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Translation Flashcards

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1
Q

What occurs in the translation process?

A

The ribosomes read the genetic code of DNA that has been rewritten in the form of mRNA and assembles proteins according to DNA instructions?

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2
Q

How are t RNAs involved in the translation process?

A

They serve as adaptors that allow specific amino acids to interact with mRNA

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3
Q

What do the ends of the tRNA bind to?

A

One end binds to the amino acid and the other end interacts with the mRNA

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4
Q

What three stages is translation broken into?

A

initiation
elongation
termination

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5
Q

What is initiation in translation?

A

binding of mRNA to ribosome and binding of the first tRNA with amino acid

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6
Q

What is the function of elongation in translation?

A

Extension of the polypeptide

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7
Q

What is the termination of translation?

A

release of the complete polypeptide and the mRNA

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8
Q

What are the differences in translation between eukaryotes and prokaryotes?

A

initiation in eukaryotes is more complex than prokaryotes

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9
Q

What are the two initiation events that must occur in Prokaryotes?

A
  1. tRNA is charged to form the amino-acyl tRNA

2. dissociation of the 70s ribosome into the large 50s and 30s subunit

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10
Q

Where is a tRNA charged?

A

when it is covalently bound to an amino acid

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11
Q

All tRNAs have the same base sequence on the what end? and what is the base sequence?

A

3’ end

CCA

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12
Q

What is the structure of the charged tRNA look like?

A

see page

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13
Q

What type of bond joins the carboxyl terminus of the amino acid to the 2’ or 3’ hydroxyl group of the terminal A of the tRNA.

A

ester bond

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14
Q

What is the enzyme that catalyzes the 2 reactions needed for tRNA charging?

A

Aminoacyl-tRNA synthetase

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15
Q

WHat is the one of the reaction required for tRNA charging?

A

The amino acid is activated by coupling it with AMP to form aminoacyl-AMP

amino acid + ATP —> aminoacyl-AMP + pyrophosphate PPi

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16
Q

How does the first reaction in tRNA charging activate aminoacyl AMP?

A

The energy releases from ATP hydrolysis is trapped in the amino acyl AMP thus activating it

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17
Q

What is the second reaction required for tRNA charging?

A

The stored energy is used to transfer the amino acid to a tRNA thus forming aminoacyl-tRNA

aminoacyl-AMP + tRNA—> aminoacyl-tRNA + AMP

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18
Q

WHat is the summary reaction for the 2 reactions required to charge tRNA?

A

aa + ATP + tRNA —> aminoacyl-tRNA+AMP+PPi

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19
Q

How many aminoacyl-synthetases are there? WHy?

A

20

One for each amino acid

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20
Q

Each synthetase has a high degree of what?

A

specificity

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21
Q

When attaching the correct amino acid to its partner tRNA how many errors are generally made?

A

very few

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22
Q

What was the first preinitiation step in prokaryotes and what is the second?

A
  1. formation of aminoacyl tRNA

2. dissociation of ribosome

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23
Q

Why is the 2nd preinitiation step required in prokaryotes and eukaryotes?

A

translation initiation complex is built on the small ribosomal subunit

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24
Q

WHat are the three initiation factors in E coli?

