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genes and gene expression > Mechanism & regulation of translation I > Flashcards

Mechanism & regulation of translation I Flashcards

1
Q

what is the central dogma?

A

theory stating that genetic information flows only in one direction, from DNA to RNA, to protein or RNA directly to protein

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2
Q

protein synthesis

A
  • crucial for cell growth, proliferation and survival
  • expensive process for the cell, therefore tightly regulated
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3
Q

what can protein synthesis regulation do?

A

control overall rates of proteinsynthesis and modulate the expression of specific transcripts

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4
Q

what can proteinsynthesis be inhibited by?

A

cell stresses and withdrawal of nutrients; serum deprivation, temperature shock, DNA damage, viral infection, hypoxia, cytokine treatment

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5
Q

prokaryotes

A
  • polycistronic
  • normally unstable
  • translation can occur on nascent transcripts
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6
Q

eurkaryotes

A
  • normally monocistronic
  • capped and polyadenylated
  • 5’ and 3’ UTR
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7
Q

what is the structure of CAP?

A
  • found at the 5’ end of all cellular mRNAs
  • seals end of mRNA protecting it from nuclease digestion
  • landing pad for elF4E
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8
Q

what is the process of capping of newly synthesised mRNAs?

A
  1. removal of 5’ terminal phosphate (triphosphatase)
  2. addition of 5’ terminal GMP (guanylyl transferase)
  3. methylation of guanine base (guanine- 7methyl transferase)
  4. methylation of ribose (in some cases)
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9
Q

what is polyadenylation?

A
  • poly A tail found at 3’ end of mRNAs
  • protects mRNA from enzymatic degradation
  • aids in transcription termination, export of mRNA from the nucleus and translation
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10
Q

what are the stages in the 3’ polyadenylation of newly synthesised mRNAs?

A
  • recognition of AUAAA sequences by specificity componens RNA cleavage by cleavage factors
  • initial poly(A) polymerisation by poly(A) polymerase; followed by binding of poly(A) binding protein (PABP)
  • more poly(A) polymerisation and binding of more PABP
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11
Q

what are the general characteristics of tRNA molecules?

A
  • single RNA strand of approx 80 nucleotides
  • helps decode mRNA sequence into a protein
  • function at specific sites in the ribosome during translation
  • aminoacyl tRNA synthestase links amino acid to 3’ end of acceptor arm to produce an aminoacyl-tRNA
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12
Q

ribosome structure

A

large and small subunits
contain 50% proteinsand 50% rRNAs (by mass)

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13
Q

function of 80s ribosomes

A
  • small and large subunits bind together during initiation
  • translation takes place in the cavity between the two subunits
  • peptidyl transferase activity associated with the large 60s subunit
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14
Q

what are the three binding sites for tRNA in ribosomes?

A

E; exit site
P; peptidyl-RNA binding site
A; aminoacyl-tRNA binding site

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15
Q

other facts

A
  • translation must go fast enough to supply protein but slow enough to avoid too many errors
  • error rate; 1 in 10^4 incorrect aa
  • ribosomes add 20 aa/second to a polypeptide chain
  • proteinsynthesis is energetically expensive
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16
Q

CAP-dependent protein synthesis; initiation

A
  • small ribosomal subunit and initiatior tRNA bind, recognition of CAP, scanning to start AUG codon
  • ternanry complex binds large ribosomal unit
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17
Q

CAP-dependent proteinsynthesis; elongation

A
  • tRNA brings aa to the ribosome in order specified by codons on mRNA
  • ribosome catalyses peptide bond formation between amino group of each aa
  • more than one ribosome is active on any one mRNA (polysome)
18
Q

