Lecture 28. Protein translation and alternative splicing Flashcards
What does RNA splicing regulate in eukaryotes ?
The complexity of gene expression products
What are gene coding sequences called in eukaryotes ?
Exons
What are exons ?
Expressed regions
What are introns ?
Intervening regions
What are gene coding sequences interrupted by ?
non-coding sequences called introns
What is pre mRNA splicing carried out by ?
A large molecular complex called the splicesome
What is the function of a spliceosome ?
Recognizes the junctions between introns and exons and can exise the introns to produce a fully splice mRNA molecule
What studies show that genes were comprised of uninterrupted stretches of coding sequences ?
Studies in bacteria
Do introns evolve rapidly ?
Yes
What is the donor site ?
5’ splice site
What is the acceptor site ?
3’ splice site
What is a major source of protein diversity in higher eukaryotes ?
Alternative splicing
When does alternative splicing occur?
When the pre mRNA is processed in different ways to produce mature transcripts lacking one or more exons
What is the product of alternative splicing ?
Splice variants
Do splice variants have the same or different function to the full length splice form ?
Different function
What is alternative splicing regulated by ?
Molecules that sit on the pre-mRNA
What do molecules that sit on the pre-mRNA prevent ?
Access of the splicing machinery to different splice sites
What is the calcitonin/neuropeptide gene ?
Two completely different proteins made in thyroid cells verses neurons
What is the DSCAM gene in drosphilia for ?
Involved in specifying neural circuitry
How many different possible protein isoforms are there in the DSCAM gene in drosphilia ?
38000
Where are mRNAs translated into proteins ?
Ribosomes
Where does the process of translation begin ?
cytoplasm
What happens in the process of translation in the cytoplasm ?
mRNA binds with ribosomes
What are ribosomes ?
Protein synthesis machines
How many binding sites do ribosomes have ?
3
What are the roles of the binding sites in ribosomes ?
- Binding of mRNA
2. Attach tRNA molecules that carry amino acids to the mRNA template and are labelled
What are the three labels on one of the domains used to attach tRNA molecules ?
- Aminoacyl site
- Peptidyl site
- Exit site
How do tRNAs bring amino acids to the ribosomes ?
in a manner dictated by the coding sequence of the mRNA
What is tRNA function defined by ?
Structure of the molecule
What is the structure of tRNAs ?
Cloverleaf shaped polynucleotide sequences
What does the tRNA tail end have ?
An acceptor stem
What is the function of the acceptor stem ?
Binds a specific amino acid
What does the head of tRNA have ?
Three nucleotides that form the anticodon
What is the function of an anticodon ?
Recognises the corresponding codon sequence of the mRNA molecule
What do tRNA anticodons bind complementary to ?
Triplet codons of the mRNA
What are the three stages of translation ?
- Initiation
- Elongation
- Termination
Initiation: When does translation begin?
When the mRNA molecule binds to the small subunit of the ribosome
Initiation: Where does mRNA molecule bind to the ribosome in prokaryotes and eukaryotes ?
30S in prokaryotes and 40S in eukaryotes
Initiation: What makes the mRNA bind to the ribosome in both eukaryotes and prokaryotes ?
- The recognition of the 5’ cap of eukaryotic mRNAs
2. A sequence upstream of the AUG codon in prokaryotic mRNAs
Initiation: What is the shine Dalgarno sequence ?
AGGAGG - A sequence upstream of the initiator AUG codon in prokaryotic mRNA
Initiation: What is protein synthesis initiated by ?
An AUG codon on the mRNA
Initiation: What does the AUG codon signal ?
Both the interaction of the ribosome with mRNA and also the tRNA with the anticodon (UAC)
Initiation: What happens once the AUG codon is recognised ?
The large ribosomal subunit associates with the mRNA and the small ribosomal subunit and protein synthesis begins
Initiation: What does the start codon always code for ?
The amino acid methionine (AUG)
Initiation: Where does the start codon enter ?
The P site on the ribosome
Initiation: Where does the second codon enter ?
The A site
Initiation: Where does the anticodon of the tRNA carrying methionine base pair with ?
