Section 29.

5Eukaryotic Protein Synthesis Differs from Prokaryotic Protein Synthesis Primarily in Translation
Initiation
The basic plan of protein synthesis in eukaryotes and archaea is similar to that in bacteria. The major structural and
mechanistic themes recur in all domains of life. However, eukaryotic protein synthesis entails more protein components
than does prokaryotic protein synthesis, and some steps are more intricate. Some noteworthy similarities and differences
are as follows:
1.
Ribosomes. Eukaryotic ribosomes are larger. They consist of a 60S large subunit and a 40S small subunit, which
come together to form an 80S particle having a mass of 4200 kd, compared with 2700 kd for the prokaryotic 70S
ribosome. The 40S subunit contains an 18S RNA that is homologous to the prokaryotic 16S RNA. The 60S
subunit contains three RNAs: the 5S and 28S RNAs are the counterparts of the prokaryotic 5S and 23S
molecules; its 5.8S RNA is unique to eukaryotes.
2.
Initiator tRNA. In eukaryotes, the initiating amino acid is methionine rather than N-formylmethionine. However, as
in prokaryotes, a special tRNA participates in initiation. This aminoacyl-tRNA is called Met-tRNAi or Met-tRNAf(the
subscript i stands for initiation, and f indicates that it can be formylated in vitro).
3.
Initiation. The initiating codon in eukaryotes is always AUG. Eukaryotes, in contrast with prokaryotes, do not use a
specific purine-rich sequence on the 5 side to distinguish initiator AUGs from internal ones. Instead, the AUG
nearest the 5 end of mRNA is usually selected as the start site. A 40S ribosome attaches to the cap at the 5 end
of eukaryotic mRNA (Section 28.3.1) and searches for an AUG codon by moving step-by-step in the 3 direction
(Figure 29.33). This scanning process in eukaryotic protein synthesis is powered by helicases that hydrolyze ATP.
Pairing of the anticodon of Met-tRNAi with the AUG codon of mRNA signals that the target has been found. In
almost all cases, eukaryotic mRNA has only one start site and hence is the template for a single protein. In
contrast, a prokaryotic mRNA can have multiple Shine-Dalgarno sequences and, hence, start sites, and it can
serve as a template for the synthesis of several proteins. Eukaryotes utilize many more initiation factors than do
prokaryotes, and their interplay is much more intricate. The prefix eIF denotes a eukaryotic initiation factor. For
example, eIF-4E is a protein that binds directly to the 7-methylguanosine cap (Section 28.3.1), whereas eIF-4A is
a helicase. The difference in initiation mechanism between prokaryotes and eukaryotes is, in part, a consequence
of the difference inRNA processing. The 5 end of mRNA is readily available to ribosomes immediately after
transcription in prokaryotes. In contrast, pre-mRNA must be processed and transported to the cytoplasm in
eukaryotes before translation is initiated. Thus, there is ample opportunity for the formation of complex secondary
structures that must be removed to expose signals in the mature mRNA. The 5 cap provides an easily
recognizable starting point. In addition, the complexity of eukaryotic translation initiation provides another
mechanism for gene expression that we shall explore further in Chapter 31.
4.
Elongation and termination. Eukaryotic elongation factors EF1 and EF1 are the counterparts of
prokaryotic EF-Tu and EF-Ts. The GTP form of EF1 delivers aminoacyl-tRNA to the A site of the ribosome, and
EF1 catalyzes the exchange of GTP for bound GDP. Eukaryotic EF2 mediates GTP-driven translocation in
much the same way as does prokaryotic EF-G. Termination in eukaryotes is carried out by a single release factor,

eRF1, compared with two in prokaryotes. Finally, eIF3, like its prokaryotic counterpart IF3, prevents the
reassociation of ribosomal subunits in the absence of an initiation complex.

