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BIOM 3500 - Midterm 2: Regulation of Gene Expression > Lecture 12 - Translation > Flashcards

Lecture 12 - Translation Flashcards

1
Q

What is translation?

A

The conversion of information in RNA into protein

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2
Q

What does the translation of mRNA into protein depend on?

A

Adapter molecules that can recognize and bind both to the codon and, at another site on their surface, to the amino acid

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3
Q

What do these adapters consist of?

A

A set of small RNA molecules known as transfer RNAs (tRNAs)

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4
Q

How is tRNA folded?

A

Four short segments of the folded tRNA are double-helical, producing a molecule that looks like a cloverleaf, which undergoes further folding to form a compact L-shaped structure that is held together by additional hydrogen bonds between different regions of the molecule

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5
Q

What part of the tRNA is crucial for its function in protein synthesis?

A

Two regions of unpaired nucleotides are situated at either end of the L-shaped molecule (one of these regions forms the anticodon and the other is a short single-stranded region at the 3’ end of the molecule where the amino acid matches the codon gets attached to the tRNA)

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6
Q

What is an anticodon?

A

A set of three consecutive nucleotides that pairs with the complementary codon in an mRNA molecule

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7
Q

What does the redundancy of the genetic cod imply?

A

There is more than one tRNA for many of the amino acids or some tRNAs can base-pair with more than one codon - both situations occur

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8
Q

Where is accurate base-pairing required?

A

Only at the first two positions of the codon

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9
Q

What explains why so many of the alternative codons for an amino acid differ only in their third nucleotide?

A

Wobble base-pairing in which the third codon position can tolerate a mismatch

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10
Q

How do tRNA molecules become linked to their appropriate amino acid partner?

A

Recognition and attachment depend on enzymes called aminoacyl-tRNA synthetases, which covalently couple each amino acid to its appropriate set of tRNA molecules

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11
Q

What does the synthetase-catalyzed reaction do?

A

Attaches the amino acid to the 3’ end of the tRNA which is coupled to the energy-releasing hydrolysis of ATP and produces a high-energy bond between tRNA and the amino acid which is used in a later stage in protein synthesis to link the amino acid covalently to the growing polypeptide chain

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12
Q

How do most synthetase enzymes select the correct amino acid?

A

By a two-step mechanism
- The correct amino acid has the highest affinity for the active site pocket of its synthetase and is therefore favoured over the other 19
- After the amino acid has been covalently linked to AMP: When tRNA binds, the synthetase tries to force the adenylated amino acid into a second editing pocket in the enzyme

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13
Q

What do most tRNA synthetase directly recognize?

A

The matching tRNA anticodon (synthetases contain 3 adjacent nucleotide-binding pockets, each of which is complementary in shape and charge to a nucleotide in the anticodon)

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14
Q

What is the fundamental reaction of protein synthesis?

A

The formation of a peptide bond between the carboxyl group at the end of a growing polypeptide chain and a free amino group on an incoming amino acid

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15
Q

How is a protein synthesized stepwise?

A

From its N-terminal end to its C-terminal end

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16
Q

Throughout the entire process, how does the growing carboxyl end of the polypeptide chain remain activated?

A

By its covalent attachment to a tRNA molecule (forming a peptidyl-tRNA)
- Each addition disrupts this high-energy covalent linkage, but immediately replaces it with an identical linkage on the most recently added amino acid

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17
Q

How is the synthesis of proteins guided?

A

By information carried by mRNA molecules

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18
Q

Where is protein synthesis performed? Why?

A

The ribosome - maintains the correct reading frame and ensures accuracy

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19
Q

Where are the large and small subunits of the ribosome assembled?

A

At the nucleolus, where newly transcribed and modified rRNAs associate with the ribosomal proteins that have been transported into the nucleus after their synthesis in the cytoplasm

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20
Q

When do the two ribosomal subunits join?

A

When they are exported to the cytoplasm

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21
Q

What does the small ribosomal subunit do?

A

Provides a framework on which the tRNAs are accurately matched to the codons of the mRNA

22
Q

What does the large ribosomal subunit do?

A

Catalyzes the formation of the peptide bonds that link the amino acids together into a polypeptide chain

23
Q

What happens once protein synthesis is initiated?

A

Two subunits of the ribosome join together on an mRNA molecule, usually near its 5’ end to initiate the synthesis of a protein
1) The mRNA is pulled through the ribosome, 3 nucleotides at a time
2) As its codons enter the core of the ribosome, the mRNA nucleotide sequence is translated into an amino acid sequence using the tRNAs as adaptors to add each amino acid in the correct sequence to the growing end of the polypeptide chain
3) When a stop codon is encountered, the ribosome releases the finished protein, and its two subunits separate again
4) These subunits can then be used to start the synthesis of another protein on another mRNA molecule

24
Q

How do the many coordinated movements required to allow for efficient translation?

A

A ribosome contains 4 binding sites for RNA molecules: one if for the mRNA and three (A, P, and E site) are for tRNAs

25
Q

When is a tRNA molecule held tightly at the A and P sites?

