What are 3 critical RNAs in protein synthesis?
- tRNA
- rRNA (80% of total cellular RNA)
- mRNA
How are pre-rRNA transcription units arranged?
In repetitive clusters, creating a membraneless nucleolus, this is where rRNA is made
What are th differences between bacterial ribosome and eukaryotic ribosome?
- In bacteria, 23S,16S and 50S give rise to two structires, coming together to form a complete 70S ribosome
- In eukaryotes, 60S and 40S come together to make 80S ribosome, eukaryotes have an additional RNA in it’s large subunit
Explain rRNA structure and how it is highly conserved?
- Folds into conserved stem-loops
we see that is in eukaryotes and prokaryotes, indicating how this structure is critical
Explain the processing of pre-tRNAs and how they “prepare” for translation
1) 5’ sequence is removed
2) Section of sequene is removed form loop
3) CCA is added to 3’ end
4) Modification of internal bases
The anticodon is critical for decoding of mRNA
What are aminoacyl-tRNA synthetases
Bind/charge tRNA to correct amino acid
Bind 2 substrates by froming ester bond between carboxy on a.a. and hydroxyl on tRNA nucleotide
What does it mean for codes to be “degenerate”?
- Amino acid can bind to more than one unique tRNA
- A single tRNA can bind to more than one single codon
- tRNAs will be bound to their amino acid and charged to be used in protein synthesis
Several codes can result in same thing
What are the two type of tRNAmet?
- Initiator tRNAmet (tRNAimet) - ONLY for initiation of polypeptide chain
- Normal tRNAmet - used whenever methionine comes up
How do we distinguish the two types of tRNAmet?
- In prokaryotes, tRNAimet has a formyl group to show it’s an initiatior
- In eukaryotes, tRNAimet is structurally different - binds to P site specifically
Explain the assembly of the pre-initiation complex in translation - what is the name of the complex created?
- EIF2 aids in binding of GTP with tRNAmet
- Complex binds to small ribosomal subunit, also bound by other initiation factors (creates 43S pre-initiation complex)
How does tRNAimet and EIF2 react to bad conditions?
- Bad conditions cause phosphorylation that blocks EIF2 from tRNAmet, blocking synthesis
What do we need for efficient translational initiation of mRNA?
Need a 5’cap - enhances synthesis
- specifically, a 7mGDPcap is related to increased translational efficiency, but no correlation with uncapped transcripts
What does eIF4 in general do?
- Interacts with cap, bringing down whole complex to 5’ end of an mRNA to be translated
- too much eIF4 causes protein overexpression, which is associated with tumour formation
- eIF4g brings small ribosomal subunit to where it needs to work, and forms a loop (pre-initiation complex +eIF4 with mRNA)
What does eIF4B do?
Works to enhance eIF4 activity
What does eIF4A do?
Works as an RNA helicase
What does eIF4E do?
Binds the 5’ cap of mRNA
What does eIF4G do?
binds PAPBC and eIF3
What is the scanning complex - what does it do?
eIFs + 40S and 80S ribosomes, and tRNA (including tRNAimet) make up the structure
It moves along mRNA, and first stops when it reaches the start codon
When happens when we get to the start codon?
- Instantaneously - we get the 48S initiation complex (as soon as scanning complex hits AUG)
- This changes the conformational change that permits association of large subunit
- All other initiation factors are released - 80S ribosome
- tRNAimet stays interacting with AUG in the Psite, leaving the A site vacant
