Chapter 32: Translation
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1 Chapter 32: Translation Voet & Voet: Pages (Parts of sections 1-3) Slide 1
2 Genetic code Translates the genetic information into functional proteins mrna is read in 5 to 3 direction Codons are base triplets representing one amino acid e.g. GCA = Ala Slide 2
3 Four features of the genetic code 1. Read in a particular frame, Starting point determines amino acid sequence! 2. Degenerate: 1 amino acid can be encoded by up to 6 codons Slide 3
4 Genetic code - Features 2 3. Non random: similar amino acids are encoded by similar codons Change of 1 base has only small effect on protein 4. Widespread, almost universal: same in most organisms from bacteria to human with very few exceptions Slide 4
5 trnas as adaptors trna anticodon can form base pairs with mrna codon At least 1 trna for 1 amino acid 3' end of trna is covalently linked to amino acid Aminoacyl-tRNA Slide 5
6 trna structure L-shaped!!! Secondary structure (base pairs): Cloverleaf structure Aminoacyl-end and anticodon are on opposite sites of the trna. Slide 6
7 Aminoacyl-tRNA Synthases One or more enzymes for each amino acid Specifically recognize trna & attach amino acid to its 3 end: 1. Activation of amino acid by forming aminoacyl-amp: 2. Transfer of amino acid to trna s 2 or 3 OH (ribose) by forming an ester bond with carboxyl group of amino acid 3. Proofreading: bonds to incorrect amino acids are hydrolyzed Slide 7
8 Wobble Hypothesis Problem: 61 sense codons versus ~ 40 different trnas Solution: 3rd codon position allows non Watson-Crick base pairs wobble base pairs Allowed wobble pairings: 5 anticodon 3 codon C G A U U A or G G U or C I U, C, or A Inosine 1 trna can recognize 1-3 codons Slide 8
9 Procaryotic Ribosome mrna trnas A-site: aminoacyl trna P-site: peptidyl trna E-site: exit deacylated trna Small Subunit (30S in prokaryotes) Large Subunit (50S in prokaryotes) Low-resolution structure from cryo-electron microscopy Slide 9
10 Ribosome structure High-resolution crystal structures available since 2000 Inner core of the ribosome is RNA Proteins 'decorate' the outer surface => The Ribosome is a Ribozyme! Slide 10
11 Overview: Translation Slide 11
12 Initiation of protein synthesis Start codon on mrna is usually AUG (Met) Requires initiation factors (IF1, IF2 & IF3) In bacteria further defined by: Shine-Dalgarno sequence which base-pairs with ribosomal RNA mrna: Start codon in P site Slide 12
13 Elongation cycle 1. Elongation factor Tu (EF-Tu) delivers aminoacyl-trna to the A-site 2. Peptide bond is formed between growing peptide chain and new amino acid Elongation factor G (EF-G) catalyzes movement of the ribosome along mrna (translocation) 3 Cycle can resume Slide 13
14 Elongation factor Tu EF-Tu as a G protein switch: EF-Tu-GTP binds aminoacyl-trna and brings it to the ribosome correct codon recognition triggers GTP hydrolysis by EF-Tu EF-Tu-GDP is recycled to EF-Tu-GTP by elongation factor Ts (EF-Ts) Slide 14
15 Peptide Bond Formation 1. Amino group of aminoacyltrna (A site) attacks ester bond between growing peptide chain and P-site trna 2. Peptide chain is transferred from P-site trna and to A-site trna (forms peptide bond) Catalyzed by ribosomal RNA, i.e. ribozyme new peptidyl-trna is in A site deacylated (uncharged) trna in P site Slide 15
16 Translocation Elongation factor G (EF-G) catalyzes Simultaneous movement of: deacylated trna from P to E site (then dissociates from ribosome) peptidyl-trna from A to P site mrna bound to the trnas EF-G also hydrolyzes GTP peptidyl-trna in the P site empty A site presenting next codon Slide 16
17 Translation Termination End of peptide chain indicated by stop codon in the A site Requires release factors (RF1, RF2, RF3, RRF) and EF-G - RF1/RF2 bind stop codon - RF3 catalyzes RF1/RF2 removal Ribosome Recycling: RRF/EF-G catalyzes removal of trna and mrna RF3 and EF-G are GTPases Slide 17
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