CAP-dependent proteinsynthesis; termination

A
  • at 3’ end of coding sequence the ribosome encounters a stop codon
  • polypeptide chain released
19
Q

initiation process

A
  • rate limiting step and tightly regulated
  • > 10 soluble eukaryotic initiation factors (eIF)
  • aim is to bring mRNA to ribosome and initiator tRNA to AUG start codon
20
Q

three main steps of initiation

A
  1. assembly of 43s pre-initiation complex
  2. binding of mRNA to 43s complex
  3. assembly of 80s initiation complex
21
Q

what is the structure of polypeptide chain eIF2

A

three subunits;
- GTP binding site; on y subunit
- phosphorylation site; found on a subunit, ser 51
- K boxes on b subunit, involved in interaction of eIF2B and eIF5

22
Q
  1. assembly of 43S pre-initiation complex
A

association met-RNA with eIF2/GTP (ternary complex)
40S trapped as monomer by eIF2 (43S)
binding of complexes to form 43S pre-initiation complex

23
Q
  1. binding of mRNA to 43S complex
A

eIF4E binds to CAP on mRNA
eIF4G binds to EIF4E
eIF4A unwinds mRNA secondary structure
43S preinitiation complex binds to eIF4G (via eIF3)- 48S preinitiation complex
scanning

24
Q

role of eIF4E

A

recognition of 5’ CAP on mRNA

25
Q

role of eIF4G

A

binds to eIF4E, eIF4A and eIF3

26
Q

role of eIF3

A

acts as a bridge between eIF4G and the 40s ribosomal subunit, hence required for mRNAs binding to the ribosome

27
Q

role of eIF4A

A

helps unwind secondary structure in 5’ end of the mRNA

28
Q

structure of eIF4G

A
  • has multiple domains for interaction with other initiation factors
  • acts as scaffold molecule that brings together other translation IFs
29
Q

interaction of 43S pre-initiation complex with 3’ end of the mRNA via poly(A) tail

A

interaction achieved by ability of poly(A) binding protein (PABP) to bind to both eIF4G and eIF4B that is part of 43S pre-in. complex
= circulatisation of the mRNA by assocation of the 5’ end with the 3’ end

30
Q

what happens after binding of 43S pre-in. comlex to CAP site?

A

scanning to the initiating AUG

31
Q

what is the effect of secondary structure in the 5’ UTR of an mRNA?

A

impedes the attachment of the 43S pre-initiation complex to the mRNA and/or reduces ability of the complex to scan along the mRNA

32
Q

IRES; internal ribosomal entry site

A

allow direct binding of the 43S complex to the mRNA without the need or CAP recognition or scanning
- IRES can replace function of some of the IFs
- some viral mRNA e.g. picornacirus use IRES mediated CAP-independent translation

33
Q
  1. assembly of the 80S initiation complex
A

40S/43S preinitiation complex scans to the AUG start codon
Met tRNA occupies the P site on the ribosome
60S subunit associated to form the 80S complex

34
Q

what does assembly of the 80S initiation complex involves?

A
  • binding of the 60S ribosomal subinit to the 43S mRNA complex
  • hydrolysis of the GTP bound to eIF2
  • release of most of the IFs from the ribosome
  • these events need eIF5 (has GTPase activity)
35
Q

stages in the formation of the first peptide bond and elongation

A
  • AA-tRNA binding; catalysed by eIF1, requires GTP
  • peptide bond formation; catalysed by the ribosome itself
  • translocation; catalysed by eEF2, requires GTP
36
Q

how is STOP codon recognised?

A

by release factor eRF
STOP codons;
- UAA ochre
- UAG amber
- UGA opal

37
Q

what happens at and after peptide formation?

A
  • hydrolysis of the last peptidyl-tRNA bond and release of the completed polypeptide chain
  • dissociation of the two ribosomal subunits from the mRNA
38
Q

what is a polysome?

A

structure that consists of multiple ribosomes attached to a single mRNA

39
Q

free ribosomes

A

synthesise soluble proteins that function in the cytosol

40
Q

ER bound ribosomes

A

synthesise proteins destined either for incorporation into cell membranes or for export from the cell

genes and gene expression (14 decks)

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