Translation and Protein Biosynthesis

Fig. 1.57. Model of the regulation of translation by insulin. Insulin ( and other growth factors) activates the Akt kinase pathway (see ch. 10), whose final result is the phosphorylation of 4E-BPl, a regulatory protein of translation initiation. The 4E-BP1 protein inactivates the initation factor eIF-4E by complex formation. eIE-4E is required, together with the proteins eIE-4A and p220, for the binding of the 40S subunit of the ribosome to the cap structure of the mRNA. If the 4E-BP1 protein becomes phosphorylated as a result of insulin-mediated activation of the PI3 kinase/Akt kinase cascade, then eIE-4E is liberated from the inactive eIP-4E 4E-BPl complex and protein biosynthesis can begin.

In all these events, the hydrogen-bonded Watson-Crick base pair is operative and is responsible for DNA reduplication, transcription, and translation. Since mispairing can occur, all these processes are checked for fidelity by several enzymes which can correct for errors [6521. At this and all other levels of DNA reduplication and protein biosynthesis, intermolecular hydrogen bonds between nucleic acids and between nucleic acids and proteins are responsible for recognition, interaction, and, finally, for information transfer. 

The beginning of secondary product formation is often directly coupled with the synthesis of the corresponding enzymes. One example is the biosynthesis of flavonoids in cell cultures of Petroselinum hortense (Fig. 7). Here the regulatory mechanisms were extensively investigated with phenylalanine ammonia-lyase (PAL) (D 22.2.1) and chalcone synthase (D 22.3.3), the key enzymes of the biosynthetic chain. Experiments with inhibitors of transcription and translation showed that the increase of enzyme activity depends on RNA and protein biosynthesis. Labeling experiments demonstrated that it is caused by an accelerated rate of enzyme synthesis. 

The important feature of this methodology is that fidelity of the amide bond formation is achieved by specific and template-directed coupling of the desired acyl transfer. Therefore, proper juxtaposition of peptidyl- and aminoacyl-oligonucleotides on a polynucleotide template control the direction of polypeptide synthesis. This is essentially the way that fidelity of translation in protein biosynthesis is taking place in natural systems. 

Cellular protein biosynthesis involves the following steps. One strand of double-stranded DNA serves as a template strand for the synthesis of a complementary single-stranded messenger ribonucleic acid (mRNA) in a process called transcription. This mRNA in turn serves as a template to direct the synthesis of the protein in a process called translation. The codons of the mRNA are read sequentially by transfer RNA (tRNA) molecules, which bind specifically to the mRNA via triplets of nucleotides that are complementary to the particular codon, called an anticodon. Protein synthesis occurs on a ribosome, a complex consisting of more than 50 different proteins and several stmctural RNA molecules, which moves along the mRNA and mediates the binding of the tRNA molecules and the formation of the nascent peptide chain. The tRNA molecule carries an activated form of the specific amino acid to the ribosome where it is added to the end of the growing peptide chain. There is at least one tRNA for each amino acid. 

Whereas DNA is mostly located in the nucleus of cells in higher organisms (with some also in mitochondria and in plant chloroplasts), RNA comes in three major and distinct forms, each of which plays a crucial role in protein biosynthesis in the cytoplasm. These are, respectively, ribosomal RNA (rRNA), which represents two-thirds of the mass of the ribosome, messenger RNA (mRNA), which encodes the information for the sequence of proteins, and transfer RNAs (tRNAs) which serve as adaptor molecules, allowing the 4-letter code of nucleic acids to be translated into the 20-letter code of proteins. These latter molecules contain a substantial number of modified bases, which are introduced enzymatically. 

Protein biosynthesis by eukaryotic ribosomes, which are larger and more complex than those of bacteria, is very similar in its basic outline to that of bacteria. The major difference is in the initiation of translation, which involves recognition of the 5 -cap on the mature mRNA, mentioned earlier, by the small ribosomal subunit. 

In eukaryotes, the cytoplasm, representing slightly more than 50% of the cell volume, is the most important cellular compartment. It is the central reaction space of the cell. This is where many important pathways of the intermediary metabolism take place—e.g., glycolysis, the pentose phosphate pathway, the majority of gluconeogenesis, and fatty acid synthesis. Protein biosynthesis (translation see p. 250) also takes place in the cytoplasm. By contrast, fatty acid degradation, the tricarboxylic acid cycle, and oxidative phosphorylation are located in the mitochondria (see p. 210). 

With all proteins, protein biosynthesis (Translation for details, see p. 250) starts on free ribosomes in the cytoplasm (1). Proteins that are exported out of the cell or into lyso-somes, and membrane proteins of the ER and the plasma membrane, carry a signal peptide for the ER at their N-terminus. This is a section of 15-60 amino acids in which one or two strongly basic residues (Lys, Arg) near the N-terminus are followed by a strongly hydro-phobic sequence of 10-15 residues (see p. 228). 

To prevent incorrect folding of the growing protein during protein biosynthesis, chaperones (see B) in the lumen of the rER bind to the peptide chain and stabilize it until translation has been completed. Binding protein (BiP) is an important chaperone in the ER. 

Like amino acid activation (see p. 248), protein biosynthesis (translation) takes piace in the cytopiasm. it is cataiyzed by compiex nucieoprotein particies, the ribosomes, and mainiy requires GTP to cover its energy requirements. 

Induced lignification is blocked by treatments which inhibit protein synthesis (10), and it seems likely therefore that the increases in activities observed for enzymes involved in lignin biosynthesis (7,8) depend on enhanced translation, and probably enhanced transcription of the appropriate genes, as has been shown for the phytoalexin response of some plants (27). However, the molecular biology of the response in wheat awaits investigation. 

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