RNA translation Flashcards
What are the stop codons?
UAA, UAG, UGA
When does tRNA splicing occur?
Happens after transcription, not during
What are the unusual nucleotides in tRNA?
deamination of A (inosine), sulfur-replaces-uracil oxygen (4-thiouridine), two methyls added to G (N,N-dimethyl G), hydrogenation of U (dihydro U)
What end of tRNA are amino acids bound to?
3’ end. Attached to acceptor stem.
How do amino acids get on tRNA’s?
Aminoacyl synthetases add them
How many aminoacyl synthetases are there?
at least 20 of them, one for each amino acid - but slightly more
What part of the amino acid is bound to tRNA’s?
Where the amino bond will be (is the R group of an ester)
What orientation is mRNA vs anticodon?
tRNA is 3’-5’ on top
mRNA is 5’-3’ on bottom
What is the wobble position?
The last (3’ end of mRNA, 5’ end of tRNA) position on codons can change. That’s why multiple codons can code for the same amino acid
What are the bases that wobble?
U: A, G, or I
C: G or I.
A: U
G: C
Only U and C can wobble
Methionine codon
AUG
What happens right before aminoacyl synthetase binds to amino acids?
Amino acid gets adenylated with ATP>ADP
What’s tRNA with amino acid on it called?
aminoacyl tRNA
What is ‘charging’ tRNA?
Adding the amino acid to it
How can aminoacyl synthetase enzyme fix mistakes?
There’s an editing site inside aminoacyl synthetase. Correct amino acid has high affinity for active site pocket; incorrect amino acid gets removed at editing site.
How are incorrect amino acids cleaved from aminoacyl synthetase?
hydrolysis
What’s the error rate of aminoacyl synthetase couplings?
1/40,000
What terminus of do peptidyl-tRNAs add to?
N-terminus: the peptidyl-tRNA’s are already on the growing polypeptide chain
How is a new amino acid added to a growing chain?
N-terminus of new aminoacyl-tRNA adds to C-terminus of growing chain
What remains on tRNA molecules after they drop off amino acids (at location where amino acids were)?
an OH group
What are ribosomes composed of?
rRNA, ribosomal proteins
Is the difference between euk and prok ribosomes qualitative or quantitative?
quantitative
What are the three sites on the large ribosomal subunit?
E site, P site, A site
Where does aminoacyl tRNA bind?
A site
What does the A site do?
Accept aminoacyl-tRNA
What does the P site do?
It’s just occupied by peptidyl-tRNA (growing polypeptide chain)
What does the E site do?
It’s the exit site
Are the sites on large subunit, small, or both?
They’re on both, but there’s a translocation that happens
What happens after A-site binding?
New peptide bond is formed
What happens after a new peptide bond is formed in translation?
Large subunit of ribosome translocates to next codon
What happens after large subunit translocation?
Small subunit translocation, and now the A site is empty
How many nucleotides can each site hold?
3 (a full codon)
How many nucleotides are the large and small subunit on total?
9 (three for each site)
What happens in translation cycles besides amino acid growth?
Proofreading
What direction is the new polypeptide chain synthesized?
N-terminus to C-terminus
How is energy to initiate protein translation acquired?
ATP and GTP used.
What is the initiator RNA and what’s it do?
methionine aminoacyl tRNA, looks around trying to find first AUG
What happens after initiator RNA binds?
Initiation factors bind
What is used for energy at each amino acid addition?
GTP
What binds when the stop codon appears?
The release factor
What happens after release factor binds?
Everything dissociates, can be reused
What happens to mRNA after it’s translated?
Stays in nucleus to get translated again or degraded
What’s the ratio of mRNA to protein it makes?
High - many ribosomes can translate mRNA at once
Why, when, and how are amino acids cleaved from aminoacyl synthetases?
If the wrong amino acid binds. Cleaved after it binds. Amino acid is hydrolytically cleaved in editing pocket.
What happens after initiator tRNA binds to P site?
Initiation factors come in with mRNA
What happens after mRNA first gets to the small ribosomal subunit with initiator tRNA bound to its P site?
Initiator tRNA moves around to find the first AUG site
What happens after initiator tRNA finds first AUG site?
Other protein factors dissociate, large subunit binds, first peptide bond forms. Initiator tRNA starts at P site.
Can the correct amino acid be put into the editing side of tRNA synthetase? How does this proofreading work?
No. The proofreading works by trying to shove the amino acid into the editing pocket; it can’t go in if it’s the right amino acid.
How does tRNA synthetase recognize the correct tRNA? What does this mean the binding point of the synthetase is similar to? Do all synthetases do this?
The synthetase binds to the tRNA codon. This means the synthetase has the same amino acid sequence as the mRNA sequence the tRNA will bind to. Some synthetases bind to the acceptor stem instead.
What is at the C-terminal end of a polypeptide chain as it’s growing? Where on the ribosome is this?
a peptidyl-tRNA in the P-site of the ribosomes.
How accurate is translation?
1 mistake per 10,000 amino acids
What is the role of the small subunit?
Provides the framework on which tRNA’s are matched to codons
What is the role of the large subunit?
Catalyze formation of peptide bonds in growing chains
How fast do eukaryotic ribosomes add amino acids?
2 amino acids per second
How fast do prokaryotic ribosomes add amino acids?
20 amino acids per second
Why does the large subunit translocate first?
It catalyzes formation of peptide bonds, and it can move as soon as it catalyzes a bond.
What does a peptidyl transferase do? Where is it located?
It’s the main enzyme that catalyzes peptide bonding. It’s located in the large subunit.
What site is the emerging polypeptide chain coming out from?
The P site.
When does a spent tRNA get ejected from the ribosomal complex?
As the small subunit moves forward to the next codon.
How does GTP/GDP factor into the ribosomal polypeptide growth cycle?
Two elongation factors come in to speed up the process with GTP>GDP.
What provides energy to power polypeptide growth?
Elongation factors hydrolizing GTP, and breaking the high-energy tRNA-amino acid bond.
What do elongation factors do besides speed up synthesis?
Increase accuracy
What do release factors do?
They terminate translation of a protein.
When can translation stop? What terminates it?
Either when the ribosome hits a stop codon, or when there was too much amino acid misincorporation (too many errors). Release factors terminate.
What amino acid is on the N-terminus of all newly made amino acids? What happens to it?
Methionine. It’s removed later by a protease.
What’s the first thing that binds to the small subunit?
initiator tRNA with methionine.
Why is leaky scanning important and what does it involve?
Sometimes initiation tRNA’s skip the first start site (AUG). This lets more types of proteins get made, it’s another level of alternative splicing
Where do stop codons have to be positioned for release factors to bind?
The A site
What gets added to the C-terminus of the last peptidyl-tRNA in a polypeptide chain, and what adds it? What happens to the polypeptide chain?
Water instead of a new amino acid, peptidyl transferase adds it. This releases the complete polypeptide into the cytoplasm.
What causes peptidyl transferase to add water to the last amino acid of a peptide chain?
Release factors that bind to the ribosome and change peptidyl transferase conformation.
What does ADEPT stand for?
Analogy, Diagram, Example, Plain-english, Technical
