1. A variety of cellular components play important roles in translation
INTRODUCTION
The translation of the mRNA codons into amino acid sequences leads to the synthesis of proteins
A variety of cellular components play important roles in translation
These include proteins, RNAs and small molecules
In this chapter we will discuss the current state of knowledge regarding the molecular features of mRNA translation
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2. THE GENETIC BASIS FOR PROTEIN SYNTHESIS
Proteins are the active participants in cell structure and function
Genes that encode polypeptides are termed structural genes
These are transcribed into messenger RNA (mRNA)
The main function of the genetic material is to encode the production of cellular proteins
In the correct cell, at the proper time, and in suitable amounts
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3. The Genetic Code
Translation involves an interpretation of one language into another
In genetics, the nucleotide language of mRNA is translated into the amino acid language of proteins
This relies on the genetic code
The genetic information is coded within mRNA in groups of three nucleotides known as codons
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4. Multiple codons may encode the same amino acid.
Three codons do not encode an amino acid. These are read as STOP signals for translation
Multiple codons may encode the same amino acid.
These are known as synonymous codons
Triplet codons correspond to a specific amino acid

5. The code is nearly universal
Special codons:
AUG (which specifies methionine) = start codon
This defines the reading frame for all following codons
AUG specifies additional methionines within the coding sequence
UAA, UAG and UGA = termination, or stop, codons
The code is degenerate
More than one codon can specify the same amino acid
For example: GGU, GGC, GGA and GGG all code for lysine
In most instances, the third base is the degenerate base
It is sometime referred to as the wobble base
The code is nearly universal
Only a few rare exceptions have been noted
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6. An overview of gene expression
Note that the start codon sets the reading frame for all remaining codons

7. A Polypeptide Chain Has Directionality
Polypeptide synthesis has a directionality that parallels the 5’ to 3’ orientation of mRNA
During each cycle of elongation, a peptide bond is formed between the carboxyl group of the last amino acid in the polypeptide chain and the amino group in the amino acid being added
The first amino acid has an exposed amino group
Said to be N-terminal or amino terminal end
The last amino acid has an exposed carboxyl group
Said to be C-terminal or carboxy terminal end
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8. Last peptide bond formed in the growing chain of amino acids
H3N+ C C N C C N C C +
H3N+ C C H H H H H O – H O – R1 O R2 O R3 O R4 O
H3N+ C C N C C N C C N C C +
H2O H H H H H H H O –
Last peptide bond formed in the
growing chain of amino acids
(a) Attachment of an amino acid to a peptide chain
CH3 OH S
CH2 OH SH
H3C
CH3
CH2
CH2 CH
CH2
CH2 H H H H
Amino-
terminal
end H N + C C N C C N C C N C C N C C O –
Carboxyl-
terminal
end 3 H O H O H O H O H O
Methionine
Serine
Valine
Tyrosine
Cysteine
Peptide bonds 5′
A U G
A G C
GU U
U A C
U G C 3′
Sequence in mRNA
(b) Directionality in a polypeptide and mRNA
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9. Nonpolar amino acids are hydrophobic
There are 20 amino acids that may be found in polypeptides
Each contains a different side chain, or R group
Each R group has its own particular chemical properties
Nonpolar amino acids are hydrophobic
They are often buried within the interior of a folded protein
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10. Nonpolar and charged amino acids are hydrophilic
They are more likely to be on the surface of the protein
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11. Levels of Structures in Proteins
There are four levels of structures in proteins
1. Primary
2. Secondary
3. Tertiary
4. Quaternary
A protein’s primary structure is its amino acid sequence
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12. Within the cell, the protein will not be found in this linear state
Rather, it will adapt a compact 3-D structure
Indeed, this folding can begin during translation
The progression from the primary to the 3-D structure is dictated by the amino acid sequence within the polypeptide
129 amino acids long
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13. Levels of Structures in Proteins
The primary structure of a protein folds to form regular, repeating shapes known as secondary structures
There are two types of secondary structures
a helix
b sheet
Certain amino acids are good candidates for each structure
These are stabilized by the formation of hydrogen bonds
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14. The short regions of secondary structure in a protein fold into a three-dimensional tertiary structure
This is the final conformation of proteins that are composed of a single polypeptide
Proteins made up of two or more polypeptides have a quaternary structure
This is formed when the various polypeptides associate together to make a functional protein
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16. Functions of Proteins
To a great extent, the characteristics of a cell depend on the types of proteins it makes
Proteins can perform a variety of functions
A key category of proteins are enzymes
Accelerate chemical reactions within a cell
Can be divided into two main categories
Anabolic enzymes  Synthesize molecules and macromolecules
Catabolic enzymes  Break down large molecules into small ones
Important in generating cellular energy
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17. STRUCTURE AND FUNCTION OF tRNA
In the 1950s, Francis Crick and Mahon Hoagland proposed the adaptor hypothesis
tRNAs play a direct role in the recognition of codons in the mRNA
In particular, the hypothesis proposed that tRNA has two functions
1. Recognizing a 3-base codon in mRNA
2. Carrying an amino acid that is specific for that codon
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18. Recognition Between tRNA and mRNA
During mRNA-tRNA recognition, the anticodon in tRNA binds to a complementary codon in mRNA
tRNAs are named according to the amino acid they bear
The anticodon is anti-parallel to the codon
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19. tRNAs Share Common Structural Features
The secondary structure of tRNAs exhibits a cloverleaf pattern
It contains
Three stem-loop structures; Variable region
An acceptor stem and 3’ single strand region
The actual three-dimensional or tertiary structure involves additional folding
In addition to the normal A, U, G and C nucleotides, tRNAs commonly contain modified nucleotides
More than 80 of these can occur
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20. Other variable sites are shown in blue as well
Found in all tRNAs
Not found in all tRNAs
Other variable sites are shown in blue as well
The modified bases are:
I = inosine
mI = methylinosine
T = ribothymidine
UH2 = dihydrouridine
m2G = dimethylguanosine
y = pseudouridine
Structure of tRNA

