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Biochemistry 2 > RNA metabolism > Flashcards

RNA metabolism Flashcards

1
Q

3 types of RNA and their functions

A
  1. mRNA: messenger RNA which encode the amino acid of one or more polypeptides specified by a gene or set of genes. Each is complementary to a DNA strand and it carries the genetic message from the chromosome to the ribosome
    1. Transfer RNA (tRNA): it read the information encoded in the mRNA and transfer the appropriate amino acid to a growing polypeptide chain during protein synthesis
    2. Ribosomoal RNA (rRNAS): they are constituents of ribosomes, the intricate cellular machines that synthesized proteins
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2
Q

difference between transcription, translation and transriptome

A
  • Transcription: an enzyme system converts the genetic information in a segment of double-stranded DNA into an RNA strand with a base sequence complementary to one of the DNA strands
    • Translation: the process in which the genetic information in an mRNA molecule specifies the sequence of amino acids during protein synthesis
      Trancriptome: the sum of all the RNA molecules produced in a cell under a given set of conditions
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3
Q

3 similiraties between DNA replication and RNA synthesis

A
  1. addition of complementary nucleotide
  2. requirement of template
  3. direction of synthesis (5prime to 3prime)
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4
Q

3 differences between DNA replication and RNA synthesis

A
  1. no primer required
  2. segment of DNA is used as a template
  3. one template strand
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5
Q

difference between DNA template strand, DNA nontemplate (coding) strand and RNA transcript

A
  1. DNA template strand: the strand that serves as template for RNA synthesis
  2. DNA nontemplate (coding) strand: it is complementary to the template and it is identical in base sequence to the RNA transcribed from the gene, with U in the RNA in place of T in the DNA
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6
Q

lenght of 2 bases and hydrogen bond between them (one purine and one pyrimidine)

A

1,08 nm

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7
Q

what is required for the RNA synthesis (3)

A

DNA template

all 4-ribo NTPs (ATP,GTP,UTP,CTP), Mg2+

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8
Q

how the new nucleotide binds to the one that is already in the RNA during the synthesis

A

The phosphate interacts with the third carbon of the existing ribose, so there is a bond between the 5prime of the incoming nucleotide and the 3prime of the existing nucleotide

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9
Q

what is the role of Mg2+ ions in RNA synthesis

A

It requires 2 Mg2+, coordinated to the phosphate groups of the incoming nucleoisde triphosphate (NTP) and to the three Asp residues, which are highly conserved in the RNA polymerase.
Roles of Mg2+:
- One ion facilitates the nucleophilic attack by the 3prime-hydroxyl group of the a phosphate of the NTP
The other ion facilitates displacement of the pyrophosphate

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10
Q

6 steps of transcription cycle (or sigma cycle) for prokaryotes

A
  1. Sigma factor and RNA polymerase core binds to the DNA promoter
  2. transcription bubbles form (a region of the promoter is unwind to create an open complex)
  3. transcription is initiated (RNA polymerase start to assembled complementary nucleotides of the DNA)
  4. promoter clearance is followed by elongation
  5. elongation continues. Sigma factor dissociates and it is replace by NusA (a protein that helps RNA plymerase to move further down the gene (elongation))
  6. transcription is terminated. NusA dissociates and the RNA polymerase is recycled

** so always, initiation, elongation, termination

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11
Q

what is the typical eukaryote Pol2 promoter

A

Inr = initiator sequence (+1) so it is the place where the first nucleotide is transcribed and it also contains TSS (transcription start site)
** it is the place where the DNA unwind

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12
Q

the promoter has the Inr, the TATA box (the major point of assembly for the proteins of the preinitiation complex of Pol2) and other sequences on the 5prime side of the TATA box. Why these sequences are important

A

Many additional sequences serve as binding sites for a wide variety of proteins that affect the activity of Pol2 and these sequences are important in regulatong Pol2 promoters and differ greatly in type and number (generally located within a few hundreds base pairs of the TATA box on the 5prime side)

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13
Q

how many RNA polymerase the eukaryotes have compare to prokaryotes and which one is very important

A

3 compare to one for prokaryotes
- RNA polymerase 2: synthesis of mRNA and specialized RNA

  • specialized RNAs: can recognize thousands of promoters that vary greatly in sequence but some Pol2 promoters have a few sequences features in common: TATA box near base pair and Inr sequence (initiator) near the RNA starts site at +1
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14
Q

what is the step in the transcription process that eukaryotes have but not prokaryotes

A

assembly: the first step is to recruit many basal transcription factos such as TBP and TF2H that will come bind to the DNA and their assembly will permit the recruitment of RNA pol2