Temporarily with the start codon
Initiation: Where does the second anticodon complementary to the mRNA codon to the A site form a temporary base pair ?
The codon of the mRNA in the A site
Initiation: What does the amino acid attached to the tRNA in the A site and the methionine in the P site form ?
A polypeptide bond
Initiation: What is the polypeptide bond catalysed by ?
Peptidyl transferase
Elongation: What does the A (aminoacyl) site do ?
Binds incoming charged aminoacyl tRNAs
Elongation: What does the P (peptidyl) site do ?
Binds charged tRNAs carrying amino acids that have formed polypeptide bonds with the growing polypeptide chain but have not yet dissociated from their corresponding tRNA
Elongation: What does the E (exit) site do ?
Releases dissociated tRNAs so that they can be recharged with free amino acids
Elongation: Why does the ribosome move along the mRNA sequence and the tRNA that resided in the A site moves over to the P site ?
Due to ratchet like conformational change
Elongation: Where does the ratchet like conformational change take place ?
Between the two subunits of the ribosome
Elongation: Where is the energy for each peptide bond formation from ?
GTP hydrolysis
Elongation: Where does the tRNA that was previously in the P site move to ?
The exit site and then leaves the ribosome
Elongation: Where is the growing peptide chain attached to the tRNA left ?
The P site
Elongation: What enters the A site ?
A new mRNA codon
Elongation: What does the tRNA carrying a complementary amino acid connect with ?
The bases of the new codon in the A site
Elongation: What forms a new peptide ?
Two adjacent amino acids
Elongation: What is GTP hydrolysis catalysed by ?
A separate elongation factor
Elongation: Where does the tRNA molecule from the P site move to ?
The E site and is released into the cytoplasm where it binds to an amino acid of the same type
Elongation: What does a tRNA complementary to the new codon in the A site do ?
Creates a new peptide bond between the amino acid and the currently formed peptide chain
Termination: When does the process of translation stop ?
Until one of the stop codons enters the A site of the ribosome
Termination: What are the three stop codons ?
- TUU
- TUG
- TGU
Termination: When does the protein chain move to the cytoplasm ?
Once the final peptide bond is created
Termination: Where is the protein chain connected ?
Connected only to the tRNA located in the P site
What is the ribosome composed of ?
Two sub-units which are comprise of ribosomal RNAs and specific ribosomal proteins
How is the ribosome formed ?
The two sub units fit to each other and form a spherical structure
What do prokaryotic ribosomes have ?
70S ribosomes
Prokaryotic ribosomes: What is their structure ?
One large (50S) and one small (30S) subunit
Prokaryotic ribosomes: What is the small subunit built from ?
A 16S RNA subunit
Prokaryotic ribosomes: How long is the small subunit ?
1540 nucleotides
Prokaryotic ribosomes: How many ribosomal proteins does the small RNA subunit bind to ?
21
Prokaryotic ribosomes: What does the large subunit contain ?
A 5S RNA and a 23S RNA
Prokaryotic ribosomes: How long is each of the RNAs in the large subunit ?
120 and 2900 nucleotides
Prokaryotic ribosomes: How many proteins does the ribosomal RNA molecules bind to ?
31
Eukaryotic ribosomes: What does the eukaryotic have ?
One large (60S) and one small (40S) subunit
Eukaryotic ribosomes: What does the ne large (60S) and one small (40S) subunit form ?
A 80S ribosome
Eukaryotic ribosomes: How many proteins does the small subunit contain ?
33
Eukaryotic ribosomes: Where does the small subunit bind to and how long is it ?
- 18S RNA
2. 1900 nucleotides long
Eukaryotic ribosomes: How many RNAs do eukaryotes have in their large subunit ?
3
Eukaryotic ribosomes: What are the 3 RNAs in the large subunit and how long are each of them ?
- 28S - 4700 nucleotides
- 5.8S - 160 nucleotides
- 5S - 120 nucleotides
Eukaryotic ribosomes: How many proteins does 3 ribosomal RNAs bind to ?
46