126 Questions
What is the primary function of aminoacyl-tRNA synthetases?
To covalently bond amino acids to tRNA
During chain elongation, what occurs between the amino acids?
Peptide bond formation
What is the term for a sequence of three bases that specifies one amino acid?
Codon
In which step of protein synthesis does the first aminoacyl-tRNA bind to the mRNA?
Chain initiation
What is the purpose of the ribosome in protein synthesis?
To provide a site for protein synthesis
What is the consequence of degeneracy in the genetic code?
Multiple codons can code for the same amino acid
What is the term for the process of forming a peptide bond between amino acids?
Peptide bond formation
What is the final step of protein synthesis?
Chain termination
If the genetic code had a one-to-one relationship between bases and amino acids, how many amino acids could be encoded?
4
What would be the consequence if the ribosome moved along the mRNA more than three bases at a time?
The code would be punctuated
What is the term for the situation where more than one triplet can encode the same amino acid?
Degenerate code
What is the significance of the 'wobble' base in the genetic code?
It allows for more room for variation in the codon
What is the term for a mutation in the DNA that does not lead to a change in the amino acid translated?
Silent mutation
What is the significance of the second base of the codon in the genetic code?
It is important for determining the type of amino acid
What is the characteristic of codons that share the same first letter?
They code for amino acids that are products of one another
How many possible genetic codes were calculated to have the effect of reducing errors in protein function?
100
What is the advantage of the genetic code having multiple codons for a single amino acid?
It decreases the error rate in protein function
What is the significance of the 64 possible codons in the genetic code?
They are all used to encode the 20 amino acids or one of the three stop signals
What is the purpose of using synthetic polyribonucleotides as messengers in laboratory systems?
To establish the code for the four possible homopolymers quickly
What is the product of poly U as a messenger in polypeptide synthesis?
Polyphenylalanine
What is the significance of the experiment using an alternating copolymer as the messenger?
It proves that the code is a triplet code
What is the purpose of the filter-binding assay?
To identify the codon assignment of tRNA molecules
How do aminoacyl-tRNAs bind to ribosomes in the presence of the correct trinucleotide?
Through strong hydrogen bonding
Why is there not a 1:1 correspondence between codons and tRNA molecules?
Because of the wobble phenomenon
Which base of an anticodon is involved in the wobble phenomenon?
The first base at the 5' end
What is the direction of reading mRNA during protein synthesis?
From the 5' to the 3' end
What is the result of using an alternating copolymer as the messenger in polypeptide synthesis?
An alternating polypeptide is produced
What is the significance of the filter-binding assay in codon assignment?
It identifies the codon assignment of tRNA molecules
Which of the following bases can form a base pair with uracil in the wobble position of the anticodon?
Adenine
When the wobble position is occupied by which of the following bases, variations in hydrogen bonding do not occur?
A (adenine) or C (cytosine)
What is the result of the wobble model on the required number of tRNAs?
Fewer tRNAs are required
Which of the following codons would be translated as a glycine in the marine alga Acetabularia?
UAA
What is the energy source for the formation of the aminoacyl-AMP bond?
Hydrolysis of ATP
What is the product of the first step of amino acid activation?
Aminoacyl-AMP
What is the chemical nature of aminoacyl-AMP?
A mixed anhydride of a carboxylic acid and a phosphoric acid
What is the result of the hydrolysis of pyrophosphate (PPi) to orthophosphate (Pi)?
Energy is released
Which of the following best describes the effect of the wobble model on the genetic code?
It minimizes the damage caused by misreading of the code
What is the consequence of rare codons having low quantities of matching tRNAs?
The speed of translation is decreased
What is the function of IF-3 protein in the initiation of protein synthesis?
Prevents premature binding of the 50S subunit
What is the source of the formyl group in the formation of N-formylmethionine-tRNAfmet?
N10-formyltetrahydrofolate
What is the function of the Shine-Dalgarno sequence in mRNA?
Acts as a ribosomal binding site
What is the consequence of the hydrolysis of GTP to GDP and Pi during initiation?
The release of initiation factors
How does the 30S subunit recognize the initiator tRNA?
Through a single C-A mismatched base pair near the acceptor stem
What is the function of IF-2 protein in the initiation of protein synthesis?
Aids in the selection of the initiator tRNA
What is the purpose of the triplet sequence 3'-UAC-5' in tRNAfmet?
Recognizes the AUG triplet in the mRNA sequence
What is the consequence of the formation of the 70S initiation complex?
