Amino acids elongation factors

Ono, Y., Skoultchi, A., Klein, A. and Lengyel, P. (1968) Peptide chain elongation discrimination against the initiator transfer RNA by microbiol amino-acid polymerization factors. Nature, 220,1304-1307. 

Increased protein synthesis Increased amino acid uptake/increased translation of mRNA Akt-mediated stimulation of system A amino acid transporter and stimulation of mRNA-translation through activation of p70S6kinase and elongation initiation factor 4 (elF4). Possible involvement of atypical PKCs... 

Elongation is a cycUc process on the ribosome in which one amino acid at a time is added to the nascent peptide chain. The peptide sequence is determined by the order of the codons in the mRNA. Elongation involves several steps catalyzed by proteins called elongation factors (EFs). These steps are (1) binding of aminoacyl-tRNA to the A site, (2) peptide bond formation, and (3) translocation. 

The now deacylated tRNA is attached by its anticodon to the P site at one end and by the open GGA tail to an exit (E) site on the large ribosomal subunit (Figure 38-8). At this point, elongation factor 2 (EE2) binds to and displaces the peptidyl tRNA from the A site to the P site. In turn, the deacylated tRNA is on the E site, from which it leaves the ribosome. The EF2-GTP complex is hydrolyzed to EF2-GDP, effectively moving the mRNA forward by one codon and leaving the A site open for occupancy by another ternary complex of amino acid tRNA-EFlA-GTP and another cycle of elongation. 

Figure 38-8. Diagrammatic representation of the peptide elongation process of protein synthesis. The small circles labeled n - 1, n, n -I-1, etc, represent the amino acid residues of the newly formed protein molecule. EFIA and EF2 represent elongation factors 1 and 2, respectively. The peptidyl-tRNA and aminoacyl-tRNA sites on the ribosome are represented by P site and A site, respectively.

The principal drawbacks to the technique are its labor-intensive nature and low protein yields, which in the best cases reach only a milligram or less. Competition with release factors at the amber stop codon often results in truncated protein as the primary product [16, 17]. Suppression efficiency rates are also affected by the character of the amino acid, which determines whether it is a good substrate for the ribosome and protein elongation factors. In addition, context effects variously ascribed to the influence of neighboring mRNA... 

The ribosome can carry two aminoacyl-tRNAs simultaneously. In the chain elongation stage, the growing polypeptide is carried on one of these tRNAs. The chain is transferred to the second tRNA, which adds its amino acid to the growing peptide, and displaces the first tRNA. The ribosome then moves one codon along the mRNA to allow the next to be read. Termination of protein synthesis involves the release of the completed polypeptide, expulsion of the last tRNA, and dissociation of the ribosome from the mRNA. This is signaled by specific termination codons (UAA, UAG, or UGA) in the mRNA and requires the participation of various release factors. 

An understanding of protein synthesis, the most complex biosynthetic process, has been one of the greatest challenges in biochemistry. Eukaryotic protein synthesis involves more than 70 different ribosomal proteins 20 or more enzymes to activate the amino acid precursors a dozen or more auxiliary enzymes and other protein factors for the initiation, elongation, and termination of polypeptides perhaps 100 additional enzymes for the final processing of different proteins and 40 or more kinds of transfer and ribosomal RNAs. Overall, almost 300 different macromolecules cooperate to synthesize polypeptides. Many of these macromolecules are organized into the complex three-dimensional structure of the ribosome. 

The third stage of protein synthesis is elongation. Again, our initial focus is on bacterial cells. Elongation requires (1) the initiation complex described above, (2) aminoacyl-tRNAs, (3) a set of three soluble cytosolic proteins called elongation factors (EF-Tu, EF-Ts, and EF-G in bacteria), and (4) GTP. Cells use three steps to add each amino acid residue, and the steps are repeated as many times as there are residues to be added. 

A large number of components are required for the synthesis of a polypeptide chain. These include all the amino acids that are found in the finished product, the mRNA to be translated, tRNAs, functional ribosomes, energy sources, and enzymes, as well as protein factors needed for initiation, elongation, and termination of the polypeptide chain. 

Requirements include all the amino acids that eventually appear in the finished protein, at least one specific type of tRNA for each amino acid, one aminoacyl-tRNA synthetase for each amino acid, the mRNA coding for the protein to be synthesized, fully competent ribosomes, protein factors needed for initiation, elongation, and termination of protein synthesis, and ATP and GTP as energy sources. 

Initiation (Figs. 29-10 and 29-11), elongation (Fig. 29-12), and termination are three distinct steps in the synthesis of a protein. A variety of specialized proteins are required for each stage of synthesis. Their sequential interaction with ribosomes can be viewed as a means of ensuring an orderly sequence of steps in the synthesis cycle. The rate of protein formation will depend upon the concentrations of amino acids, tRNAs, protein factors, numbers of ribosomes, and kinetic constants. The formation of specific proteins can also be inhibited by translational repressors, proteins that compete with ribosomes for binding to target mRNAs.287... 

Overview of reactions in protein synthesis. (aab aa2, aa3 = amino acids l, 2, 3.) Protein synthesis requires transfer RNAs for each amino acid, ribosomes, messenger RNA, and a number of dissociable protein factors in addition to ATP, GTP, and divalent cations. First the transfer RNAs become charged with amino acids, then the initiation complex is formed. Peptide synthesis does not start until the second aminoacyl tRNA becomes bound to the ribosome. Elongation reactions involve peptide bond formation, dissociation of the discharged tRNA, and translocation. The elongation process is repeated many times until the termination codon is reached. Termination is marked by the dissociation of the messenger RNA... 

At the conclusion of the initiation process, the ribosome is poised to translate the reading frame associated with the initiator codon. The translation of the contiguous codons in mRNA is accomplished by the sequential repetition of three reactions with each amino acid. These three reactions of elongation are similar in both prokaryotic and eukaryotic systems two of them require nonribosomal proteins known as elongation factors (EF). Interestingly, the actual formation of the peptide bond does not require a factor and is the only reaction of protein synthesis catalyzed by the ribosome itself. 

In the next step a second tRNA molecule, associated with a protein named elongation factor Tu (EF-Tu), comes in carrying the appropriate amino acid and binds to the ribosome. A peptide bond forms between the two amino acids held on the ribosome. The first tRNA molecule now has lost its amino acid, and the two covalently bonded amino acid residues are linked to the second tRNA. At this point the first... 

Fig. 9. Comparison of amino acid sequences of bovine mouse rat a, a (partial), retinal rod and cone cells < bacterial elongation factor EFTu and c-Ha-ras, aligned to maximize matching. This leads to the definition of an aavp of 3% amino acids (numbers above sequences). Boxes surround identical or conserved residues, (c) and (p), residues ADP-ribosylated by CTX and PTX, respectively. —A—, — C—, —E— and —G—, sequences known to be intimately involved in guanine nucleotide binding by EFTu (From Refs. 175 and 176).

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