Ribosome base-pairing with mRNA

Figure 13.3 The process of protein synthesis on the ribosome. The strand of mRNA is shown associated with the small subunit of the ribosome. The aminoacyl-tRNA molecules are shown associated with the large subunit of the ribosome and base-paired with mRNA codons. A peptide bond is in the process of formation between the two associated amino acids, extending the growing polypeptide chain by one unit. On the left, a tRNA is shown leaving the ribosome, having donated its amino acid to the growing chain. On the right, an aminoacyl-tRNA molecule is shown entering the ribosome. It is next in line to contribute its amino acid to that chain.

Figure 29.22. Transfer RNA-Binding Sites. (A) Three tRNA-binding sites are present on the 70S ribosome. They are called the A (for aminoacyl), P (for peptidyl), and E (for exit) sites. Each tRNA molecule contacts both the SOS and the 50S subunit. (B) The tRNA molecules in sites A and P are base paired with mRNA.

Stage 2 Initiation The mRNA bearing the code for the polypeptide to be made binds to the smaller of two ri-bosomal subunits and to the initiating aminoacyl-tRNA. The large ribosomal subunit then binds to form an initiation complex. The initiating aminoacyl-tRNA base-pairs with the mRNA codon AUG that signals the beginning of the polypeptide. This process, which requires GTP, is promoted by cytosolic proteins called initiation factors. 

Initiation The components of the translation system are assembled, and mRNA associates with the small ribosomal subunit. The process requires initiation factors. In prokaryotes,a purine-rich region (the Shine-Dalgarno sequence) of the mRNA base-pairs with a complementary sequence on 16S rRNA, resulting in the positioning of the mRNA so that translation can begin. The 5 -cap on eukaryotic mRNA is used to position that structure on the ribosome. The initiation codon is 5 -AUG-3. ... 

The function of the leader sequence is to fine tune expression of the trp operon based on the availability of tryptophan inside the cell. It does this as follows. The leader sequence contains four regions (Fig. 2, numbered 1-4) that can form a variety of base-paired stem-loop ( hairpin ) secondary structures. Now consider the two extreme situations the presence or absence of tryptophan. Attenuation depends on the fact that, in bacteria, ribosomes attach to mRNA as it is being synthesized and so translation starts even before transcription of the whole mRNA is complete. When tryptophan is abundant (Fig. 2a), ribosomes bind to the trp polycistronic mRNA that is being transcribed and begin to translate the leader sequence. Now, the two trp codons for the leader peptide lie within sequence 1, and the translational Stop codon (see Topic HI) lies between sequence 1 and 2. During translation, the ribosomes follow very closely behind the RNA polymerase and synthesize the leader peptide, with translation stopping eventually between sequences 1 and 2. At this point, the position of the ribosome prevents sequence 2 from interacting with sequence 3. Instead sequence 3 base-pairs with sequence 4 to form a 3 4 stem loop which acts as a transcription terminator. Therefore, when tryptophan is present, further transcription of the trp operon is prevented. If, however, tryptophan is in short supply (Fig. 2b), the ribosome will pause at the two trp codons contained within sequence 1. This leaves sequence 2 free to base pair with sequence 3 to form a 2 3 structure (also called the anti-terminator),... 

In eukaryotes, methionyl-tRNAjMet binds to the small ribosomal subunit. The 5 cap of the mRNA binds to the small subunit and the first AUG codon base-pairs with the anticodon on the methionyl-tRNA 61. 

Proteins are synthesized in the amino-to-carboxyl direction, and mRNA is translated in the 5 —>3 direction. The start signal on prokaryotic mRNA is usually AUG preceded by a purine-rich sequence that can base-pair with 16 5 rRNA. In prokaryotes, transcription and translation are closely coupled. Several ribosomes can simultaneously translate an mRNA, forming a polysome. 

P-site The peptidyl site on the ribosome to which Met-tRNA is brought to base pair with the mRNA sequence AUG. It is also the site to which the peptidyl RNA is moved in a process known as translocation following the formation of a new peptide bond. 

The transcription(copying in mRNA reciprocal code) and translation(using the mRNA to place amino acids in the proper sequence) of the DNA code to protein products proceeds through a complicated series of steps first involving the formation of a messenger RNA (mRNA) having a base sequence complementary to that of the parent DNA strand. The mRNA then associates with ribosomal RNA (rRNA) - protein complexes. Transfer RNA (tRNA) molecules bearing specific amino acids are then base-paired with the mRNA. Many enzyme-catalyzed reactions later, a protein product is formed. 

