Protein Synthesis Flashcards
What is the ribosome composed of?
Large subunit- catalyses the formation of the peptide bond Small subunit that matches the tRNA anticodon to the codons of the mRNA
What is the large subunit made of?
60S subunits have three RNAs 5S, 5.8S and 28S (120, 160 4700 nucleotides) and ~49 proteins
What is the small subunit made of?
18S rRNA (1900 nucleotides) and ~33 proteins
What does the S of the subunits mean?
Stands for Svedberg- a coefficient of sedimentation relating to the rate of sedimentation of the particle and is not additive
What end do the codons fit onto the tRNA?
3’ prime end
Decribe the role of EIF4 and 5
eIF4 and 5 these recognise and bind to the cap structure
Deecribe the initiation process(5)
- The cap binding protein associates with the cap, the poly A binding proteins associate with the poly A tail- This causes circularisation of the RNA
- Initiation factors assemble on the small subunit and have a GTP bound to it
- This binds to a special met-tRNA, this initiation tRNA (tRNAi) is the only one that can bind to the P(peptidyl)
- The small subunit binds to the 5’ cap of the mRNA but the start codon can be a lot further down so the ribosome needs to move along until it finds the AUG start codon which has to be near the Kozak sequence which is ACCATGG. Scanning uses ATP
- Scanning stops once the start codon has been found
How is the process of scanning involved in initiation?(2)
- Scanning is the process whereby the ribosome tries to find the start codon of the mRNA. This involves the consumption of energy via the hydrolysis of ATP.
- If the ribosome is 100 bases away from the start codon 100 molecules of ATP will be used up.
When does the large subunit bind to the mRNA?
After the subunit has attached to the mRNA
Describe the translation/ elongation process(6)
- EIF3 and eiF2 gets released after hydrolysing GTP which allows the 60S ribosomal subunit to join the 40S preinitiation complex which forms the 80S ribosome around the RNA.
- Met-tRNAi is associated with P site and a second charged tRNA enters the A site with the first of the elongation factors EF1.
- when the covalent bond forms between methionine and the next amino acid, peptidyl transferase moves the amino acid in the P site to the A site
- the tRNA on the P site is now empty
- the tRNA in the A site has both amino acids attached to it.
- P site tRNA now moves to the E site so it can go and get recharged so a new tRNA can enter the A site.
Viruses can inactivate EIF3 and EIF4 so antiviral proteins cannot be synthesized.
What must happen to the tRNA for it to be reused after elongation?
The tRNA which has left the E site it must be recharged by aminoacyl tRNA synthetase which requires ATP, producing AMP and pyrophosphate.
What is a polysome?
In most cells, the RNA has a few ribosomes attached to it so many ribosomes are carrying out protein synthesis simultaneously on one piece of RNA.
Describe the process of termination(4)
- Ribosome encountering a stop codon UAG, UGA or UAA
- The Entry of a stop codon to the Amino acyl or A site results in the gaining of a protein called release factor 1 (or eRF1)
- This triggers Hydrolysis of GTP and the terminal peptidyl tRNA bond releasing the polypeptide chain
- dissociation of the ribosomal subunits releasing the mRNA allowing for recycling of the factors and the protein machinery
Summarise the energy consumption of protein synthesis in each stage and tRNA recharging
1) Charging tRNA with amino acids → 1 ATP 2) Initiation of polypeptide synthesis → GTP + nATP
3) Elongation of the polypeptide → 2 x GTP per amino acid
4) Termination of synthesis → GTP
How can antibiotics kill bacteria without disrupting human cells?(2)
Bacterial and mammalian ribosomes are different so certain substances can be used to solely target specific ribosomes and only affect protein synthesis in bacteria without affecting eukaryotic protein synthesis
Recall the proportions of RNA type
rRNA- 80-85% tRNA- 10-15% mRNA-2-5%
Recall 5 antibiotics and the ribosome subunit they target
Streptomycin- Small ribosomal subunit- Inhibition of initiation misreading of genetic code
Tetracyclines- Small ribosomal subunit- Inhibition of aminoacyl tRNA binding to ribosome
Chloramphenicol- Large ribosomal subunit- Inhibition of peptidyl transferase activity
Erythromycin-Large ribosomal subunit- Inhibition of translocation
Neomycins- Multiple sites- Several effects
Recall the expense of different stages of protein synthesis
1) Charging tRNA with amino acids → 1 ATP
2) Initiation of polypeptide synthesis → GTP + nATP
3) Elongation of the polypeptide → 2 x GTP per amino acid
4) Termination of synthesis → GTP
What is the central dogma of molecular biology?
DNA-RNA-Protein
What is a UTR?
Untranslated region near the beginning of the mRNA, they are exons.
What is the role of 5’UTR?
determines the rate at which the protein is synthesized
What are the Ribosomal subunits?
60S subunit and 40S subunit which combined make up 80S ribosome
where in the ribosome does protein synthesis happen?
the cleft of the ribosome
what happens once the start codon is found?
- 60S binds to 40S forming complete 80S ribosome.
→EIF2 hydrolyses the GTP to form GDP and releases pyrophosphate.
→All EIFs are released from the ribosome.
What do viruses do that prevent antiviral proteins from being made?
inactivate EIF3 and EIF4
what is translocation and what does it require?(3)
- tRNAs moving from the A & P sites to the P & E sites respectively.
- uses up one GTP
- the enzyme translocase and EF2.
what enzyme recharges tRNA?
→aminoacyl tRNA synthase
what is produced when tRNA is recharged
AMP and pyrophosphate
What is the name of the eukaryotic initiation factor that binds to the 40S subunit at the beginning of initiation, and what is its purpose?
EIF3, and it’s there to prevent the association of the 40S subunit with the 60S subunit.
what is synonymous alteration in genes?
when a nucleotide changes in the chain, but the coding remains the same.
what are the two subunits of a ribosome called, and what rRNA (and proteins) are they made up of?
→smaller 40S subunit, and a larger 60S subunit.
→The 40S is made up of 18S rRNA (and 33 proteins), →60S is made up of 5S, 5.8S and 28S rRNA (and 49 proteins).
what are the three sites in the ribosome called?
Aminoacyl tRNA binding site
→ Peptide binding site
→ Exit
(APE)
what is the 48S initiation complex?
→40S ribosomal subunit
→EIF3
→met-tRNAi
→EIF2
→GTP
what are the components of a prokaryotic ribosome?
→70S ribosome
→50S and 30S subunits
→50S: 23S and 5S rRNA (31 proteins)
→30S: 16S (21 proteins)
what are the components of a eukaryotic ribosome?
→80S ribosome
→60S and 40S subunits
→60S: 5S, 5.8S and 28S (49 proteins)
→40S: 18S (33 proteins)
what is EIF3 and what is it for?
→ eukaryotic initiation factor
→prevent the association of the 40S subunit with the 60S subunit.
What does Streptomycin?
Inhibition of initiation, misreading of genetic code
Targets small ribosomal unit of bacteria
What does chloramphenical do?
- inibition of peptidyl transferace activity
- targets large subunit
what is EIF3 and what is it for?
→ eukaryotic initiation factor
→prevent the association of the 40S subunit with the 60S subunit.
