Cellular location: In eukaryotic cells, the ribosomes either are free in the cytosol or are in close association with the endoplasmic reticulum (which is then known as the rough endoplasmic reticulum, or RER). RER-associated ribosomes are responsible for synthesizing proteins (including glycoproteins; see p. 166) that are to be exported from the cell, incorporated into membranes, or imported into lysosomes (see p. 169 for an overview of the latter process). Cytosolic ribosomes synthesize proteins required in the cytosol itself or destined for the nucleus, mitochondria, or peroxisomes. [Note: Mitochondria contain their own ribosomes (55S) and their own unique, circular DNA. Most mitochondrial proteins, however, are encoded by nuclear DNA, synthesized completely in the cytosol, and then targeted to mitochondria.]
F. Protein factors
Initiation, elongation, and termination (or, release) factors are required for polypeptide synthesis. Some of these protein factors perform a catalytic function, whereas others appear to stabilize the synthetic machinery. [Note: A number of the factors are small, cytosolic G proteins and thus are active when bound to guanosine triphosphate (GTP) and inactive when bound to guanosine diphosphate (GDP). See p. 95 for a discussion of the membrane-associated G proteins.]
G. Energy sources
Cleavage of four high-energy bonds (see p. 73) is required for the addition of one amino acid to the growing polypeptide chain: two from ATP in the aminoacyl-tRNA synthetase reaction, one in the removal of PPi and one in the subsequent hydrolysis of the PPi, to two Pi by pyrophosphatase, and two from GTP, one for binding the aminoacyl-tRNA to the A site and one for the translocation step (see Fig. 32.13, p. 457). [Note: Additional ATP and GTP molecules are required for initiation in eukaryotes, whereas an additional GTP molecule is required for termination in both eukaryotes and prokaryotes.] Translation, then, is a major consumer of energy.
IV. CODON RECOGNITION BY TRANSFER RNA
Correct pairing of the codon in the mRNA with the anticodon of the tRNA is essential for accurate translation (see Fig. 32.6). Most tRNA (isoaccepting tRNA) recognize more than one codon for a given amino acid.
A. Antiparallel binding between codon and anticodon
Binding of the tRNA anticodon to the mRNA codon follows the rules of complementary and antiparallel binding, that is, the mRNA codon is read 5′→3′ by an anticodon pairing in the opposite (3′→5′) orientation (Fig. 32.9). [Note: Nucleotide sequences are always written in the 5′ to 3′ direction unless otherwise noted. Two nucleotide sequences orient in an antiparallel manner.]
B. Wobble hypothesis
The mechanism by which a tRNA can recognize more than one codon for a specific amino acid is described by the wobble hypothesis, which states that codon–anticodon pairing follows the traditional Watson-Crick rules (G pairs with C and A pairs with U) for the first two bases of the codon but can be less stringent for the last base. The base at the 5′-end of the anticodon (the first base of the anticodon) is not as spatially defined as the other two bases. Movement of that first base allows nontraditional base-pairing with the 3′-base of the codon (the last base of the codon). This movement is called wobble and allows a single tRNA to recognize more than one codon. Examples of these flexible pairings are shown in Figure 32.9. The result of wobble is that 61 tRNA species are not required to read the 61 codons that code for amino acids.
V. STEPS IN TRANSLATION