The release of initiation factors
What is the function of IF-1 protein in the initiation of protein synthesis?
Binds to IF-3 and IF-2, and facilitates their action
What is the purpose of the editing site in the tRNA synthetase?
To hydrolyze the incorrectly acylated aminoacyl-tRNA
What is the source of greater specificity in the recognition of tRNAs by aminoacyl-tRNA synthetases?
The specific binding sites on the tRNA
What is the purpose of the three binding sites for tRNA on the 50S subunit of the 70S ribosome?
To facilitate the synthesis of the polypeptide chain
What is the significance of the two-stage reaction in aminoacylation?
It enables selectivity to operate at two levels: amino acid and tRNA
What is the role of the peptidyltransferase center in the ribosome?
It forms a peptide bond between amino acids
What is the consequence of the convergent evolution of Class I and Class II aminoacyl-tRNA synthetases?
They have different structures and functions
What is the significance of the specific binding of mRNA and aminoacyl-tRNAs to the ribosome?
It ensures the accuracy of translation
What is the direction of polypeptide chain growth in protein synthesis?
From the N-terminal end to the C-terminal end
What is the term for the extra level of proofreading by the synthetase?
The second genetic code
What is the significance of the Mg2+ requirement for the synthetase enzyme?
It enables the formation of an aminoacyl-AMP
What is the result of the recognition of the correct tRNA by the aminoacyl-tRNA synthetase?
The fidelity of translation
What is the main function of EF-Tu during chain elongation?
To guide the aminoacyl-tRNA into the A site and align the anticodon with the mRNA codon
What is the consequence of EF-Tu binding an activated tRNA too well?
The tRNA is not delivered to the ribosome
What is the result of the peptide bond formation reaction during chain elongation?
A dipeptidyl-tRNA at the A site and an uncharged tRNA at the P site
What is the role of EF-G during the translocation step?
To move the mRNA with respect to the ribosome
What is the purpose of puromycin in experiments studying chain elongation?
To mimic the 3' end of an aminoacyl-tRNA and study chain elongation
What is the significance of the structure of the ribosome and EF-Tu complex?
It provides insight into the mechanism of EF-Tu-mediated aminoacyl-tRNA delivery
What is the outcome of the translocation step during chain elongation?
All of the above
What is the purpose of the P site during chain elongation?
To hold the growing peptide chain
What is the function of EF-Ts during chain elongation?
To regenerate EF-Tu-GTP
What is the significance of the 3.6-angstrom resolution structure of the ribosome and EF-Tu complex?
It provides insight into the mechanism of EF-Tu-mediated aminoacyl-tRNA delivery
What was the significance of the discovery of RNA-based peptidyl transferase?
It provided evidence for an 'RNA world' where RNA carried the message and processed it.
What is the primary function of the poly A binding protein (Pab1p)?
To link the poly A tail to eIF4G
What is the term for a complex of mRNA with several ribosomes attached to it?
Polyribosome
Which of the following factors can influence the selection of the start codon?
The presence of mRNA secondary structure
What is the main difference between prokaryotic and eukaryotic translation?
Eukaryotes have a more complex process of chain initiation.
What is the purpose of the 5' cap and 3' poly A tail in eukaryotic mRNA?
They are essential for translation.
What is the term for the complex formed by the mRNA and the seven eIFs?
The 48S preinitiation complex
What is the primary function of puromycin in protein synthesis?
To bind to the P site and block translocation
Which of the following is a characteristic of the eukaryotic ribosome?
It has an A site and a P site, but no E site
What is the role of eIF4E in eukaryotic translation?
It is a cap-binding protein that helps to recruit the ribosome.
What is the role of RF-3 in protein synthesis?
To facilitate the activity of RF-1 and RF-2
What is the function of the antibiotic chloramphenicol?
To inhibit peptidyl transferase activity in prokaryotes
What is the consequence of the lack of cell compartmentalization in prokaryotes?
It allows for simultaneous transcription and translation.
Which of the following is a characteristic of the eukaryotic elongation factor eEF2?
It causes translocation of the ribosome
What is the significance of the conserved sequence Pro-X-Thr in RF-1?
It controls the codon specificity of RF-1
What is the significance of the scanning mechanism in eukaryotic translation?
It helps to locate the start codon on the mRNA.
What is the function of selenocysteine in proteins?
To clear reactive oxidative molecules during thyroid hormone activation
What is the function of the release factor in eukaryotic translation?
