Translation: part II
Explain how 16S rRNA Plays an Active Role in Translation
• 16S rRNA plays an active role in the functions of the 30S subunit. - It interacts directly with mRNA (3' domain), with the 50S subunit, and with the anticodons of tRNAs in the P and A sites.
Describe how 30s Ribosomal RNA Pervades Both Ribosomal Subunits
• 30S subunit. • 5' domain forms the 30S body; the central domain forms the platform; the 3' major domain forms the head; and the 3' minor domain runs along the intersubunit face for the 30S subunit. • 4 domains.
Describe how 50s Ribosomal RNA Pervades Both Ribosomal Subunits
• 50S subunit. • 3 main features. • Central protuberance formed by 5S rRNA. • Stalk formed by L7 protein. • Notch: interaction with 30S subunit.
Describe the Elongation Factor Tu Loads Aminoacyl-tRNA into the A Site
• EF-Tu ( an elongation factor) is a monomeric G protein whose active form (bound to GTP) binds to aminoacyl-tRNA. • The EF-Tu-GTP-aminoacyl-tRNA complex binds to the ribosome A site. • Ef-Tu is released in the GDP- bound form. Ef-Tu is only associated with the ribosome during the process of aminoacyl- tRNA entry. • Ef-Ts (guanine nucleotide excahnge factor) is required to mediate the replacement of GDP by GTP
Describe how (interaction between 16S and 23S rRNAs) Ribosomal RNA Pervades Both Ribosomal Subunits
• Each rRNA has several distinct domains that fold independently. • Virtually all ribosomal proteins are in contact with rRNA. • rRNAs provide the main contacts between the ribosome subunits (interaction between 16S and 23S rRNAs). • Most of the contacts between ribosomal subunits are made between the 16S and 23S rRNAs. • Interactions between 16S and 23S rRNA are localised (1 domain in 23S rRNA and 2 domains in 16S rRNA)
Describe how Ribosomes Have Several Active Centers
• Interactions involving rRNA are a key part of ribosome function. • The environment of the tRNA- binding sites is largely determined by rRNA.
Explain how 23S rRNA Has Peptidyl Transferase Activity
• Peptidyl transferase activity resides exclusively in the 23S rRNA. • Peptide bond formation requires an attack by the amino group of one amino acid on the carboxyl group of another amino acid. • This requires acid-base catalysis in which an H atom is transferred to a basic residue.
Describe how the Translocation Moves the Ribosome
• Ribosomal translocation moves the mRNA through the ribosome by three bases. • Translocation moves deacylated tRNA into the E site and peptidyl-tRNA into the P site, and empties the A site. • The hybrid state model proposes that translocation occurs in two stages, in which the 50S moves relative to the 30S, and then the 30S moves along mRNA to restore the original conformation.
Explain how The Polypeptide Chain Is Transferred to Aminoacyl-tRNA
• The 50S subunit has peptidyl transferase activity as provided by an rRNA ribozyme. • The ribosome remains in place while the polypeptide chain is elongated. • The nascent polypeptide chain is transferred from peptidyl-tRNA in the P site to aminoacyl-tRNA in the A site. • Peptide bond synthesis generates deacylated tRNA in the P site and peptidyl-tRNA in the A site.
Describe the process of terminate codons
• The class 1 release factors (RF1 and RF2) respond to specific termination codons and hydrolyze the polypeptide- tRNA linkage. • The class 1 release factors are assisted by class 2 release factors (RF3) that depend on GTP. • The mechanism is similar in bacteria (which have two types of class 1 release factors) and eukaryotes (which have only one class 1 release factor). • Final step. RRF - ribosome recycling factor, acts together with EF-G in a reaction that hydrolyzes GTP.
Describe Three Codons Terminate Translation
• The stop codons UAA (ochre), UAG (amber), and UGA (sometimes called opal) terminate translation. • In bacteria, they are used most often with relative frequencies UAA>UGA>UAG.
Explain the longation Factors Bind Alternately to the Ribosome
• Translocation requires EF-G, whose structure resembles the aminoacyl- tRNA-EF-Tu-GTP complex. • Binding of EF-Tu and EF-G to the ribosome is mutually exclusive. • Translocation requires GTP hydrolysis, which triggers a change in EF-G, which in turn triggers a change in ribosome structure.
Explain why Termination Codons Are Recognized by Protein Factors
• Two stages are involved in ending translation. The termination reaction itself (RF1 and RF2, release of the protein chain from the last tRNA) and the post-termination reaction (RF3, release of the tRNA. mRNA and dissociation of the ribosome). • Termination codons are recognized by protein class 1 release factors, not by aminoacyl- tRNAs. • RF1 - The bacterial release factor that recognizes UAA and UAG as signals to terminate polypeptide translation. • RF2 - The bacterial release factor that recognizes UAA and UGA as signals to terminate polypeptide translation. • RF1 and RF2 recognise termination codons and activate the ribosome to hydrolyse the peptidyl tRNA. • RF3 - A polypeptide translation termination factor related to the elongation factor EF-G. - RF3 interacts with RF1 or RF2. - It functions to release the factors RF1 or RF2 from the ribosome when they act to terminate polypeptide translation. RRF is the ribosome recycling factor
Describe how Translation Occurs by Initiation, Elongation, and Termination
• initiation - The stages of translation up to synthesis of the first peptide bond of the polypeptide. • elongation - The stage of translation in which the polypeptide chain is extended by the addition of individual subunits. • termination - A separate reaction that ends translation by stopping the addition of subunits and (typically) causing disassembly of the synthetic apparatus.