21. Charging of tRNAs
The enzymes that attach amino acids to tRNAs are known as aminoacyl-tRNA synthetases
There are 20 types
One for each amino acid
Aminoacyl-tRNA synthetases catalyze a two-step reaction involving three different molecules
Amino acid, tRNA and ATP
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22. Charging of tRNAs
The aminoacyl-tRNA synthetases are responsible for the “second genetic code”
The selection of the correct amino acid must be highly accurate or the polypeptides may be nonfunctional
Error rate is less than one in every 100,000
Sequences throughout the tRNA including but not limited to the anticodon are used as recognition sites
Many modified bases are used as markers
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23. The amino acid is attached to the 3’ end by an ester bond

24. tRNAs and the Wobble Rule
The genetic code is degenerate
With the exception of serine, arginine and leucine, this degeneracy always occurs at the codon’s third position
To explain this pattern of degeneracy, Francis Crick proposed the wobble hypothesis
In the codon-anticodon recognition process, the first two positions pair strictly according to the A – U /G – C rule
However, the third position can actually “wobble” or move a bit
Thus tolerating certain types of mismatches
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25. RIBOSOME STRUCTURE AND ASSEMBLY
Translation occurs on the surface of a large macromolecular complex termed the ribosome
Bacterial cells have one type of ribosome
Found in their cytoplasm
Eukaryotic cells have two types of ribosomes
One type is found in the cytoplasm
The other is found in organelles
Mitochondria ; Chloroplasts
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26. RIBOSOME STRUCTURE AND ASSEMBLY
Unless otherwise noted the term eukaryotic ribosome refers to the ribosomes in the cytosol
A ribosome is composed of structures called the large and small subunits
Each subunit is formed from the assembly of
Proteins
rRNA
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28. Functional Sites of Ribosomes
During bacterial translation, the mRNA lies on the surface of the 30S subunit
As a polypeptide is being synthesized, it exits through a hole within the 50S subunit
Ribosomes contain three discrete sites
Peptidyl site (P site)
Aminoacyl site (A site)
Exit site (E site)
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30. STAGES OF TRANSLATION
Translation can be viewed as occurring in three stages
Initiation
Elongation
Termination
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31. Initiator tRNA
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32. The Translation Initiation Stage
The mRNA, initiator tRNA, and ribosomal subunits associate to form an initiation complex
This process requires three Initiation Factors
The initiator tRNA recognizes the start codon in mRNA
In bacteria, this tRNA is designated tRNAfmet
It carries a methionine that has been covalently modified to N-formylmethionine
The start codon is AUG, but in some cases GUG or UUG
In all three cases, the first amino acid is N-formylmethionine
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33. Component of the 30S subunit
The binding of mRNA to the 30S subunit is facilitated by a ribosomal-binding site or Shine-Dalgarno sequence
This is complementary to a sequence in the 16S rRNA
Component of the 30S subunit
Hydrogen bonding
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34. IF1 and IF3 bind to the 30S subunit.
The mRNA binds to the 30S subunit.
The Shine-Dalgarno sequence is
complementary to a portion of the
16S rRNA.
Portion of
16S rRNA
IF3
IF1
Shine-
Dalgarno
sequence
(actually 9
nucleotides long)
Start
codon 3′ 5′
IF2, which uses GTP, promotes
the binding of the initiator tRNA
to the start codon in the P site.
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35. This marks the end of the first stage
