- Initiated by GTP hydrolysis
- initiation factors (eukaryotic IFs) help assemble the 40S ribosomal subunit with the initiator tRNA and are released when the mRNA and the ribosomal 60S subunit assemble with the complex.
- Eukaryotes: 40S + 60S →80S (Even).
- PrOkaryotes: 30S + 50S → 70S (Odd).
- ATP—required for tRNA Activation (charging with aminoacid ).
- GTP— required for tRNA Gripping and Going places (translocation).
- Aminoacyl-tRNA binds to A site (except for initiator methionine)
- rRNA (“ribozyme”) catalyzes peptide bond formation, transfers growing polypeptide to amino acid in A site
- Ribosome advances 3 nucleotides toward 3′ end of mRNA, moving peptidyl tRNA to P site (translocation)
- Mnemonic APE
- A site = incoming Aminoacyl-tRNA.
- P site = accommodates growing Peptide.
- E site = holds Empty tRNA as it Exits.
- Stop codon is recognized by release factor, and completed polypeptide is released from ribosome.
- Posttranslational modifications
- Removal of N- or C-terminal propeptides from zymogen to generate mature protein (e.g., trypsinogen to trypsin).
- Covalent alterations
- Phosphorylation, glycosylation, hydroxylation, methylation, acetylation, and ubiquitination.
- Chaperone protein
- Intracellular protein involved in facilitating and/or maintaining protein folding.
- In yeast, some are heat shock proteins (e.g., Hsp60) that are expressed at high temperatures to prevent protein denaturing/misfolding.
Which one of the following statement is not true regarding translation:
- Requires tRNAs, ribosomes, mRNAs, amino acid pool and energy
- ATP is the only essential source of energy
- A polypeptide is synthesized from amino terminus to carboxyl terminus
- It proceeds in cytoplasm
ANS: B. Translation process requires both ATP and GTP as energy source. The process of protein synthesis by mRNA is called translation. This process requires tRNAs, ribosomes, mRNAs, amino acid pool and free energy. Polypeptide is synthesized from amino -carboxyl terminal end. mRNA is read from 5-3 direction. Proteins are synthesized on ribosomes, which are present on the rough endoplasmic reticulum in the cytoplasm.
Binding of an amino acid to a specific tRNA reaction is catalyzed by:
- Amino acyl rRNA synthetases
- Aminoacyl tRNA polymerases
- Amino acyl tRNA synthetases
- Amino acyl mRNA synthetases
ANS: 40. C. Amino acids are attached to their tRNAs by highly specific enzymes known as aminoacyl tRNA synthetases. There must be atleast 20 different amino acyl tRNA synthetases. In the first step of the activation reaction the synthetase enzyme attaches carboxyl group of an amino acid to the phosphate portion of ATP (Adenosine tri phosphate) with the hydrolysis of pyrophosphate.
The Wobble hypothesis refers to:
- Less stringency between the third base of the codon and the complementary anticodon
- No effect on mutations
- Codon and anticodon base pairing is parallel
- None of the above
ANS: A. Wobble hypothesis refers to the less stringency between the third base of the codon and complementary anticodon. These unusual base pairs can form partly because of flexibility of the bases in the anticodon loop of the tRNA, mRNA and tRNA base pairing is antiparallel because of this, base pairing is always between the first base of the anticodon tRNA and third base of the codon in mRNA. The first base of the anticodon is G (Guanosine), it can pair with either its normal partner C (Cytosine) or with U (Uracil). The wobbling hypothesis and the degeneracy of codon together will reduce the effect of mutation.
Which rRNA (Ribosomal ribo nucleic acid) of eukaryotes has no equal sized rRNA in the prokaryoes:
- 23S rRNA
ANS: 42. D.
- Prokaryotes and eukaryotes having large subunits 50S and 60S and small subunits 30S and 40S respectively.
- The large subunit of prokaryotes has two rRNAs 5S and 23S, whereas large subunit of eukaryotes has three rRNAs 5.8S, 5S and 28S. 5S and 23S rRNA of prokaryotes are equal to the 5S and 28S rRNA of eukaryotes. 5.8S rRNA has no equal rRNA in the prokaryotes.
- The small subunits in eukaryotes and prokaryotes has single ribosomal rRNA that is 18S larger than prokaryotic 16S rRNA.
Which amino acid is first incorporated in initiation of protein synthesis in prokaryotes:
- N-formyl methionine
- N-formyl glycine
ANS: B. In prokaryotes N-formyl methionine is the first amino acid incorporated in initiation of protein synthesis.