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15
Q

5 steps of transcription in Eukaryotes

A
  1. Pol2 is recruited to the DNA by transcription factors
  2. the transcription bubble forms: it happens at the Inr where the DNA is unwind by the helicase activity of the TF2H
  3. CTD (carboy-terminal domain that is presents in every protein) is phosphorylated by TF2H during initiation. The polymerase escapes the promoter (which means that the transcription is initiated)
  4. transcription elongation is helped by elongation factors (factors that are not involve in the assembly and that helps RNA Poly2 to move forward and elongate) when TF2H and TF2E dissociate
  5. Elongation factors dissociates. CTD is dephosphorylated as transcription terminates, a process facilitated by termination factors
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16
Q

what are the three processes that happen to RNA (primary transcript) before it becomes a mature mRNA

A
  1. 5prime capping
  2. RNA slicing
  3. Poly(A) tail
17
Q

why is the link in 5prime capping is unususal (2)

A
  1. it happens between 5prime-5prime (instead of 5prime-3prime)
  2. the bond is not a phosphodiester bond but a triphosphate bond (involve 3 phosphate groups instead of one) and this is to protect RNA from ribonucleases
18
Q

which genes do not have introns and what is the lenght of introns and extrons

A

histone genes
exons are short (less than bp)
introns are long (up to 20K bp)

19
Q

in the 5prime capping, what is the name of the molecule that link to the first 5prime of the nucleotide

A

7-methyl-guanosine

20
Q

4 enzymes involve in the generation of the 5prime cap

A

phosphohydrolase
guanyltransferase
guanine-7-methyltransferase
2prime-O-methyltransferase

21
Q

how the cap is make and attach to the mRNA

A
  1. there is the cap-synthesizing enzymes that are associated with the CTD of pol2 so it will synthesized the cap but the problem is that it does not bound it to the mRNA
  2. the cap-binding complex will come and the cap-synthesizing complex will leave and the CBC will make the cap bound to the mRNA and it will leave after
22
Q

for introns that can go to the self-splicing, what are their sequences at 3 and 5prime end

A

5prime end: UpA

3 prime end: GpU

23
Q

what are the 2 steps in self-splicing to cleave the intron

A
  • ** self-splicing means that there is no energy uses!!
    1. GTP comes in and the 3prime of this GTP has an OH like in ribose nucleotide and this 3prime OH acts as a nucleophilic attack that breaks the phosphidiester bound between UpA (it attacks the 5prime of the A)
    2. 3prime OH is now attach to the U of the exons and it becomes and this 3prime OH become the nucleophile, completing the reaction (it will attack the GpU phosphodiester bond) so the result will be UpU (because A and G are gone with the introns)
24
Q

examples of genes that do self-splicing (does not require energy)

A

mitochondrial genes and in the nuclear genome

25
Q

characteristics of splicing by spliceosome (when it occurs, what are the proteins, what are 3 primordial elements in the structure of the introns)

A
  • occurs in most mRNAs
  • spliceosome: RNA-proteins called snRNP’s (small nuclear ribonucleotids)
  • 5 snRNP: U1,U2,U4,U5,U6
  • it requires ATP
    3 important structure of the intron:
    1. At 5prime end of the intron, we have the nucleotide GU : it is called a donor site
    2. 3prime, we have the AG : it is called receptor site
    ** these sequences marks the sites where the splicing occurs
    3. there is a structure upstream of the 3prime end that is important : the sequence upstream that contains the A (in red) and this A is very important: the upstream sequence with the A is called branching site
26
Q

whar are the steps of the splicing by spliceosome

A
  1. U1 and U2 comes where U1 binds to the 5prime (GU) and the U2, which requires ATP to bind, will bind to the A in the branching sequence.
  2. U4,U5,U6 come in
  3. additional of all of it will lead to the formation of the spliceosome (the principle is very similar to the pre-initiation complex in the transcription) so as soon as the assembly is complete, the GU of the 5 prime side will bind to the A of the branching sequence and it will form a lariat
  4. the OH on the 5prime exon will do a nucleophilic attack to the 3prime end of the extrons, which cause their binding and the release of the introns
27
Q

some components of the splicing are tethered to what (bind to what)

A

CTD of RNA polymerase2 (like the 5prime cap) and we remember that for the transcription to occurs, the CTD has to be phosphorylated so splicing is tightly coordinated with the transcription process

28
Q

steps of the poly(a) tail formation

A
  • ** Pol2 synthesized RNA beyond the segment of the transcript containing the cleavage signal sequences(including AAUAAA)
    1. The RNA is cleaved by the endonuclease component of a large enzyme complex (associated with the CTD of RNA polymerase2) at a point 10 to 30 nucleotides 3prime to AAUAAA.
    2. Cleavage generates the free 3prime OH that defines the end of the mRNA, to which A residues are immediately added by polyadenylate polymerase (80 to 250 nucleotides long)
29
Q

role of poly(A) tail

A

The poly A tail serves as a binding sites for one or more specific proteins. It probably helps protect mRNA from enzymatic destruction

30
Q

what alternative splicing leads to

A

multiple mRNAs from one gene

Biochemistry 2 (17 decks)

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