The genetic information in DNA is converted into the linear sequence of amino acids in polypeptides in a two-phase process. During transcription, RNA molecules are synthesized from a DNA strand through complementary base pairing between the bases in DNA and the bases in free ribonucleoside triphosphate molecules. During the second phase, called translation, mRNA molecules bind to ribosomes that are composed of rRNA and ribosomal proteins. Transfer RNA-aminoacyl complexes position their amino acid cargo in the catalytic site within the ribosome in a process that involves complementary base pairing between the mRNA codons and tRNA anticodons. Once the amino acids are correctly positioned within the catalytic site, a peptide bond is formed. After the mRNA molecule moves relative to the ribosome, a new codon enters the ribosome s catalytic site and base pairs with the appropriate anticodon on another aminoacyl-tRNA complex. After a stop codon in the mRNA enters the catalytic site, the newly formed polypeptide is released from the ribosome. 

Translation begins as each ribosome binds an mRNA molecule and proceeds to convert its base sequence into a polymer of amino acids linked by peptide bonds. Each amino acid is specified by a code word, called a codon, that consists of three sequential bases. The actual transfer of information occurs when each mRNA codon interacts and forms complementary base pairs with a three-base sequence in a transfer RNA (tRNA) molecule called an anticodon. 

Initiation. Translation begins with initiation, when the small ribosomal subunit binds an mRNA. The anticodon of a specific tRNA, referred to as an initiator tRNA, then base pairs with the initiation codon AUG. Initiation ends as the large ribosomal subunit combines with the small subunit. There are two sites on the complete ribosome for codon-anticodon interactions the P (peptidyl) site (now occupied by the enitiator tRNA) and the A (aminoacyl) site. In both prokaryotes and eukaryotes, mRNAs are read simultaneously by numerous ribosomes. An mRNA with several ribosomes bound to it is referred to as a polysome. In actively growing prokaryotes, for example, the ribosomes attached to an mRNA molecule may be separated from each other by as few as 80 nucleotides. 

During Initiation, the ribosomal subunits assemble near the translation start site in an mRNA molecule with the tRNA carrying the amino-terminal methionine (Met-tRNAi ) base-paired with the start codon (Figure 4-25). 

As shown in Figure 26.10, a purine-rich sequence, called the Shine-Dalgarno sequence, is centered 10 nucleotides upstream of the AUG codon in most bacterial mRNAs. It was discovered that the 30 end of bacterial 16S rRNA has a complementary sequence which can base pair with the Shine-Dalgarno sequence to position the 30S subunit relative to the initiator AUG codon. Ribosome positioning in eukaryotes is more difficult to predict, but there are two general rules for identifying the most likely eukaryotic AUG initiator codon (1) it is often the first AUG encountered by the ribosome after it binds the 50 mRNA cap and "scans" in the 30 direction and (2) there are preferred contexts for the initiator AUG which have been defined by the consensus sequence GCCPuCCAUGG. 

Translation is accomplished by the anticodon loop of tRNA forming base pairs with the codon of mRNA in ribosomes Stop codons act to stop translation... 

EF-Tu will bind to any aminoacylated tRNA other than tRNA the initiator tRNA (step c. Fig. 29-12), and carry it to the ribosome (step d), where it binds into the A site. There it is selected if it forms a proper base pair with the mRNA codon in the A site or is rejected if it does not. This decoding process involves both an initial step and a proofreading step. The aminoacyl-tRNA binds both to the decoding site in the 16S RNA and to the peptidyltransferase site in the 23S RNA. (See discussions on p. 1687.) The decoding site is on the platform at the upper end of helix 44 (Fig. 29-2). Nucleotide G1401 plays a crucial roie.375 vVhen one of the isoacceptor species of E. coU tRNA is irradiated with ultraviolet light, the... 

Elongation of the polypeptide involves three steps (a) binding of an aminoacyl-tRNA to the A site on the ribosome where it base-pairs with the second codon on the mRNA (b) formation of a peptide bond between the first and second amino acids and (c) translocation, movement of the mRNA relative to the ribosome, so that the third mRNA codon moves into the A site. These three elongation steps are repeated until a termination codon aligns with the site on the ribosome where the next aminoacyl-tRNA would normally bind. Release factors bind instead, causing the completed protein to be released from the ribosome. 

A purine-rich mRNA sequence, three to nine nucleotides long (called the Shine-Dalgamo sequence), which is centered about 10 nucleotides upstream of (to the 5 side of) the start codon, base-pairs with a sequence of complementary nucleotides near the 3 end of the 16S rRNA of the 30S ribosomal subunit. This interaction plus the association of fMet-tRNAf with the AUG in the P site of the ribosome sets the mRNA reading frame. 

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