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Biochemistry 2 > RNA metabolism > Flashcards

RNA metabolism Flashcards

1
Q

How does RNA differ from DNA

A

RNA has a hydroxyl group on carbon 2

Uracil but not thymine
Single stranded

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2
Q

RNA is both ___ and ___ (functions)

A

Genetic

Catalytic

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3
Q

Three major classes of RNA

A

mRNA
tRNA
rRNA

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4
Q

what is transcriptome

A

Collection of all transcripts in the cell

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5
Q

Three well understood rules of RNA

A

-Messenger RNA encode the amino acid sequences of all the polypeptides found in the cell

  • Transfering and matching aminoacids to triplets in mRNA
  • Ribosomoal RNA are the constituents of ribosomes
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6
Q

RNA can act as ____ in viruses

A

Genetic material

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7
Q

How do microRNA regulate the expression of genes?

A

Via binding to specific nucleotide sequence

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8
Q

Similarities between DNA replication and RNA synthesis

A
  • Addition of complementary nucleotide
  • Requirement of template
  • Direction of synthesis ( 5’ to 3’)
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9
Q

Differences between DNA replication and RNA synthesis

A
  • No primer required
  • Segment of DNA is used as template
  • One strand act as a template
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10
Q

RNA transcript is complementary to ____ and identical to ___

A

Complementary to DNA template strand

Identical to DNA nontemplate (coding) strand

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11
Q

Polymerase II reads the template strand strand in what direction

A

In 3’ to 5’

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12
Q

In what direction usually RNA transcript , DNA template strand and DNA coding strand are written

A

RNA transcript and DNA coding strand->5’ to 3’

DNA template strand ->3’ to 5’

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13
Q

How do nucleotide complement to each other

A

A purine are paired with pyrimidine , A+T->2 bonds

G+C-> 3 bonds

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14
Q

Pyrimidine is ___

Purine ___

A

Pyrimidine->2 rings(adenine, guanine)

Purines->2 ring (uracil,thymine,cytosine)

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15
Q

You always have ___ amount of purines and pyrimidines

A

Equal

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16
Q

In what form do nucleotides come

A

In triphosphate group

ribo-NTPs

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17
Q

There are three phosphates in NTPs: alpha, beta and gamma. What phosphate is connected to the sugar in the nucleotide

A

Alpha phosphate is connected to 5th carbon of the sugar ring

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18
Q

What is required for RNA synthesis

A

DNA template, all 4-ribo-NTPs and Mg

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19
Q

Explain how nucleotides are added to the growing RNA strand

A

The reaction involves two Mg2+ ions, coordinated to the phosphate groups of the incoming nucleoside
triphosphates (NTPs) and to three Asp residues, which are highly conserved in the RNA polymerases of all species. One
Mg2+ ion facilitates attack by the 3′-hydroxyl group on the α phosphate of the NTP; the other Mg2+ ion facilitates
displacement of the pyrophosphate. Both metal ions stabilize the pentacovalent transition state.

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20
Q

What domain of the polymerase has the site where the actual bonding between two nucleotides occur

A

Beta site

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21
Q

How does polymerase decide which DNA strand will be the template?

A

It can copy from any of the strand , but always it copies or “reads” in the direction of 3’ to 5’ and RNA will be from 5’ to 3’ direction

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22
Q

What is TSS

A

transcription start site (+1)- the first nucleotide that is transcribed into RNA

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23
Q

Similar looking sequences are classified as

A

Consensus sequences

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24
Q

Transcription cycle is also called

A

Sigma cycle

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25
Q

Three main stages in sigma cycle

A

Initiation
Elongation
Termination

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26
Q

What are initial TFs that help to bind RNA polymerase to RNA sequence in transcription ?

A

sigma 70 or sigma factor in prokaryotic system

Like TBP in eukaryotic system

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27
Q

Explain the sigma cycle in prokaryotes

A

1) The binding of sigma factor to the promoter, brings in RNA polymerase
2) Two together they initiate the transcription process
3) As the transcription continues, polymerase leaves the promoter region and goes forward and sigma factor leaves( because his role is done)
4) To continue the transcription , NusA comes and helps the polymerase to continue transcription ( elongation)
5) When the transcription stops, NusA leaves and RNA polymerase is free to work again(termination)

28
Q

Three types of RNA polymerase

A

Pol I: synthesis of rRNA
Pol II: synthesis of mRNA and specialized RNA(microRNA,non-coding RNA)
Pol III: synthesis of tRNA, 5s rRNA and specialized RNA

29
Q

What is unique to transcription in eukaryotes

A

Initiator sequence which is at position +1( the first nucleotide transcribed) that has the transcription start site

30
Q

What is meant by various regulatory sequences

A

In DNA promoter sequence to which TFs will come and bind and will regulate the transcription and multiple TFs can come

31
Q

How many polymerases do eukaryotes and prokaryotes have

A

Eukaryotes- 3

Prokaryotes -1 ( only Pol II)

32
Q

How many subunits are there in RNA pol II in humans

A

12

33
Q

What one extra step do transcription have comparing to prokaryotes

A

Assembly ( before initiation), because we need the preinitiation complex

34
Q

RNA exit cleft where and what does it do

A

In RNA polymerase II and RNA is exiting through this cleft

35
Q

What are two first proteins that come in the eukaryotic system when making a transcript?