To catalyze the hydrolysis of the bond between the C-terminal amino acid and the tRNA
What is the role of the 43S preinitiation complex in eukaryotic translation?
It is the initial stage of translation where the ribosome and mRNA are assembled.
What is the term for the tRNA that allows translation to continue through a stop codon?
Suppressor tRNA
What is the significance of the absence of a Shine-Dalgarno sequence in eukaryotes?
It means that the start codon is located differently.
What is the unique feature of selenocysteine synthesis?
It is synthesized while bound to its tRNA
What is the recent evidence that challenges the traditional view of eukaryotic translation?
That the nucleus has all the components necessary for translation
What is the function of the O-phosphoseryl-tRNA kinase in selenocysteine synthesis?
To phosphorylate the b-hydroxyl group of Ser-tRNASec
What is the consequence of the mutation of the putative RNA bases involved in the catalytic mechanism of peptidyl transferase?
It has no effect on the efficiency of peptidyl transferase.
What is the significance of the ribosome being a ribozyme?
It is an RNA molecule that catalyzes peptide bond formation
What is the percentage of the cell's protein synthesis that is suggested to occur in the nucleus?
10-15%
What was the significance of the Tetrahymena snRNP discovery?
It showed that RNA can also catalyze reactions
What was the significance of the X-ray crystallography study of the large ribosomal subunit?
It determined the complete structure of the large ribosomal subunit to 2.4-Å resolution
What is the consequence of the misacylation of tRNASec by the seryl-tRNA synthetase?
It leads to the formation of a peptide bond between selenocysteine and the growing polypeptide chain
What is the significance of the discovery that CUG can be used as a start codon in peptide synthesis?
It demonstrates the flexibility of start codon usage in eukaryotic translation.
What is the primary function of leader sequences in proteins destined for export?
To direct proteins to their proper destination
What is the consequence of a single break in the peptide backbone of a protein?
The protein is rapidly degraded
What is the role of ribosomes in protein folding?
To act as a chaperone and prevent misfolding
What is the significance of posttranslational modifications in protein synthesis?
They are involved in protein maturation and activation
What is the fate of abnormal proteins formed from errors in transcription or translation?
They are degraded rapidly
What is the significance of the dynamic state of protein turnover?
It enables the rapid adaptation of cells to changing environments
What is the consequence of the absence of posttranslational modifications in protein synthesis?
Proteins are inactive or malfunctioning
What is the role of the Golgi apparatus in protein synthesis?
To direct proteins to their final destination
What is the significance of protein degradation in the regulation of gene expression?
It is an essential control mechanism at the level of protein degradation
What is the primary function of ubiquitin in protein degradation?
To condemn proteins to destruction in a proteasome
What is the name of the enzyme that transfers ubiquitin to the targeted protein?
Ubiquitin-protein ligase (E3)
What is the consequence of having an acetyl group at the N-terminus of a protein?
The protein is protected from ubiquitin-mediated degradation
What is the role of tRNA in the degradation of proteins with an acidic residue at the N-terminus?
It is part of the destruction pathway for these proteins
What is the result of the wobble base in the anticodon of tRNA?
It breaks Watson-Crick base-pairing rules
What is the significance of the 'second genetic code'?
It refers to the specificity of amino acid-tRNA synthetases
What is the consequence of having the wrong amino acid loaded onto a tRNA?
It is quickly hydrolyzed by the synthetase
What is the role of the ubiquitin-protein ligase (E3) in the degradation of proteins?
It transfers ubiquitin to the targeted protein
What is the significance of the N-terminal amino acid in protein degradation?
It influences the susceptibility of the protein to ubiquitin-mediated degradation
What is the role of the lysosome in protein degradation?
It is a degradative subcellular organelle
What is the role of the Shine-Dalgarno sequence in prokaryotic translation?
To align the ribosome for correct translation initiation
What is the significance of EF-Tu in E. coli?
It adds a level of complexity to the elongation of proteins, leading to proofreading
What is the primary difference between prokaryotic and eukaryotic translation initiation?
The number of initiation factors and elongation factors
Why do proteins often require the help of a chaperone to arrive at the correct structure?
Because of possible interactions with other proteins before the nascent protein chain is complete
What is the role of ubiquitin in protein degradation?
To target proteins for destruction
What is the significance of the 5' cap and 3' poly A tail in eukaryotic mRNA?
They are involved in the formation of the initiation complex
Why are damaged proteins degraded very quickly?
Because they are a threat to cellular function and integrity
What is the role of the Kozak sequence in eukaryotic translation initiation?