Aminoacyl tRNA synthetase links methionine to the tRNA and formyl from N10 formyl tetra hydrofolate is added to the amino group of methionine, this reaction is catalyzed by the enzyme transformylase to form N-formyl methionyl tRNA. Methionine is the first amino acid for initiation of protein synthesis in eukaryotes
- Which one of the following is the initiator anticodon of met- tRNA:
Pre initiation complex is formed between the small ribosomal unit, protein factor and an initiator tRNA carrying a methionine.
AUG is the initiator codon of mRNA, which pair with the anticodon UAC of initiator methionine tRNA (met tRNA). Initiation complex is formed by the association of larger subunit to form translation machinery.
Which of the following is a key structural feature, normally present on mRNA, if missing. would not allow a translation initiation complex to form?
(A) Intron–exon secondary structure
(C) The 5′ cap
(E) The poly-A tail
The answer is C: The 5′ cap. The 5′ cap of mRNA is recognized by initiation factors (specifi cally eIF4E) to allow ribosome assembly on the mRNA. The absence of a cap would not allow a translation initiation complex to form Introns are not found in mature mRNA (they are removed by splicing in the nucleus); thus, intron–exon secondary structure would not be present. Pseudouridine is found only in tRNA, not in mature mRNA. Thymine, while found in tRNA by posttranscriptional processing, is not found in mRNA (uracil is placed into the mRNA when an adenine is in the template strand), and the poly-A tail, at the 3′ end of the mRNA, adds stability to the mRNA, but it does not play a role in translation initiation.
Under conditions of active exercise, protein synthesis is reduced in the muscle. Under these conditions, which aspect of translation is inhibited?
(A) Inability to initiate translation
(B) Inability to elongate during translation
(C) Inability to terminate translation
(D) Inability to synthesize mRNA
(E) Inability to produce rRNA
2 The answer is A:
Inability to initiate translation. As muscle works, and AMP levels rise, the muscle wants to preserve its ATP for muscle contraction, and not to use it for new protein synthesis. The increase in AMP levels leads to the activation of AMP-activated protein kinase, which will phosphorylate and inactivate eIF4E (eukaryotic initiation factor 4E), which is a necessary component in recognizing the 5′ cap structure of the mRNA to allow ribosome assembly on the mRNA (see the figure in the answer to the previous question). The activation of the AMP-activated protein kinase does not alter elongation or the termination of translation. It does not block overall transcription, either of mRNA or rRNA (although it may lead to an inhibition of ribosomal biogenesis as well as the transcription of certain specific genes).
A mutant ribonuclease that specifi cally cleaves the large rRNA molecule into many pieces, destroying its secondary structure and its ability to bind to ribosomal proteins. This cell line has greatly reduced the rates of protein synthesis. large ribosomal RNA is required for which of the following steps of protein synthesis ?
(D) Peptide bond formation
(E) tRNA activation and charging
4 The answer is D:
Peptide bond formation. It is the large ribosomal RNA that catalyzes peptide bond formation, using peptides and amino acids in the “A” and “P” sites on the ribosome. Destroying the secondary structure of this rRNA via the aberrant ribonuclease will limit the ability of the ribosome to create peptide bonds. The large, ribosomal RNA molecule is not essential for the initiation, termination, elongation (moving the ribosome along the mRNA after peptide bond formation has occurred), or tRNA activation and charging.
The molecular mechanism responsible for toxic effects of diphtheritic toxin is which of the following?
(A) Activation of protein kinase A
(B) Activation of an elongation factor for translation
(C) Glycosylation of a G protein
(D) Inhibition of protein kinase A
(E) Inhibition of an elongation factor for translation
7 The answer is E:
Inhibition of an elongation factor for translation. The child has diphtheria, which is caused by a bacterium (Corynebacterium diphtheriae), which produces a toxin that leads to the inhibition of eEF2 (eukaryotic elongation factor 2), which is required for the movement of tRNA from the “A” site to the “P” site. The toxin catalyzes the ADP-ribosylation (using NAD+ as a substrate) of eEF2 to bring about this inhibition. If one treats such a child with nicotinamide (the reaction product resulting from the loss of ADP-ribose from NAD+), one can reverse and block the ADP-ribosylation reaction catalyzed by the toxin. The toxin has no effect on protein kinase A, nor does it glycosylate a G protein. Diphtheria causes sore throat, fever, swollen nodes (bull neck), weakness, hoarseness, painful swallowing, and chills. The hallmark of the disease is a thick, gray membrane covering the pharynx.