tRNAfMet
Initiator tRNA
GTP
IF2
IF3
IF1 3′ 5′
IF1 and IF3 are released.
IF2 hydrolyzes its GTP and is released.
The 50S subunit associates.
70S initiation complex
tRNAfMet
This marks the end of the first stage E P A
70S
initiation
complex 3′ 5′
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36. The Translation Initiation Stage
In eukaryotes, the assembly of the initiation complex is similar to that in bacteria
However, additional factors are required
Note that eukaryotic Initiation Factors are denoted eIF
The initiator tRNA is designated tRNAmet
It carries a methionine rather than a formylmethionine
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37. The start codon for eukaryotic translation is AUG
Scanning ribosome may pass over the first AUG
But in most cases, the start codon for eukaryotic translation is usually the first AUG after the 5’ Cap!
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38. Translational initiation in eukaryotes can be summarized as such:
A number of initiation factors bind to the 5’ cap in mRNA
These are joined by a complex consisting of the 40S subunit, tRNAmet, and other initiation factors
The entire assembly moves along the mRNA scanning for the right start codon
Once it finds this AUG, the 40S subunit binds to it
The 60S subunit joins
This forms the 80S initiation complex
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39. mRNA scanning

40. The Translation Elongation Stage
During this stage, the amino acids are added to the polypeptide chain, one at a time
This process, though complex, can occur at a remarkable rate
In bacteria  amino acids per second
In eukaryotes  6 amino acids per second
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41. Thus, the ribosome is a ribozyme!
The 23S rRNA (a component of the large subunit) is the actual peptidyl transferase
Thus, the ribosome is a ribozyme!
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42. tRNAs at the P and A sites move into the E and P sites, respectively
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43. The Translation Elongation Stage
16S rRNA (a part of the 30S ribosomal subunit) plays a key role in codon-anticodon recognition
It can detect an incorrect tRNA bound at the A site
It will prevent elongation until the mispaired tRNA is released
This phenomenon is termed the decoding function of the ribosome
It is important in maintaining the high fidelity in mRNA translation
Error rate: 1 mistake per 10,000 amino acids added
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44. The Translation Termination Stage
The final stage occurs when a stop codon is reached in the mRNA
In most species there are three stop or nonsense codons
UAG
UAA
UGA
These codons are not recognized by tRNAs, but by proteins called release factors
Indeed, the 3-D structure of release factors mimics that of tRNAs
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45. The Translation Termination Stage
Bacteria have three release factors
RF1, which recognizes UAA and UAG
RF2, which recognizes UAA and UGA
RF3, which does not recognize any of the three codons
It binds GTP and helps facilitate the termination process
Eukaryotes only have one release factor
eRF, which recognizes all three stop codons
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47. Bacterial Translation Can Begin Before Transcription Is Completed
Bacteria lack a nucleus
Therefore, both transcription and translation occur in the cytoplasm
As soon an mRNA strand is long enough, a ribosome will attach to its 5’ end
So translation begins before transcription ends
This phenomenon is termed coupling
A polyribosome or polysome is an mRNA transcript that has many bound ribosomes in the act of translation
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48. Coupling between transcription and translation in bacteria
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