A

IF-1
IF-2
IF-3

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25
WHich three initiation factors of E coli participate in ribosomal dissociation?
IF-1 | IF-3
26
What is the function of IF-1 of Ecoli?
promotes the dissociation of the intact 70s ribosome
27
What is the function of IF-3 of Ecoli?
Binds to the 30s subunit to prevent its reassociation with the 50s subunit
28
What is ribosome cycling?
The process when each intact ribosome dissociates into its subunits at the end of each round of translation
29
What is the 30s initiation complex?
It is the initiation complex for translation. It is formed on the 30s subunit and contains mRNA, aminoacyl-tRNA, and initiation factors
30
WHat is the order of the IF binding to the initiation complex?
IF3---> IF2---> IF1
31
Can IF-1 bind to the 30s subunit by itself?
No
32
What does the presence of all 3 IFs on the initiation complex allow to occur?
It allows the mRNA and the first aminoacyl tRNA binding to occur
33
WHat is almost always the first codon?
AUG
34
What happens when AUG is in the interior of the message what does it code for?
methionine
35
WHen AUG appears at the beginning of the message what does it code for?
The initiation/start codon
36
What des an initiating AUG signal the binding of?
a special aminoacyl-tRNA that contain N-formyl-methionine (fMet-tRNA or tRNA^fMet)
37
WHat does fMet-tRNA stand for?
aminoacyl tRNA that contains an N-formyl-methionine
38
What is the structure for methionine compared to the structure of N-formyl-methionine?
see sheet
39
What are other initiating codons?
GUG and UUG
40
What initiating codon occurs 90% of the time in Ecoli?
AUG
41
In any event what is the first amino acid that is incorporated into a polypeptide? And what happens during protein maturation?
- fMet | - the amino acid is typically removed during protein synthesis
42
What does GUG code for?
valine
43
What does UUG code for?
leucine
44
How is an initiation codon recognized by the cell(ribosome) versus an ordinary codon?
The mRNA contains a conserved sequence that is located upstream from the start codon that is called the Shine-Dalgarno sequence and it base pairs with a complementary base sequence located on the 3'end of the 30s ribosomal subunit rRNA
45
What other factors does GTP bind to?
fMet-tRNA mRNA initiation factors 30s ribosomal subunit
46
How many molecules of GTP is bound per mole of fMet-tRNA?
one molecule of GTP
47
When is this GTP hydrolyzed?
GTP is hydrolyzed when the 50s subunit links with the 30s complex to form the complete intact ribosome and the IF2 has dissociated from the 30s subunit ribosomal subunit
48
What does the formation of the complete 70s ribosome require?
the 50s and the 30s ribosomal subunits to join together
49
WHat two initiation factors are dissociate from the complex?
IF-1 and IF3
50
GTP hydrolysis drives the release of what initiation factor?
IF2
51
What two things form the GTPase?
IF-2 and the 70s complex
52
What are the general steps in the formation of the 30s initiation complex in prokaryotes?
1. Binding of all three initiation factors to the 30s subunit 2. Once bound the initiation factors direct the binding of the mRNA and fMet-tRNA 3. the 30s initiation complex consists of the 30s ribosomal subunit+mRNA+fMET-tRNA+GTP + IF-1 + IF-2 + IF-3
53
What are the steps in the formation of the 70s initiation complex in prokaryotes?
1. 50s ribosomal subunit binds to the 30s initiation complex 2. Dissociation of the IF-1 and IF-3 3. Hydrolysis of GTP on the 30s subunit 4. The hydrolysis of GTP drives the release of IF-2 5. 70s initiation complex is formed ready to elongation
54
What are the two major difference in eukaryotic initiation of translation compared to the prokaryotic initiation of translation?
1. initiation in eukaryotes uses methionine rather than N-formyl methionine 2. There is no Shine Delgarno sequence to guide the ribosomes tot he translation start site. Instead the 5'-cap directs the initiation factors to bind and search for an initiation codon.
55
Prokaryotic mRNA is polycistronic what does this mean?
They contain information for multiple genes. Each cistron has its own initiation codon and ribosome binding site.
56
How prevalent is polycistronic mRNA in eukaryotes?
Not very common...rare
57
How does the eukaryotic ribosome locate the AUG start site?
The eukaryotic ribosome will locate the 5'-end of the message and will scan downstream along mRNA until the first AUG is located. The AUG must be in the correct sequence context to be considered the start site.
58
WHat are some correct sequence context that AUG will be recognized by the eukaryotic ribosome as the start site?
1. A purine at the -3 2. A G at +4 3. The "A" in the AUG is at position +1
59
Why do eukaryotic require more initiation factors than the bacteria?
The bacteria have a very direct way to find the start codon using the Shine Delgarno sequence. The more indirect method to find the AUG start codon in EUkaryotes requires more factors 12 to be exact.
60
What are the steps in the eukaryotic initiation of translation?
