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Berg JM, Tymoczko JL, Stryer L. Biochemistry. 5th edition. New York: W H Freeman; 2002.

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Biochemistry. 5th edition.

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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:


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.


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).


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 in RNA 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.


Elongation and termination. Eukaryotic elongation factors EF1α and EF1βγ are the counterparts of prokaryotic EF-Tu and EF-Ts. The GTP form of EF1α delivers aminoacyl-tRNA to the A site of the ribosome, and EF1βγ 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.

Figure 29.33. Eukaryotic Translation Initiation.

Figure 29.33

Eukaryotic Translation Initiation. In eukaryotes, translation initiation starts with the assembly of a complex on the 5′ cap that includes the 40S subunit and Met-tRNAi. Driven by ATP hydrolysis, this complex scans the mRNA until the first AUG (more...)

29.5.1. Many Antibiotics Work by Inhibiting Protein Synthesis

Image caduceus.jpg The differences between eukaryotic and prokaryotic ribosomes can be exploited for the development of antibiotics (Table 29.4). For example, the antibiotic puromycin inhibits protein synthesis by causing nascent prokaryotic polypeptide chains to be released before their synthesis is completed. Puromycin is an analog of the terminal aminoacyl-adenosine part of aminoacyl-tRNA (Figure 29.34).

Table 29.4. Antibiotic inhibitors of protein synthesis.

Table 29.4

Antibiotic inhibitors of protein synthesis.

Figure 29.34. Antibiotic Action of Puromycin.

Figure 29.34

Antibiotic Action of Puromycin. Puromycin resembles the aminoacyl terminus of an aminoacyl-tRNA. Its amino group joins the carbonyl group of the growing polypeptide chain to form an adduct that dissociates from the ribosome. This adduct is stable because (more...)

It binds to the A site on the ribosome and inhibits the entry of aminoacyl-tRNA. Furthermore, puromycin contains an α-amino group. This amino group, like the one on aminoacyl-tRNA, forms a peptide bond with the carboxyl group of the growing peptide chain. The product, a peptide having a covalently attached puromycin residue at its carboxyl end, dissociates from the ribosome.

Streptomycin, a highly basic trisaccharide, interferes with the binding of formylmethionyl-tRNA to ribosomes and thereby prevents the correct initiation of protein synthesis. Other aminoglycoside antibiotics such as neomycin, kanamycin, and gentamycin interfere with the decoding site located near nucleotide 1492 in 16S rRNA of the 30S subunit (Section 29.3.9). Chloramphenicol acts by inhibiting peptidyl transferase activity. Erythromycin binds to the 50S subunit and blocks translocation. Finally, cyclohexamide blocks peptidyl transferase activity in eukaryotic ribosomes, making a useful laboratory tool for blocking protein synthesis in eukaryotic cells.

Image ch29fu14.jpg

29.5.2. Diphtheria Toxin Blocks Protein Synthesis in Eukaryotes by Inhibiting Translocation

Image caduceus.jpg Diphtheria was a major cause of death in childhood before the advent of effective immunization. The lethal effects of this disease are due mainly to a protein toxin produced by Corynebacterium diphtheriae, a bacterium that grows in the upper respiratory tract of an infected person. The gene that encodes the toxin comes from a lysogenic phage that is harbored by some strains of C. diphtheriae. A few micrograms of diphtheria toxin is usually lethal in an unimmunized person because it inhibits protein synthesis. The toxin is cleaved shortly after entering a target cell into a 21-kd A fragment and a 40-kd B fragment. The A fragment of the toxin catalyzes the covalent modification of an important component of the protein-synthesizing machinery, whereas the B fragment enables the A fragment to enter the cytosol of its target cell.

A single A fragment of the toxin in the cytosol can kill a cell. Why is it so lethal? The target of the A fragment is EF2, the elongation factor catalyzing translocation in eukaryotic protein synthesis. EF2 contains diphthamide, an unusual amino acid residue of unknown function that is formed by posttranslational modification of histidine. The A fragment catalyzes the transfer of the adenosine diphosphate ribose unit of NAD+ to a nitrogen atom of the diphthamide ring (Figure 29.35). This ADP-ribosylation of a single side chain of EF2 blocks its capacity to carry out translocation of the growing polypeptide chain. Protein synthesis ceases, accounting for the remarkable toxicity of diphtheria toxin.

Figure 29.35. Blocking of Translocation by Diphtheria Toxin.

Figure 29.35

Blocking of Translocation by Diphtheria Toxin. Diphtheria toxin blocks protein synthesis in eukaryotes by catalyzing the transfer of an ADP-ribose unit from NAD+ to diphthamide, a modified amino acid residue in elongation factor 2 (translocase). Diphthamide (more...)

By agreement with the publisher, this book is accessible by the search feature, but cannot be browsed.

Copyright © 2002, W. H. Freeman and Company.
Bookshelf ID: NBK22531


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