
Q

What is the function of the beta subunits in the core RNA polymerase of E.coli?

A

Catalytic activity

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13
Q

What is the function of the alpha subunits in the core RNA polymerase of E.coli?

A

Binds transcription factors

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14
Q

What is the function of the omega subunit in the core RNA polymerase of E.coli?

A

Assembly and stability

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15
Q

What are the functions of the sigma factor in the RNA polymerase holoenzyme?

A
  • Increases affinity for promoters

- Decreases nonspecific DNA binding

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16
Q

How does transcription initiation occur in E.coli?

A
  • Sigma factor subunit in transcription bubble scans for promoter
    - Promoter binds and DNA melts
    - Primer synthesised (start of mRNA)
    - When RNA primer is 18 nucleotides long, the sigma factor is released and NusA protein binds
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17
Q

What are the intrinsic terminators in E.coli?

A
  • RNA stem-loop structure
    - G-C rich (stable) region at stem base
    - 3’ U-rich tail
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18
Q

What is the method of rho-independent transcription termination in E.coli?

A
  • The polymerase pauses at the hairpin structure of intrinsic terminator
    - rU-dA base-pairs destabilise the RNA-DNA hybrid within the polymerase
19
Q

What is the method of rho-dependent transcription termination in E.coli?

A
  • rho is an ATP-dependent RNA helicase
    - rho binds to C-rich terminator sequences within the RNA and then catches up with the polymerase
    - rho uses its helicase activity to unwind the RNA/DNA base-pairing
20
Q

What are the functions of the three RNA polymerases in eukaryotic cells?

A

pol I -rRNA
pol II - mRNA
po III - 5S, tRNA

21
Q

What is unique about the catalytic subunit of pol II in eukaryotic cells?

A

It has an extended C-terminal domain

22
Q

What is the method of transcription initiation in eukaryotes?

A
  • Preinitiation complex assembled at pol II promoters
    - TATA box binding protein in TFIID binds to the DNA followed by TFIIB
    - pol II binds to the DNA with TFIIF
    - TFIIE and TFIIH bind
    - The helicase activity of TFIIH melts the DNA duplex, allowing priming
23
Q

What is the torpedo model of transcription termination in eukaryotes?

A
  • Transcript cleaved by cleavage/polyadenylation complex
    - Xrn2 breaks downstream fragment
    - Polymerase displaced
24
Q

What are the three types of mRNA processing?

A
  • Capping of the 5’ end
    - Removal of introns (splicing)
    - Polyadenylation of the 3’ end
25
Q

How does RNA processing occur in eukaryotes?

A

The C-terminal domain on pol II is differentially phosphorylated during transcription, allowing coupling of transcription with RNA processing events

26
Q

What is the structure of the mRNA cap in eukaryotes?

A

On the 5’ end. Two nucleotides bound 5’ to 5’ so can’t be hydrolysed

27
Q

What is the method of 3’ end processing in eukaryotic mRNAs?

A
  • 3’ end formed by coupled cleavage and polyadenylation
    - poly(A) polymerase adds nontemplated adenylate residues to the 3’ end
    - Cleavage and polyadenylation occurs 3’ of the consensus sequence AAUAAA
28
Q

How are intronic and exonic sequences distinguished?

A

Through the recognition of highly conserved splice site sequences

29
Q

How are spliceosomes formed?

A

They are assembled and disassembled from snurps (small nuclear RNPs)

30
Q

What is the mechanism of splicing?

A
  • Involves 2 transesterification reactions
    - The 2’ hydroxyl group of the branchpoint adenosine attacks the 3’ phosphate of the 5’ exon
    - The 5’-2’ phosphodiester bond gives a looped lariat
    - The generated 3’ hydroxyl group attacks the 5’ phosphate of the 3’ eon, releasing the lariat
31
Q

What are the possible noncanonical base-pairings?

A
  • A modified G nucleotide at the first position of the anticodon can recognise codons ending with C or U
    - Inosine base-pairs with A, C or U
32
Q

What are the 21st and 22nd amino acids?

A
  • Selenocysteine can be formed from cysteine to make selenoproteins
    - The codon UAG can encode pyrrolysine which is made from lysine in some archaeabacteria
33
Q

What is the method of tRNA generation?

A
  • RNase P generates the 5’ end by removing the additional genetic information to the dies
    - The 3’ end is generated by endo- and/or exonucleases
    - tRNA nucleotidyltransferase adds CCA to the 3’ end
    - Any introns get removed
34
Q

How are tRNAs folded into an L-shape?

A
  • Coaxial stacking of helices and base -pairing between the ends of the TPsiC and D loops produce a flat, L-shaped molecule
    - The anticodon loop and aminoacyl group are positioned at opposite ends of the molecule
35
Q

How are tRNAs charged?

A

They are charged with the appropriate amino acid by aminoacyl-tRNA synthetases

36
Q

What is the structure of a ribosome?

A
  • Two subunits (Large and small) and 3 tRNA binding sites
    • Codon/anticodon binding occurs on the small subunit at the decoding centre
      - Peptide bond formation occurs on the large subunit at the peptidyltransferase centre
      - Has a polypeptide exit tunnel in the large subunit
37
Q

What is the method of ribosome synthesis in eukaryotes?

A
  • Transcription and early assembly
    • Modification
    • Processing
    • Transport out of the nucleolus to the cytoplasm and late assembly
38
Q

What are the three ribosomal tRNA binding sites?

A
  • The A (aminoacyl) site
    • The P (peptidyl) site
    • The E (exit) site
39
Q

What is the method of the translation elongation cycle in ribosomes?

A
  • Amino acids and mRNA at P and A
    • Bond forms between them and P detaches
    • mRNA moves up so first codon at E instead of P and amino acids move to P as another attaches to A
40
Q

What is the role of GTPases in translation elongation?

A
  • aa-tRNA brought to the ribosome by EF-Tu (A GTPase) and a GTP is hydrolysed
    • EFG (Another GTPase) translocates the amino acid and a GTP is hydrolysed
41
Q

How is the initiation codon recognised?

A

Using the prokaryotic initiator tRNA (tRNAMetf) in which the Met is formylated. In eukaryotes the initiator tRNA is not modified (Meti not Metf)

42
Q

What is the method of translation initiation in E.coli?

A
  • Initiator tRNA binds the P sire and is bound by initiation factor IF2 in a ternary complex
    • 30S/mRNA/tRNA complex formed and 50S subunit associates
    • Initiation complex formed by GTP hydrolysis by IF2
43
Q

What is the method of translation initiation in eukaryotes?

A
  • Ternary complex formed as initiator tRNA binds to GTP-charged eIF2
    • Ternary complex binds 40S and other factors
    • Complex binds 5’ end of mRNA through interactions with cytoplasmic cap binding complex, eIF4F
    • Pre-initiation complex scans mRNA until it finds an AUG codon in the Kozak sequence
    • GTP hydrolysis by eIF2 and initiation factors released
    • 60S bound using eIF5
44
Q

What is the method of translation termination?

A
  • Stop codons recognised by protein termination factors
    • Release factors 1 or 2 bind, triggering peptide hydrolysis
    • RF3 hydrolyses GTP allowing release of release factor
      - Initiation factors, ribosome recycling factor and EF-G dissociate the ribosome

MBB164 - Molecular Biology (4 decks)

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