Protein Synthesis II Flashcards
1) Define the terms triplet codes and degenerate when referring to the genetic code
- Triplet codes: 3 bases code for 1 amino acid
There are 20 amino acids to be produced so 4^3 = 64 provides a sufficient amount of combinations to form at least 20 amino acids - Degenerate: different codons can code for the same amino acid -> there are 3 special termination codons
- AUG : methionine (start codon)
- UAG, UGG, UGA (stop codons)
2) Describe the structure of a tRNA molecule
- 75-90 nucleotides
- cloverleaf structure due to extensive internal base-pairing
- unusual bases (contain uracil, not thymine)
- anticodon on central loop which reads the codons on mRNA and forms a complementary codon
- all tRNA molecules look similar -> only the amino acid they carry changes, some can recognise more than one codon
3) Define the term ‘wobble mechanism’
- the last base on the codon loop of tRNA can form non Watson-Crick base pairs
- does not alter the amino acid sequence of the protein
- allows a single tRNA to recognise more than one codon [allows a faster system]
- e.g. GCC, GCG, GCU and GCA all code for alanine (however there are some exceptions like serine)
4) Name the enzyme which ‘loads’ tRNA molecules with amino acids
Aminoacyl-tRNA synthetases (each one is specific)
- (each tRNA can only accept a single amino acid that is appropriate for its anticodon sequence)
5) State the reaction for the activation of amino acids by binding to tRNA
Amino acid + tRNA -> Amino acyl-tRNA
- ATP –> AMP + PPi
-energy for addition provided by ATP
6) How are the 20 different types of tRNA classified?
- Different types of tRNA classified by the amino acid they carry, e.g. tRNA^phe or tRNA^leu
7) How are activated amino acids (bonded with tRNA) named?
- e.g. tRNA^phe bonded to phenylalanine: Phe-tRNA^phe
8) Which enzyme is responsible for ensuring the correct tRNA is bonded to the correct amino acid?
tRNA synthetase
9) Describe the ribosome in E.Coli (prokaryote)
- 70s complex
- two sub-units 50s and 30s are combined so overall surface area is smaller (70) than sum (80)
- 3 binding sites present on ribosome (EPA for exit, polymerisation and accepting sites)
10) State the 3 stages of translation
- Initiation
- Elongation
- Termination
11) Describe the first stage of translation (initiation) in prokaryotes
- 30s subunit binds to 2 initiation factors: IF1 and IF3
- mRNA binds to the ribosome
- fmet-tRNA^fmet binds GTP-1F2 (GTP at 5’ end of mRNA) and enters the ‘P’ site of the 30s subunit
- fmet-tRNA^fmet anticodon loop lines up with AUG initiation codon
- 50s subunit binds to the 30s subunit (mRNA in between) and GDP + Pi released
12) What is the Shine-Dalgarno sequence and what is its function?
- A sequence on the bacterial mRNA strand which is a ribosomal binding site
- Upstream of (before) AUG start codon which binds to the 16s rRNA of the ribosome
13) What is the 16s rRNA of a ribosome?
- 16s rRNA is the anchor sequence (e.g. -35, -10 boxes) to the Shine-Dalgarno sequence on the mRNA strand, which ensures in-frame translation -> correct triplet codes translated each time
14) How can several proteins be formed from one mRNA strand?
- several Shine-Dalgarno sequences preceeding several AUG codons
- the ribosome will move along the strand, its 16s rRNA binding at each SDS, and translation will occur to produce proteins
15) Describe the second stage of translation in prokaryotes (elongation)
- Amino-acyl tRNA binds to an elongation factor (EF) and delivers its amino acid to the ribosome [at A site]
- Proof-reading pause occurs (GTP hydrolysis provides energy for peptide bond formation)
- Enzyme peptidyl transferase (in ribosome) causes the tRNA molecule to shift left (to ‘P’ site) and also catalyses the formation of peptide bonds between AAs
- Translocation : ribosome moves over so A site is empty, and the discharged tRNA^fmet (in ‘E’ site) leaves
16) What happens after the discharged tRNA has exited from the E site of the ribosome?
The next amino-acyl tRNA will bind to another elongation factor, bringing the next amino acid to the A site of the ribosome
17) Describe the final stage of translation in prokaryotes (termination)
- When a stop codon is reached (UAA, UGA, UAG), a release factor binds to the A site (at the termination codon) and the last tRNA leaves.
- The amino acid polypeptide then breaks off
18) Give 6 differences between prokaryotic and eukaryotic protein synthesis
- P: mRNA is unmodified -> translated as soon as its synthesised while E: immature ‘hnRNA’ is modified by capping/polyadenylation/splicing before translation
- P: 30s+50s > 70s ribosome, E: 40s+60s ->80s ribosome
- P: SDS interacts with 16rRNA for ribosome binding while E: interaction between ribosome and cap at 5’ end
- P: initiator tRNA is fmet-tRNA^fmet, E: met-tRNA^met
- P: mRNA can code for >1 protein, E: only 1 protein
- P: free polyribosomes in cytoplasm, E: “ “ & also on RER
19) Explain how some antibiotics are selective and give an example
- some kill bacteria by distinguishing between eukaryotic and prokaryotic transcription and translation
- alpha-aminitin : inhibits RNA polymerase II, involved in eukaryotic protein synthesis
20) Why do most antibiotics have minimal effects for eukaryotes?
- few drugs cross the inner mitochondrial membrane and there is a low rate of transcription
21) Give 6 examples of antibiotics that affect protein synthesis in prokaryotes and/or eukaryotes
- Actinomycin: binds DNA at transcription initiation complex and prevents elongation by RNA polymerase
- Rifamycin: inihibition of RNA synthesis, binds to RNA polymerase
- Streptomycin: affects initiation (30s subunit) and causes misreading of codons
- Erythromyocin: binds to 50s subunit, prevents relocation of ribosome
- Chloramphenicol: inhibits peptidyl transferase by binding to the 50s subunit
- Tetracyclines: inhibits binding of amino acyl-tRNA to the ribosome
