lec28 Flashcards
where does the aa attach on trna
-3’ end
modifications on the trna
-D loop, anticodon loop, T loop
-modified bases T, D, Y
-happens right afetr transcription
-modifications To stabilize tRNA’s 3d structure, improve codon-anticodon pairing, help recognition by trna synthetase
What enzyme charges tRNA and how?
Aminoacyl-tRNA synthetase:
Binds amino acid + ATP → aminoacyl-AMP
Binds correct tRNA
Transfers amino acid to 3’ A of CCA tail
→ Forms high-energy aminoacyl-tRNA (charged tRNA)
ATP → AMP + PPi (2 phosphates lost)
Synthetase ensures correct aa for tRNA → codon indirectly selects aa via anticodon.
What binds in the active site of aminoacyl-tRNA synthetase?
Both ATP and the amino acid fit into a deep cleft (active site pocket) of the enzyme. This ensures correct positioning for forming aminoacyl-AMP and attaching the aa to the tRNA.
the tRNA and the mRNA
interactions are antiparallel. t or f
true
mrna moves 5’ to 3’
trna binds antiprallel so its 3 to 5
stop codons
There are 3 stop codons, UAA, UAG, and UGA.
These codons signal the end of a gene. They
terminate protein synthesis. They do not
encode an amino acid.
orf
ORF = OPEN READING FRAME, i.e. a long sequence
of nucleotide triplets with no stop codonsfr
point vs frameshift mutation
frameshift is when one base is added or deleted. changes reading frame. sometimes it can cause a stop codon somewhere in the middle whihc can result in a truncated protein that will not fold and likely degrade
point is when one base is swapped
first, second nad third reading frame
second is skipping one base and then reading a codon, third is skipping two bases and then reading codon
peptide formation:
- inititation
- elongation
- termination
- ribosome recycling
What do reconstitution experiments tell us about ribosomes?
Ribosomal RNA and proteins can be purified and reassembled into functional ribosomes.
This shows:
All essential components are known
Ribosomes can self-assemble
Components from different organisms can sometimes be mixed and still form active ribosomes
prokaryotic vs eukaryotic ribosomes
prok: 30s+50s=70s
euk: 40s+60s=80s
What makes up most of the ribosome and what does it do?
rRNA makes up ~2/3 of the ribosome’s mass.
It forms the ribosome’s shape, helps it bind mRNA, and carries out the catalytic activity (like making peptide bonds).
Ribosomal proteins are smaller and attach to specific rRNA regions.
What is the Shine-Dalgarno sequence?
A sequence on prokaryotic mRNA (e.g., AGGAGG) just before the start codon.
It base-pairs with the 16S rRNA of the 30S ribosome to position the start codon in the P site.
Helps the ribosome start translation in the right place.
Formation of the prokaryotic ribosome initation complex:
Formation of the 70S prokaryotic ribosome:
30S small subunit binds to IF1, IF2 (with GTP), IF3
It also binds charged initiator tRNA (fMet-tRNAᵢ^Met)
mRNA binds — Shine-Dalgarno sequence pairs with 16S rRNA to line up the start codon
This triggers release of IF3 (so 50S can join)
GTP hydrolysis (by IF2) helps the 50S large subunit attach
IF1 and IF2 are released
🎯 Now you have the full 70S initiation complex:
P site has the charged tRNA
A and E sites are empty
16s rRNA
Prokaryotic 16S rRNA (inside the small
ribosomal subunit, 30S) forms base pairs
with the mRNA containing the
Shine–Dalgarno sequence (on the mRNA)
usually UGGAG, AGGAG, AAGGG, GGAG etc
Ester Bond Formation (tRNA charging)
The 3′-OH of the tRNA attacks the carboxyl group (–COOH) of the amino acid. This condensation reaction releases water (H₂O) and forms an ester bond:
tRNA–O–C(=O)–AA
The bond links the amino acid to the tRNA’s 3′ end, creating a charged tRNA.
Peptidyl Transferase Reaction
The A-site amino acid’s NH₂ attacks the P-site carbonyl C, forming an unstable tetrahedral intermediate that collapses into a peptide bond. Elongation uses 2 GTP per cycle — one for bringing in the charged tRNA, and one for ribosome translocation.
ribosome transocation
Ribosome translocation is the step where the ribosome shifts one codon (3 bases) along the mRNA (5′ → 3′ direction) after a peptide bond is formed.
🔄 It moves:
The A-site tRNA → P-site
The P-site tRNA → E-site (and exits)
The next codon enters the A-site
🧪 This step uses 1 GTP, hydrolyzed by elongation factor EF-G in prokaryotes (or eEF2 in eukaryotes).
2 gtp used in elongation
one to promote binding of charged trna (AA-tRNA^AA) to A site and one to faciliatate translocation of ribosome in 3’ direction
how does tetracycline work
Tetracycline antibiotic binds 16S rRNA on
the 30S (small) subunit of the bacterial ribosome,
blocking the tRNA from entering the ribosome
which way does protien synthesis
n->c terminal.
exit of new polypeptide frm ribosome (prok)
hydrphobic tunnel in the 50s sub at the site of peptide bond formaion
termination of translation
A stop codon enters the A-site → Release factor (e.g., RF2) binds → GTP is hydrolyzed → Polypeptide is released from the P-site tRNA → Ribosome, mRNA, and tRNAs disassemble.