Prokaryotic Translation:
1. It occurs on 70 S ribosomes.
2. It is a continuous process as both transcription and translation occur in cytoplasm.
6. It is a faster process, adds about 20 amino acids per second.
7. It requires 3 initiation factors IFI. IF2. IF3.
8. After translation, formyl group from first formylated methionine is removed, retaining
methionine in the polypeptide chain.
9. It requires two release factors RF1 (for UAG and UAA) and RF2 (for UAA and UGA) in the
termination.
10. mRNA life is short (some seconds to some minutes) as mRNA is less stable.
Eukaryotic Translation:
1. It occurs on 80 S ribosomes.
2. It is a discontinuous process as transcription occurs in nucleus while translation takes place in
cytoplasm.
3. mRNA is monicistronic
4. First amino acid is met (methionine).
5. Initiation codon is AUG. occasionally GUG or CUG.
6. It is a slower process that adds one amino acid per second, thus a slower process.
7. It requires a set of 9 initiation factors elF 1, 2, 3, 4A. 4B, 4C, 4D, 5. 6.
8. The whole of initiating methionine is removed from the polypeptide chain.
9. It requires single release factor eRF 1.
10. mRNA has a life of few hours to few days. It is quite stable.
Reverse Transcription
Reverse transcription is the process by which a reverse transcriptase enzyme mediates the creation of a DNA
complement (complementary DNA or cDNA) from an RNA strand. The discovery and use of reverse transcriptases has
greatly improved knowledge in the area of molecular biology. Reverse transcriptases are used for gene expression
analysis, to create cDNA libraries from mRNA and, along with other enzymes, allow cloning, sequencing, and
characterization of RNA.
Importance:
RTs bring about the replication of many groups of viruses, mobilization of rectrovirus and other transposable elements,
and many others that pioneer of this field are yet to understand [8].
Reverse transcriptase plays a vital role in viral replication. Bacterial and Eukaryotic Reverse transcriptases (RT) are all
RNA-dependent DNA polymerases which produce complementary DNA (cDNA) [2]. Never the less, the cDNAs produced
by these two rectroelement systems are fundamentally different [2].
In eukaryotes, RT replicates the entire genone or the transposable genetic units including the gene for RT [2]. This is done
to ensure that RT plays its role of producing identical complementary DNAs (cDNAs) of the original units, and is able to
produce infectious viruses or transposable units when integrated back into the host DNA [2].
One Gene One Polypeptide
The theory that each gene is responsible for the synthesis of a single polypeptide. It was originally stated as the one
gene-one enzyme hypothesis by the US geneticist George Beadle in 1945 but later modified when it was realized that
genes also encoded nonenzyme proteins and individual polypeptide chains. It is now known that some genes code for
various types of RNA involved in protein synthesis.
The Genetic Code Is Degenerate and Universal
Given the different numbers of letters in the mRNA and protein alphabets, scientists theorized that combinations of
nucleotides corresponded to single amino acids. Nucleotide doublets would not be sufficient to specify every amino acid

because there are only 16 possible two-nucleotide combinations (4 2). In contrast, there are 64 possible nucleotide triplets
(43), which is far more than the number of amino acids. Scientists theorized that amino acids were encoded by nucleotide
triplets and that the genetic code wasdegenerate. In other words, a given amino acid could be encoded by more than one
nucleotide triplet. This was later confirmed experimentally; Francis Crick and Sydney Brenner used the chemical mutagen
proflavin to insert one, two, or three nucleotides into the gene of a virus. When one or two nucleotides were inserted,
protein synthesis was completely abolished. When three nucleotides were inserted, the protein was synthesized and
functional. This demonstrated that three nucleotides specify each amino acid. These nucleotide triplets are called codons.
The insertion of one or two nucleotides completely changed the triplet reading frame, thereby altering the message for
every subsequent amino acid (Figure). Though insertion of three nucleotides caused an extra amino acid to be inserted
during translation, the integrity of the rest of the protein was maintained.
In addition to instructing the addition of a specific amino acid to a polypeptide chain, three of the 64 codons terminate
protein synthesis and release the polypeptide from the translation machinery. These triplets are called nonsense codons,
or stop codons. Another codon, AUG, also has a special function. In addition to specifying the amino acid methionine, it
also serves as the start codon to initiate translation. The reading frame for translation is set by the AUG start codon near
the 5' end of the mRNA.
The genetic code is universal. With a few exceptions, virtually all species use the same genetic code for protein synthesis.
Conservation of codons means that a purified mRNA encoding the globin protein in horses could be transferred to a tulip
cell, and the tulip would synthesize horse globin. That there is only one genetic code is powerful evidence that all of life on
Earth shares a common origin, especially considering that there are about 10 84 possible combinations of 20 amino acids
and 64 triplet codons.
Degeneracy is believed to be a cellular mechanism to reduce the negative impact of random mutations. Codons that
specify the same amino acid typically only differ by one nucleotide. In addition, amino acids with chemically similar side
chains are encoded by similar codons. This nuance of the genetic code ensures that a single-nucleotide substitution
mutation might either specify the same amino acid but have no effect or specify a similar amino acid, preventing the
protein from being rendered completely nonfunctional.