Which of the following is a drug that blocks prokaryotic peptide bond formation ?
8 The answer is C:
- Chloramphenicol. Chloramphenicol blocks peptide bond formation in prokaryotic ribosomes (with no effect on eukaryotic ribosomes).
- Cycloheximide has the same effect as chloramphenicol in eukaryotic cells but has no effect on prokaryotes.
- Rapamycin leads to the blockage of translation initiation, not peptide bond formation.
- Puromycin is a chain terminator, stopping protein synthesis but not directly inhibiting peptide bond formation.
- Rifampin inhibits prokaryotic mRNA synthesis and has no direct effect on translation.
- Rifampin might be used as prophylaxis for household contacts of meningococcal meningitis but is not as effective a treatment for the actual disease.
A young patient was prescribed erythromycin for a bacterial infection, but he developed hearing loss due to use of this drug. This occurred due to which of the following?
(A) Inhibition of mitochondrial protein synthesis
(B) Inhibition of mitochondrial RNA synthesis
(C) Inhibition of mitochondrial DNA replication
(D) Weakening and tearing of the eardrum
(E) Increased neuronal signaling in the inner ear
9 The answer is A:
Inhibition of mitochondrial protein synthesis. Ototoxicity (hearing loss) occurs with a subset of antibiotics because, in addition to affecting prokaryotic ribosomes, the drugs also have an effect on mitochondrial ribosomes. Mitochondria contain their own DNA, RNA polymerase, and protein synthesizing apparatus (recall that it is thought that during evolution, bacteria invaded eukaryotic cells and formed a symbiotic relationship, with the bacteria eventually becoming the mitochondria), which is very similar to the prokaryotic apparatus. Thus, certain drugs will affect mitochondrial protein synthesis, and the effects seem to be greatest on those organs that have high energy needs (such as neuronal tissue). Erythromycin does not affect mitochondrial RNA synthesis or DNA replication. It does not affect the ear drum, nor does it increase neuronal signaling in the inner ear.
An adult male is diagnosed with atypical pneumonia. His physician prescribes clarithromycin, which is specific for prokaryotic cells. Which of the following best explains the mechanism of prokaryotic specificity?
(A) The drug binds to the 50S ribosomal subunit of bacteria and inhibits f-met-tRNAi binding
(B) The drug binds to the 30S ribosomal subunit of bacteria and blocks initiation of protein synthesis
(C) The drug binds to the 50S ribosomal subunit of bacteria and blocks translocation
(D) The drug binds to the 30S ribosomal subunit of bacteria and blocks peptide bond formation
(E) The drug binds to both ribosomal subunits and prevents bacterial ribosome assembly
11 The answer is C:
- The drug binds to the 50S ribosomal subunit of bacteria and blocks translocation. Clarithromycin (an antibiotic in the macrolide family with erythromycin and azithromycin) is specific for the large ribosomal subunit of prokaryotes (it will not bind to eukaryotic ribosomes).
- When this drug binds to the large ribosomal subunit, translocation of the ribosome (movement along the mRNA) is blocked, which blocks overall protein synthesis. tRNA binding is not affected by clarithromycin, nor is there a blockage of the formation of an initiation complex.
- It is the large subunit (50S) that contains the peptidyl transferase activity, which is also not blocked by this agent.
Rapamycin aids in preventing an immune response to the transplant via which of the following mechanisms?
(A) The drug inhibits ribosome subunit assembly
(B) The drug inhibits cap formation
(C) The drug specifically inhibits RNA polymerase III
(D) The drug inhibits initiation of protein synthesis
(E) The drug inhibits antibody-specific transcription factors from binding to DNA
12 The answer is D:
The drug inhibits initiation of protein synthesis.
- Rapamycin inhibits the mammalian target of rapamycin (mTOR), which is a protein kinase.
- One of the many targets of mTOR is eIF4E binding protein (eIF4E is a required initiation factor for protein synthesis).
- When not phosphorylated, the binding protein binds tightly to eIF4E and prevents it from participating in the formation of the translational initiation complex, thereby blocking protein synthesis.
- When phosphorylated at multiple locations by mTOR, the binding protein falls off the initiation factor and allows translational initiation complexes to form.
- In the presence of rapamycin, mTOR has no kinase activity, and the binding protein remains bound to eIF4E, thereby inhibiting protein synthesis. The drug does not affect ribosome assembly. The drug also has no effect on transcription or RNA processing.