Roper - Ribosomes Flashcards
How big is a ribosome?
It is a similar size to viruses
It constitutes 25% of the mass of a bacterium.
What is a ribosome made of?
A ribozyme decorated with proteins.
Proteins modulate the properties of hte ribosome but RNA carry out the condensation reactions
What does S stand for when describing the size of ribosomal subunits?
Svedberg, the inventor of analytical ultracentrifugation - it measures the buoyant molecular weight of biological molecules
What subunits are prokaryotic ribosomes made up of?
30S and 50S to make 70S
What subunits are eukaryotic ribosomes made up of?
40S and 60S to make 80S
these are larger and more complex that prokaryotes
What are the three types of RNA molecules used by ribosomes?
tRNAs (transfer), rRNAs (ribosomal) and mRNAs (messenger)
What drug targets ribosomes?
Puromycin
What structural features do ribosomal subunits have that contributes to its function?
The 50S subunit has an A, P and E site.
The 30S subunit has the tunnel through which mRNA feeds.
What is the A site?
where aminoacyl-tRNA bind, carrying the next amino acid
What is the P site?
where peptidyl-tRNA (with phosphodiester bond already formed with the previous amino acid) binds
What is the E site?
where the tRNA that was previously in the P site exits, now without any amino acid
What is the anticodon stem loop?
where the tRNA interacts with mRNA
How far apart is the anticodon stem loop from the site where peptide bond formation happens?
80 angstroms apart - very far which indicates how big the ribosome is and shows how the two processes are separated by huge molecular distances
What does the anticodon stem loop always end in and why?
CCA
The phosphate on the 2 or 3’ hydroxyl of adenine (last base on tRNA molecules) is what binds to amino acids.
What makes tRNA molecules so unstable?
tRNA synthetase puts amino acids on the 2’ phosphate on adenine but it can hop between both because they are chemically equivalent.
Crystal ribosome structures were published in 2000. What contributed to the increase in resolution of ribosome structures before the 2000s?
- X-ray sources and computing.
- Crystal Quality and source.
- X-ray Detectors
- Crystal Freezing techniques
- Phasing strategies
How has Electron Microscopy contributed to the increase in resolution of ribosome structures?
EM provided early micrographs, early ideas of shape.
Now Cryo-EM is the prominent technique to solve new ribosome structures
What did we learn from early structures of the ribosome?
That the interface between the two ribosomal subunits is almost entirely RNA.
Proteins in ribosomes are unusual; they have a globular head domain which tends to be on the outside, with long meandering structures which reach into the core of the ribosome but none get within 20 angstroms from the peptidyl transferase site.
What do we know about the structure of RNA in ribosomes?
Adenine residues are highly conserved in the interface between subunits and RNA helices.
What is the RNA morphology in the 30s subunit?
the 16s rRNA molecule has a distinct morphological shape in the subunit where the 5’ end forms the body, the central part forms the platform, the major 3’end forms the head and the minor 3’ end straddles the interface.
What is the RNA morphology in the 50s subunit?
the 23s rRNA is more intricately woven through the proteins to form a monolithic (strong, sturdy, singular) structure.
What have the structures we learned about peptide bond synthesis from got in common?
came from bacterial ribosome crystallised at pH 6.0, more acidic than optimum pH of 7.5
this could affect the bases so structures need to be taken with pinch of salt
What did Steitz and Moore 2003 show?
- Only RNA was needed - protein free 23s RNA could do it invitro later.
- Tunnel through the 50S where polypeptides go through as synthesised + end of the tunnel is where tRNA’s CCA end binds to mRNA.
- Photoaffinity labelling studies show that rRNA’s U-2584 and U-2585 are close to site of peptide bond formation - part of a highly conserved internal loop in the 5th domain of the 23sRNA
What did Steitz and Moore 2003 show?
- Only RNA was needed - protein free 23s RNA could do it invitro later - closest protein is 18 Angstroms away
- Tunnel through the 50S where polypeptides go through as synthesised + end of the tunnel is where tRNA’s CCA end binds to mRNA.