1. eIF-3 binds to the 40s subunit and eIF-6 binds to the 60s subunit to prevent premature 40s and 60s reassociation 2. eIF2 helps bind the 40s ribosomal subunit to the aminoacyl-tRNA (Met-tRNA) 3. eIF-4 allows the 40s subunit to bind to the 5'end of the mRNA 4. eIF-1 and eIF-1A stimulate the scanning of the mRNA for the initiation codon 5. eIF-5 stimulates the binding of the 60s ribosomal subunit
61
What is eIF-3 and its function?
eukaryotic initiation factor 3 and it binds to the small 40s ribosomal subunit to inhibit its reassociation with the large 60 ribosomal subunit
62
WHat is the function of eIF-2?
Its function is to bind the initiating aminoacyl-tRNA to the ribosome and is similar to the prokaryotic IF-2
63
WHat does eIF-2 require to function and how is this supplied to it?
It requires GTP. GTP is hydrolyzed to GDP when the factor dissociates from the ribosome. For the factor to function again GTP must replace the GDP and this is accomplished by eIF2B
64
What is the function of eIF-4?
It is a cap binding proteins that allows the 40s ribosomal subunit to bind tot he 5'-end of the mRNA
65
WHat is the function of eIF-1 and eIF-1A?
They are required for the ribosome to scan mRNA for the initiation codon
66
What is the function of eIF-5?
Stimulates the association of the 60s subunit
67
What is the function of eIF-6?
It is a second anti-association factor that prevents the premature association of the 40s ribosomal subunit and the 60s ribosomal subunit
68
What is elongation of a polypeptide in translation?
It occurs as a three step process that is repeated over and over until the polypeptide being synthesized is complete
69
When elongation starts in prokaryotes what is the initial structure?
mRNA and fMet-tRNA are bound to the intact 70s ribosome
70
What are the three binding sites for the aminoacyl-tRNA on the ribosome?
1. P(peptidyl) site 2. A (aminoacyl) site 3. E (exit) site
71
Out of the three binding sites on the ribosome where is the fMet-tRNA bound?
P site
72
Additional amino acids are added in elongation according to what? And what type of bond is formed between each amino acid?
Amino acids are added by the sequence dictated by the codon sequence on the mRNA. Peptide bonds are formed between each amino acid.
73
What is the first step in elongation in translation?
fMet-tRNA(AUG start codon) occupies the P-site, the second codon and its "matching" tRNA occupy the A-site
74
What is the binding of the aminoacyl-tRNA dependent on?
The EL-TU and GTP
75
What is the second step in the elongation process?
The large ribosomal subunit acts as peptidyl transferase, which moves fMet from its tRNA in the P-site to the aminoacyl-tRNA in the A-site. A peptide bond forms between the fMEt and the second amino acid that is in the A site (which is still attached to its tRNA).
76
What is peptidyl transferase?
a ribosome not a protein
77
WHat is the third step in elongation?
Translocation. The mRNA shifts position in the ribosome so that the dipeptidyl-tRNA in the A site is now in the Psite and the deacylated tRNA in the Psite is now displaced and leaves the ribosome. The next mRNA codon moves into the Asite and the process repeats.
78
What is translocation dependent on?
EF-G and GTP
79
WHen does the elongation process stop?
WHen it reaches a STOP codon in the message
80
What is the error rate per amino acid added in elongation?
0.001%
81
WHy is there such a low error rate in elongation?
proofreading process
82
How is the proofreading process accomplished?
The proofreading relies on the weakness of incorrect codon-anticodon base pairing to ensure dissociation of the incorrect tRNA will occur faster than when GTP hydrolysis.
83
WHat accounts for the faster timing of the dissociation of the incorrect tRNA?
differences in reaction rates.
84
WHen does termination begin?
WHen the ribosome encounters a STOP codon.
85
What are the three nonsense codons that appear in the genetic code as STOP codons?
UAG UAA UGA
86
Does tRNA recognize the STOP codon?
no
87
WHat recognized the STOP codons?
release factor(RF) proteins
88
What is RF-1 specific for?
UAA and UAG
89
What is RF-2 specific for?
UAA and UGA
90
What is the function of RF-3?
It is a GTP-binding protein that promotes the binding of RF-1 and RF-2 to the ribosome
91
What do the release factors as a whole do?
induce the release of the completed polypeptide from the ribosome.
92
How do the RFs bind to the stop codon?
They have specific amino acids that will bind to the stop codon.
93
What is the tripeptide sequence RF-1 contains? And what parts of the tripeptide sequence bind where?
Pro-Ala-Thr Pro binds to the A of UAG Thr binds to the G of UAG
94
WHat is the tripeptide sequence of RF-2? And what parts of the tripeptide sequence bind tot he amino acid?
Ser-Pro-Phe Ser binds to the G of UGA Phe binds to the A of UGA
95
RF-1 and RF-2 both can bind to what stop codon?
UAA
96
What is the release factor in eukaryotes?
eRF1 which can bind to all three STOP codons

Molecular Final (4 decks)

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