A

TBP and TFIIB

and sometimes TBP comes with TFIID and TFIIA

36
Q

The assembly in transcription is similar to

A

PIC in DNA replication

37
Q

After all TFs come and bind , then

A

Polymerase II comes, bind to the promoter

more specifically preinitiation complex) and can start the transcription

38
Q

How many nucleotides are usually open when transcribing and who does that

A

Approximately 17 nucleotides
Polymerase II does it
And now it is called transcription bubble

39
Q

What is needed to be done to “move” polymerase II and start transcription

A

Modifications should take place in CTD( carboxyl-terminal domain) for the protein. It should be phosphorylated

40
Q

What does TFIIH is doing in transcription

A

It phosphorylates CTD

41
Q

After CTD is phosphorylated what happens

A

The polymerase escapes the promoter(start of the transcription) , as well as several TFs leave the system at all

42
Q

WHen TFs leave, ___ come

A

Elongation factors come to help the transcription

43
Q

How does termination occur

A

Elongation factor leave
Termination factor come, they help pol II to stop transcribing, CTD is dephosphorylated, and the whole system is disassembled

44
Q

What three modifications will be done to mRNA before it becomes mature and goes out in the cytoplasm

A
  1. 5’ cap
  2. Polyadenylation
  3. Slicing
45
Q

When does 5’ capping occur

A

Very early , only when 20-30 nucleotides are polymerized ( pol II is till working)

46
Q

What is the purpose of 5’ capping

A

To protect from ribonucleases

47
Q

What is attached to mRNA during 5’ capping

A

7 methylguanosine

48
Q

What is the first nucleotide ( usually)

A

Adenine

49
Q

What is the linkage in 5’ cap

A

5’,5’ -triphosphate linkage

50
Q

How does 5’ capping occur by steps

A

1)gamma phosphate in Adenine ( which is in triphospahte form) is removed by phosphohydrolase

2) Guanylyltransferase attaches guanosine to the first nucleotide and removes two phosphate from guanosine ( as it comes in GTP form)
3) guanine-7-methyltransferase that adds methyl group to guanosine

In some cases this enzyme can add methyl groups to the first few nucleotides

51
Q

What part of PIC complex participate in 5’ capping

A

CTD

Synthesis of the cap is carried out by enzymes tethered to the CTD of Pol II.

52
Q

Explain what happens at CTD

A

Cap synthesizing complex come in, performs ll the 5’ capping stuff
As soon as 5’ cap is done , cap synthesizing system leaves as its function is done

And then CBC( cap binding complex) comes and binds 5’ cap to CTD, and this CBC will be there until the transcription is terminated)

53
Q

What is the exception for introns

A

Histone gens do not have introns

54
Q

Exons are ___, introns are __( length)

A

Exons-short

Introns-long

55
Q

Explain self-splicing

A

Occurs in mitochondrial genes and do not require ATP

the 3′-hydroxyl group of guanosine is used as a
nucleophile in the first step of the splicing pathway. The guanosine 3′-hydroxyl group forms a normal
3′,5′-phosphodiester bond with the 5′ end of the intron . The 3′ hydroxyl of the exon that is
displaced in this step then acts as a nucleophile in a similar reaction at the 3′ end of the intron. The
result is precise excision of the intron and ligation of the exons.

56
Q

What is typical to the gene that can undergo self splicing

A

It should have U- A at 5’ end

and G-U and 3’ end

57
Q

What is the name of RNA prteoins that act as splicesome

A

snRNP’s(snurp’s)- small nuclear RNA that never leave nucleus

58
Q

5 snurp’s

A

U1, U2, U4,U5 and U6

59
Q

Splicing by spliceosome occurs with __ introns and self splicing with ___

A

Self-splicing-> group 1 and 2(mRNA,tRNA and rRNA of bacteria, lower eikaryotes, higher plants)
Splicing with spliceosome-> group 3 (mRNA) and 4(tRNA)

60
Q

Spliceosomes has two parts :

A

Protein part and RNA part

61
Q

Splicing by spliceosome require

A

ATP

62
Q

3 important structures in spliceosome splicing

A
  • 5’ end of intron that has GU(donor site)
  • 3’ end of intron that has AG(accepting site)
  • Upstream from 5’ end there is a very important branching site with very important A in it
63
Q

What happens when splicing with spliceosome

A 
The U1 ( binds to donor site) and U2 (binds to A, branching site) snRNPs bind(ATP is used), then the remaining snRNPs (the U4-U6 complex and U5->U 5 goes to 3' end, acceptor site) bind to form an inactive
spliceosome.( the addition always require ATP)
The donor site and A site are brought together ( U1,U 4 dissociates and this requires ATP)

Lariat is formed ( a loop) between G of GU and A, a connection.

This frees up 3’ OH of the exon, which can now attack 5’ end of the exon , connect with it

And the rest Us leave with the lariat

64
Q

Where does splicing with snurps occur

A

on CTD

The same CBC cap is attached as well.

65
Q

Where do poly A tail is added

A

Transcribed beyond cleavage signal sequence , after 180 to 380 bases after the end sequence

66
Q

What is the cleavage signal( nucleotide sequence)

A

AAUAAA

67
Q

How does polyA tail is attached

A

Polyadenylate polymerase, endonuclease and many other proteins recognize the cleavage sequence and come to the transcription site.
Endonuclease removes polymerase II and extra nucleotides
Then polyadenylate polymerase adds multiple adenylate groups with usage of ATP

Biochemistry 2 (17 decks)

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