BIOL-L211 Lecture 32 Outline of Last Lecture I. Recruiting Ribosome to mRNA transcriptII. Initiation of Prokaryotic TranslationIII. Initiation of Eukaryotic TranslationIV. Eukaryotic TranslationOutline of Current Lecture I. Translation ElongationII. Translation TerminationCurrent LectureTranslation III. Translation ElongationA. No significant difference (that we will cover) between eukaryotes and prokaryotesB. Major Steps:1. Codon in A site determines which tRNA is recruited and loaded2. Amino acids are linked by a peptide bond3. Polypeptide from tRNA in P site shifts to the charged tRNA in the A site4. Ribosome is transferred toward 3' end of mRNAa. A new codon is then in the A siteb. Empty tRNA is ejected via the E site5. Repeat. Until stop codon.C. Elongation Factor EF-TuThese notes represent a detailed interpretation of the professor’s lecture. GradeBuddy is best used as a supplement to your own notes, not as a substitute.1. EF-Tu: elongation factor responsible for binding and bringing charged tRNAs to A site2. EF-Tu-GTP ensures tRNA binds at appropriate time3. GTP hydrolyzes when tRNA binds correct codon, which causes the release of EF-Tu-GDP4. Accommodation: rotation of charged tRNA into the peptidyl transferase centerD. Peptide Bond Formation1. Incorrect codon-anticodon pairing causes:a. Reduced EF-Tu GTPase activityb. No accommodation2. For Peptide Bond to form, correctly charged tRNA must be in A site AND have gone through accommodation3. Bond between amino acids is called a peptide bond and is formed through a dehydration reaction (loss of water)4. rRNA in large subunit catalyzes peptide bond formation5. Protein directionality: N-terminus to C-terminusE. Elongation Factor EF-G1. Translocation: Movement of tRNA from A site to P site, and movement of uncharged tRNA to the E site (shift down one codon)2. After peptide chain is moved to A site tRNA, tRNAs shift to hybrid state3. Hybrid State: state in which 3' ends of the tRNAs are in new positions in the large ribosomal subunit while anticodons are still bound to the codons in their initial locations in the small ribosomal subunit4. EF-G-GTP: binds ribosome and uses GTP hydrolysis to GDP to change its conformationa. tRNAs are then fully transferred to respective new sitesb. EF-G dissociates (note: in both EF-Tu and EF-G, GTP hydrolysis causes dissociation)c. A site is left open againII. Translation TerminationA. Stop codon in A site triggers termination (no corresponding tRNAs)B. Class I release factors (RFs): bind stop codon (as opposed to tRNA), and cause the release of the polypeptide chain from the tRNA in the P site1. Prokaryotes: Have DIFFERENT Class I RFs to recognize each of the three stop codons2. Eukaryotes: Have only one Class I to recognize all three stop codonsC. Class II release factors: exchange GDP for GTP causing class I RFs to leaveD. GTP hydrolysis causes dissociation of class II RFsE. Ribosome Recycling:1. tRNAs and mRNA are removed from ribosome2. Ribosome breaks apart into its large and small subunits (IF3 stimulates ribosome dissociation)3. Ribosome Recycling Factor (RRF): enters A site (which has stop codon)a. EF-G binds RRF and moves it to the P siteb. RRF in the P site causes release of tRNAs in BOTH P and E sites4. RRF and EF-G released from ribosome5. mRNA releasedF. Interpreting Stop Codons1. Exon Junction Complex: (result of splicing) proteins that bind around 20 nucleotides upstream from the exon-exon junctions, and guide the ribosome as it proceedsa. EJC contact = continue translatingb. Translation terminates when it hits the stop codon with no EJC proteins downstream G. Nonsense-mediated mRNA Decay1. Degradation of mRNA with a stop codon in the improper location (early) in eukaryotes (as splicing only occurs in eukaryotes)2. If there are EJC proteins following a stop codon, proteins come and degrade mRNA3. If there are no EJC proteins following a stop codon, translation
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