Eukaryotic Translation Flashcards
___% of a mammalian cell’s energy production is used for the process of protein translation
30%
major stages of protein translation
- load amino acids onto tRNAs
- initiation
- elongation
- termination
key design principles of protein translation
- load ribosome onto mRNA at right location = correct start site
- prevent ribosome form starting before fully assembled = placeholder proteins
- need to move along mRNA to continue translating = use proteins like ratchets and physically push mRNA and new peptides through ribosome
- when translation is done, make sure all components get taken apart FULLY to start next round = peptide release, tRNA release, ribosome subunits
- use similar shaped components when possible = tRNAs, elongation & termination factors share similar structures because they all need to fit well in the A-site
Prokaryotic translation
- transcription & translation simultaneous
- mRNAs unstable = few secs to mins
- cap-independent initiation
- 3 initiation factors = IF1, IF2, and IF3
- polycistronic = can code for more than one protein per mRNA
eukaryotic translation
- eukaryotic transcription &trnalsation are discontinuous processes
- mRNAs = quite stable = few hrs to days
- cap-dependent and cap-independent initiation
- 9 initiation factors are involved = eIF, 1, 2, 3, 4A, 4B, 4C, 4D, 5 & 6
- monocistronic (& polyc??)
4 complexes of initiation
- 43S pre-initiation complex (43S PIC) = complex ready to accept mRNA
- eIF4-mRNA complex = prepping mRNA so positioned properly for when it bonds to 43S PIC
- 48S initiation complex = ribosome binding to mRNA and scanning for start codon
- 80S elongation complex = bringing 60S to start translation
step 1 of translation initiation (prepping ribosome)
binding of ______ and _____ causes conformational changes that opens up channel allowing mRNA access
eIF1 (E) and eIFA (A)
- blocks A and E site access by tRNAs
eIF2
- ternary complex
- essential for delivery of initiator Met-tRNA to 40S
- eIF2B is a guanine exchange factor required to make eIF2 competent for reloading with Met-tRNAi^Met
the only tRNA that binds directly to P-site of ribosome
Met-tRNAi^Met
largest initiation factor with 13 subunits
eIF3
- forms many contacts that stabilizes the 43S PIC
> multiple initiation factors and the ribosome subunits
> mRNA and the 5’-m7G cap
> subunits near the mRNA entry and exit tunnel
analogous to a double-sided tape
eIF3
= seals entry and exit + protects ribosomes from starting initiation and physically blocks to stop other factors from coming in
eIF5
- GTPase activating protein (stimulates GTP hydrolysis of eIF2y
- eIF5 activity repressed by eIF1 in 43S PIC & 48S initiation complex
> to prevent premature hydrolysis of GTP
initiation factors in 43S complex
eIF1
- with eIF1A induces confirmation of 40S
- binds near E-site, close enough to P site; AUG codon recognition
eIF1A
- with eIF1 = confirmation of 40S
- binds to A site; 60S subunit assembly, and inhibits premature elongation
eIF2
- delivery Met-tRNA^Met to P-site of 40S
eIF3
- large scaffolding protein complex
eIF5
- stimulates GTPase activity of eIF2
what is the purpose of the 5’ cap on mRNA?
- to regulate nuclear export of mature mRNAs
- to inhibit degradation of mRNA by exonucleases
- to assist in the assembly of an mRNA-ribosome complex during translation initiation
eIF4
eIF4A
- RNA helicase
- unwinds the mRNA hairpin
eIF4E
- cap-binding protein
- recognize cap on mRNA
- differentiates between bacterial and human
eIF4G
- scaffold
- binds to eIF3
- binds to polyA binding protein (PABP)
NOTE: eIF4B enhances helicase activity of eIF4A
Low levels of tRNAi reduces…
Translation
Leaky scanning
When a weak Kozak consensus cause the 48 S to scan past an AUG to next site with higher frequency
eIF5B
GTPase that mediates joining of 40S and 60S
What happens upon 60S joining 40S?
40S bound eIFs leave EXCEPT eIF1A as it provides dock for eIF5B
What happens at start of elongation?
60S stimulated GTP hydrolysis of of eIF5B
eIF5B and eIF1A both dissociate
3 important steps in translation elongation
- eEF1 loads aminoachl-tRNA to A site
- Peptidyl transfer rxn = extends polypeptide chain
- Translocation
Translation termination important points
- eRF1 looks like tRNA but instead of adding another AA, cuts polypeptide chain off
- ABCE1 breaks apart subunits and prevents them from coming back together
eEF1 keeps trying but nothing happening because stop codon is unrecognized
Ribosome recycling
ABCE1 splits ribosome in 2 so that the subunits can be recycled for next round of translation
Tetracycline
More specifically inhibits prokaryote translation
Binds to small subunit and blocks tRNAs from enteringA site