A

Only if its anticodon forms a base with a complimentary codon (allowing for wobble) on the mRNA molecule that is threaded through the ribosome

26
Q

What does the close proximity of the A and P sites allow for?

A

Maintains the correct reading frame on the mRNA because their 2 tRNA molecules can be forced to form base pairs with adjacent codons on the mRNA molecule

27
Q

What are the major steps for adding each new amino acid to the elongating chain once protein synthesis has been initiated?

A

1) tRNA binding - an aminoacyl-tRNA molecule binds to a vacant A site on the ribosome
2) Peptide bond formation
3) Large subunit translocation - leaves the 2 tRNAs in their hybrid site: P on the large subunit, A on the small subunit, and E on both
4) Small subunit translocation - carries its mRNA a distance of 3 nucleotides through the ribosome to “reset” the ribosome with a fully empty A site, ready for the next aminoacyl-tRNA molecule to bind

28
Q

What comes as the result of the two translocation steps?

A

The entire ribosome moves 3 nucleotides along the mRNA and is positioned to start the next cycle

29
Q

What do the elongation factors do?

A

Enter and leave the ribosome during each cycle, each hydrolyzing GTP to GDP and undergoing conformational changes in the process

30
Q

Why is GTP hydrolysis coupled with elongation factors?

A

To speed up protein synthesis and ensure that all changes occur in the “forward” direction, helping translation to proceed efficiently

31
Q

Why is the site at which protein synthesis begins on mRNA crucial?

A

Since it sets the reading frame for the whole length of the message

32
Q

Why is the initiation step of translation important?

A

Because for most genes, it is the last point at which the cell can decide whether the mRNA is to be translated to produce a protein

33
Q

What does the translation of an mRNA begin with?

A

The codon AUG, and a special tRNA (initiator tRNA)

34
Q

What does the initiator tRNA always carry? Why?

A

The amino acid methionine because it results in all newly made proteins having methionine at their N-terminus

35
Q

Why is the initiator tRNA specially recognized by initiation factors?

A

Because it has a nucleotide sequence distinct from that of the tRNA that normally carries methionine

36
Q

In eukaryotes, in addition to the initiator tRNA-methionine complex, what else is loaded into the small ribosomal subunits?

A

Additional proteins called eukaryotic initiation factors (eIFs)

37
Q

Of all the aminoacyl-tRNAs in the cell, what is the methionine-charged initiator tRNA capable of?

A

Tightly binding the small ribosome subunit without the complete ribosome being present and unlike other tRNAs, it binds directly to the P site

38
Q

How does the small ribosomal subunit then bind to the 5’ end of an mRNA molecule?

A

It is recognized by virtue of its 5’ cap that has previously bound two initiation factors, eIF4E and eIF4G

39
Q

What does the small ribosomal subunit move along the mRNA for?

A

To search for the first AUG from the 5’ to 3’ direction

40
Q

What facilitates the movement of the small ribosomal subunit?

A

Additional initiation factors that act as ATP-powered helicases

41
Q

At this point, why do the initiation factors dissociate?

A

To allow the large ribosomal subunit to assemble with the complex and complete the ribosome

42
Q

When is protein synthesis therefore ready to begin?

A

When the initiator tRNA remains at the P site, leaving the A site vacant

43
Q

What marks the end of translation?

A

Stop codons

44
Q

How do stop codons mark the end of translation?

A

Signaling to the ribosome to stop translation because they are not recognized by a tRNA and do not specify an amino acid

45
Q

What do release factor proteins do?

A

Bind to any ribosome with a stop codon positioned in the A site, forcing the peptidyl transferase in the ribosome to catalyze the addition of a water molecule instead of an amino acid to the peptidyl-tRNA, freeing the carboxyl end of the growing polypeptide chain from its attachment to a tRNA molecule

46
Q

Where is the completed protein chain immediately released to?

A

The cytoplasm

47
Q

What is one way to avoid translating broken mRNAs?

A

Having both the 5’ cap and the poly-A tail be recognized by the translation-initiation machinery before translation begins

48
Q

What does nonsense-mediated mRNA decay do?

A

Eliminates defective mRNAs before they move away from the nucleus

49
Q

When is the nonsense-mediated mRNA decay mechanism brought into play?

A

When the cell determines that an mRNA molecule has a nonsense (stop) codon in the “wrong” place - usually if mRNA has been improperly spliced

50
Q

How does the nonsense-mediated mRNA decay mechanism begin?

A
  • As its 5’ end emerges from a nuclear pore, the mRNA is met by a ribosome, which begins to translate it
  • As translation proceeds, the exon junction complexes (EJCs) that are bound to the mRNA at each splice site are displaced by the moving ribosome
  • By the time the ribosome reaches the normal stop codon within the last exon, it will stall and no more EJCs will be bound to the mRNA
  • mRNA “passes inspection” and is released into the cytosol
51
Q

What happens if the ribosome reaches a stop codon earlier when the EJCs remain bound?

A

The mRNA molecule is rapidly degraded

52
Q

What gene is involved in nonsense-mediated decay?

A

unc54-r293 allele

BIOM 3500 - Midterm 2: Regulation of Gene Expression (4 decks)

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