H O

FIGURE 8.06 Charging Transfer RNA with the Amino Acid

This two-step procedure begins (A) by attachment of the amino acid to adenosine monophosphate (AMP) to give aminoacyl-AMP or aminoacyl-adenylate. This involves splitting ATP and the release of inorganic pyrophosphate. Then, in the second step (B), the amino acid is transferred to the hydroxyl group of the ribose at the 3'-end of the tRNA, yielding AMP as a byproduct.

FIGURE 8.07 Glutamine tRNA Bound to its Aminoacyl tRNA Synthetase

Structure of glutaminyl-tRNA synthetase bound to tRNA(Gln) and a glutaminyl adenylate analog. The analog is in orange and is shown in a space-filling representation. The tRNA is depicted in dark blue. Domains of the enzyme are color-coded as follows: active-site Rossman fold, green; acceptor-end binding domain, yellow; connecting helical subdomain, red; proximal beta-barrel, light blue; distal beta-barrel, orange. The image was made in PyMol by John Perona, Department of Chemistry and Biochemistry, University of California at Santa Barbara.

FIGURE 8.08 Components of a Bacterial Ribosome

The ribosome is composed of 30S and 50S subunits. These in turn are composed of ribosomal RNA and numerous proteins. The 30S subunit is built from 16S rRNA together with 21 proteins. The 50S subunit contains 5S and 23S ribosomal RNA plus 34 proteins.

FIGURE 8.08 Components of a Bacterial Ribosome

The ribosome is composed of 30S and 50S subunits. These in turn are composed of ribosomal RNA and numerous proteins. The 30S subunit is built from 16S rRNA together with 21 proteins. The 50S subunit contains 5S and 23S ribosomal RNA plus 34 proteins.

FIGURE 8.09 3-D Structure of a Ribosome by EM

This structure was deduced from negatively stained electron microscope images of a bacterial 70S ribosome.

FIGURE 8.09 3-D Structure of a Ribosome by EM

This structure was deduced from negatively stained electron microscope images of a bacterial 70S ribosome.

FIGURE 8.10 3-D Structure of a Ribosome by X-Ray

Views of the structure of the Thermus thermophilus 70S ribosome. A. B. C and D are successive 90° rotations about the vertical axis. (A) view from the back of the 30S subunit. H, head; P, platform; N, neck; B, body. (B) view from the right-hand side, showing the subunit interface cavity, with the

30S subunit on the left and the 50S on the right. The anticodon arm of the A-tRNA (gold) is visible in the interface cavity. (C) View from the back of the 50S subunit. EC, the end of the polypeptide exit channel. (D) View from the left-hand side, with the 50S subunit on the left and the 30S on the right. The anticodon arm of the E-tRNA (red) is partly visible. The different molecular components are colored for identification: cyan, 16S rRNA; grey, 23S rRNA; light blue, 5S rRNA; dark blue, 30S proteins; magenta, 50S proteins. From Yusupov et al., Crystal Structure of the Ribosome at 5.5 A Resolution. Science 292 (2001) 883-96.

FIGURE 8.11 Secondary Structure of a Ribosomal RNA

The 16S rRNA from the small ribosomal subunit of E. coli is complex with extensive secondary structure, forming loops and stems. Red indicates regions of base-pairing.

FIGURE 8.11 Secondary Structure of a Ribosomal RNA

The 16S rRNA from the small ribosomal subunit of E. coli is complex with extensive secondary structure, forming loops and stems. Red indicates regions of base-pairing.

m

The peptide bond linking amino acids in the growing protein is made by the largest ribosomal RNA, which acts as a ribozyme.

These rRNA molecules have highly defined secondary structures with many stems and loops (Fig. 8.11). Although it was originally believed to have a largely structural role, recent work indicates that the rRNA is responsible for most of the critical reactions of protein synthesis. In particular, the 23S rRNA of the large subunit is a ribozyme that catalyzes the synthesis of the peptide bonds between the amino acids; i.e., it is the peptidyl transferase. Indeed, X-ray crystallography of the 50S subunit has shown that no ribosomal proteins are close enough to the catalytic center to take part in the reaction. Alteration by mutation of the catalytic residues in typical ribozymes either abolishes activity completely or reduces it by many-fold. However, the peptidyl-transferase center of 23S rRNA behaves in an atypical manner. Alteration of A2451 or G2447 (E. coli numbering) did not greatly reduce catalytic activity, although these residues are present in the catalytic center. These results suggest that the ribosome does not operate via direct chemical catalysis. Rather, the ribosome acts by correctly positioning the two substrates. The activated aminoacyl-tRNA then reacts spontaneously with the end of the growing polypeptide chain.

Since the genetic code is read in groups of three bases, any nucleic acid sequence contains three possible reading frames.

Three Possible Reading Frames Exist

Before mRNA is translated into protein, the issue of reading frames must be dealt with. The bases of mRNA are read off in groups of three, with each codon corresponding to one amino acid. How are the base sequences divided into codons? For any given nucleotide sequence there are three alternatives, depending on what is considered the start. Consider the following sequence:

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