To surround the correct AUG start codon
Study Notes
Protein Biosynthesis
- Protein biosynthesis is a complex process that requires ribosomes, messenger RNA (mRNA), transfer RNA (tRNA), and protein factors.
- The ribosome is the site of protein synthesis.
- mRNA and tRNA, bound to the ribosome, ensure the correct order of amino acids in the growing protein chain.
Activation of Amino Acids
- Amino acid activation involves the formation of an aminoacyl-tRNA, which is a covalent bond between an amino acid and tRNA.
- The process occurs in two steps, catalyzed by aminoacyl-tRNA synthetase.
- The first step forms an aminoacyl-AMP, and the second step transfers the aminoacyl moiety to tRNA.
The Genetic Code
- The genetic code is a triplet, nonoverlapping, commaless, degenerate, and universal code.
- A sequence of three bases (codon) specifies one amino acid.
- The code is degenerate, meaning that more than one triplet can encode the same amino acid.
- The genetic code has withstood billions of years of natural selection, and its degeneracy acts as a buffer against deleterious mutations.
Determining the Genetic Code
- The genetic code was determined through experiments using synthetic polyribonucleotides as messengers.
- Homopolynucleotides and alternating copolymers were used to establish the code for the four possible homopolymers.
- The filter-binding assay was used to establish the codon assignments.
Codon-Anticodon Pairing and Wobble
- Codon-anticodon pairing occurs during protein synthesis, with the codon forming base pairs with the complementary anticodon of tRNA.
- The wobble position, the first base of the anticodon, allows for variations in hydrogen bonding.
- The wobble model provides insight into the degeneracy of the code, with some tRNAs able to recognize more than one codon.
Amino Acid Activation and the "Second Genetic Code"
- The aminoacyl-tRNA synthetase contributes to the accuracy of the translation process.
- The "second genetic code" refers to the specificity of the aminoacyl-tRNA synthetase, which ensures that the right amino acid pairs with the right tRNA.
- The synthetase has an editing site that detects and corrects incorrectly acylated tRNAs.
Prokaryotic Translation
- Prokaryotic translation differs from eukaryotic translation.
- The process can be divided into stages: chain initiation, chain elongation, and chain termination.
- Ribosomal architecture facilitates the binding of mRNA and aminoacyl-tRNAs.
- The initial N-terminal amino acid of all proteins in prokaryotes is N-formylmethionine (fmet).
Chain Initiation
- Chain initiation in prokaryotes requires the formation of an initiation complex, involving mRNA, the 30S ribosomal subunit, fmet-tRNAfmet, GTP, and three protein initiation factors.
- The initiation complex ensures the correct start of polypeptide synthesis.### Protein Synthesis
- Initiation of Translation: The process of protein synthesis starts with the initiation of translation, where a 30S ribosomal subunit, mRNA, and fmet-tRNAfmet form a 30S initiation complex.
- Shine-Dalgarno Sequence: A purine-rich leader sequence (5'-GGAGGU-3') in the mRNA, which lies about 10 nucleotides upstream of the AUG start signal, acts as a ribosomal binding site.
- Hydrolysis of GTP: The hydrolysis of GTP to GDP and Pi favors the process of initiation by providing energy, and the initiation factors are released.
Chain Elongation
- Three Binding Sites: The 50S subunit of the 70S ribosome has three binding sites for tRNA: P (peptidyl) site, A (aminoacyl) site, and E (exit) site.
- Aminoacyl-tRNA Binding: The second amino acid specified by the mRNA binds to the A site, and a triplet of tRNA bases forms hydrogen bonds with a triplet of mRNA bases.
- Elongation Factors: EF-P, EF-Tu, and EF-Ts (temperature-unstable and temperature-stable elongation factors, respectively) are required for chain elongation.
- Peptide Bond Formation: A peptide bond is formed in a reaction catalyzed by peptidyl transferase, which is a part of the 50S subunit.
Chain Termination
- Stop Signals: The codons UAA, UAG, and UGA are the stop signals, and they are recognized by proteins called release factors.
- Release Factors: RF-1 and RF-2 bind to the A site and affect the activity of the peptidyl transferase, leading to the hydrolysis of the bond between the carboxyl end of the peptide and the tRNA.
The 21st Amino Acid
- Selenocysteine: Selenocysteine is a cysteine residue with a selenium atom instead of sulfur, and it is the 21st amino acid found in proteins from eukaryotes and prokaryotes.
- Synthesis of Selenocysteine: Selenocysteine is synthesized while bound to its tRNA, and it requires specialized elongation factors, such as SelB in bacteria and EFsec in humans.
The Ribosome Is a Ribozyme
- Catalytic Ability of RNA: The discovery of the self-splicing ability of the Tetrahymena snRNP showed that RNA can catalyze reactions, and the structure of the large ribosomal subunit revealed that RNA has catalytic ability.
- Peptidyl Transferase: The peptidyl transferase active site is located in the ribosome, and it is responsible for the formation of peptide bonds.
Polysomes
- Polysomes: A complex of mRNA with several ribosomes attached to it, each bearing a polypeptide in various stages of completion.
- Coupled Translation: In prokaryotes, translation begins shortly after mRNA transcription, and the mRNA is still being transcribed while it is being translated.
Eukaryotic Translation
- Differences from Prokaryotic Translation: Eukaryotic translation differs from prokaryotic translation in the details of initiation, elongation, and termination.
- Initiation Factors: Thirteen initiation factors, designated as eIF, are involved in eukaryotic translation.
- Chain Initiation: The assembly of a 43S preinitiation complex, followed by the recruitment of mRNA, and the formation of a 48S preinitiation complex.
- Elongation Factors: Eukaryotic elongation factors, eEF1 and eEF2, are involved in peptide chain elongation.
Posttranslational Modification of Proteins
- Processing of Polypeptides: Newly synthesized polypeptides are frequently processed before they reach their biologically active form.
- Cleavage of Precursors: Specific bonds in precursors can be hydrolyzed, and leader sequences can be removed.
- Addition of Cofactors: Cofactors, such as heme groups, can be added to the polypeptide.
- Covalent Modifications: Amino acid residues can be covalently modified, such as the conversion of proline to hydroxyproline.
Folding of Proteins
-
Proper Folding: The primary structure of the protein conveys enough information to specify its three-dimensional structure.
-
Ribosomes and Folding: Research indicates that the ribosomes themselves are involved in protein folding.### Protein Synthesis and Degradation
-
A 2011 study in Science found that the presence of a ribosome can confer the ability for a protein to fold correctly, potentially acting as its own chaperone to prevent misfolding.
-
Translation efficiency is influenced by various factors, including tRNA abundance, codon order, and mRNA secondary structure.
-
Proteins are in a dynamic state, with a 50% turnover every three days, and abnormal proteins are degraded quickly.
-
The cell uses specific degradation pathways, such as lysosomes and proteasomes, to target proteins for destruction.
Protein Degradation Mechanisms
- Proteins are directed to lysosomes by specific signal sequences, often added in a posttranslational modification step.
- Proteasomes are found in both prokaryotes and eukaryotes, and specific pathways exist to target proteins for destruction.
- In eukaryotes, the most common mechanism for targeting proteins for destruction is through ubiquitinylation.
- Ubiquitin is a small, highly conserved polypeptide (76 amino acids) that condemns a protein to destruction in a proteasome.
Ubiquitinylation Process
- Three enzymes are involved in ubiquitinylation: ubiquitin-activating enzyme (E1), ubiquitin-carrier protein (E2), and ubiquitin-protein ligase (E3).
- The ligase transfers the ubiquitin to free amino groups on the targeted protein, either the N-terminus or lysine side chains.
- Proteins must have a free α-amino group to be susceptible to ubiquitin-mediated degradation.
- The nature of the N-terminal amino acid influences its susceptibility to ubiquitinylation.
Protein Quality Control
- The cell monitors protein quality control through a combination of chaperone and protein degradation processes.
- Proteins that are unfolded or misfolded can go through the chaperone process for correct folding or be degraded through the ubiquitination system.
- Damaged proteins are degraded very quickly.
Additional Topics
- The genetic code was determined through a variety of techniques, including the use of synthetic mRNA sequences and the filter-binding experiment of Nirenberg.
- The "second genetic code" refers to the specificity with which aminoacyl-tRNA synthetases load amino acids onto tRNA molecules.
- In prokaryotic translation, the correct AUG start codon is identified by its proximity to a consensus sequence called the Shine-Dalgarno sequence.
- EF-Tu is an important elongation factor in E. coli, as it delivers aminoacyl-tRNA to the ribosome only when the codon and anticodon match.
Test your understanding of protein synthesis, including the role of aminoacyl-tRNA synthetases, peptide bond formation, and codon sequences. Quiz covers the steps of protein synthesis and the consequences of degeneracy in the genetic code.
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