PowerPoint PresentationSlide 2Slide 3Slide 4Slide 5Slide 6Slide 7Slide 8Slide 9Slide 10Slide 11Slide 12Slide 13Slide 14Slide 15Slide 16Slide 17Peptide Bond Synthesis is Catalyzed by the 23S RNA not by a protein enzyme – the Ribosome is a Ribozyme (peptidyl trnasferase)!Slide 19Slide 20Slide 21Slide 22Slide 23Slide 24Slide 25Slide 26Slide 27Slide 28Slide 29Slide 30Slide 31Slide 32Slide 33Slide 34Slide 35Slide 36Slide 37Slide 38Slide 39Slide 40Energy Requirements for TranslationSlide 42Slide 43Translation in EukaryotesSlide 45Slide 46Slide 47Slide 48Slide 49Slide 50Slide 51Slide 52Slide 53Slide 54Slide 55Mutation and Mutagenesis Some TerminologyPoint Mutations - Consequences for ProteinsInsertion/Deletion MutationsSlide 59Lecture 25 - Chapters 39-40TranslationLecture 25 - Chapters 39-40Translationhttp://bioap.wikispaces.com/Ch+17+Collaborationhttp://bioap.wikispaces.com/Ch+17+CollaborationribosomespeptidesmRNA1OutlineOutline•Ribosomal subunits•Initiation complex formation•Elongation•Termination•Prokaryotic vs. eukaryotic protein synthesis•Synthesis of secreted proteins•Regulation of protein synthesis•Mutations•Ribosomal subunits•Initiation complex formation•Elongation•Termination•Prokaryotic vs. eukaryotic protein synthesis•Synthesis of secreted proteins•Regulation of protein synthesis•Mutations223S RNA5S RNA16S RNA21 S protein34 L proteinThe Prokaryotic RibosomeThe Prokaryotic Ribosome(60S)(60S)(80S)(80S)(40S)(40S)In eukaryotesIn eukaryotes3The ribosome has three binding sites for tRNAsThe ribosome has three binding sites for tRNAsA = aminoacyl-tRNA siteP = peptidyl-tRNA siteE = exiting tRNA siteA = aminoacyl-tRNA siteP = peptidyl-tRNA siteE = exiting tRNA siteDecoding CenterDecoding CenterPeptidyl transferase CenterPeptidyl transferase Center4Initiation in ProkaryotesInitiation in Prokaryotes1. Ribosome Binding Site on mRNA (sometimes called a Shine-Dalgarno sequence) base pairs with 16S rRNA of 30S subunit2. Initiation codon - AUG3. Initiation factors (IF): IF-1, IF-2(GTP), IF-34. Initiator tRNA - fMet-tRNAiMet1. Ribosome Binding Site on mRNA (sometimes called a Shine-Dalgarno sequence) base pairs with 16S rRNA of 30S subunit2. Initiation codon - AUG3. Initiation factors (IF): IF-1, IF-2(GTP), IF-34. Initiator tRNA - fMet-tRNAiMet5Initiation Site (purine-rich)Shine-Dalgarno Sequences (Ribosome Binding Site)Initiation Site (purine-rich)Shine-Dalgarno Sequences (Ribosome Binding Site)UAC6Table 30-7, p. 967E.coli Initiation FactorsE.coli Initiation FactorsIF1: Binds to 30S A site and prevents tRNA bindingIF2: G-protein that binds fMet-tRNA(initiator tRNA), which docks at P site of 30SIF3: Binds to 30S E site; prevents 50S bindingIF1: Binds to 30S A site and prevents tRNA bindingIF2: G-protein that binds fMet-tRNA(initiator tRNA), which docks at P site of 30SIF3: Binds to 30S E site; prevents 50S binding7Initiator tRNAInitiator tRNA8Formation of the 30S initiation complexFormation of the 30S initiation complexIF3IF3RBSRBS9Formation of the 70S initiation complexFormation of the 70S initiation complex50S50S30S30SmRNAPPAAEE10E. coli Elongation Factors (EF)E. coli Elongation Factors (EF)EF-Tu: G protein that binds aminoacyl-tRNA and delivers it to the A siteEF-Ts: Guanine-nucleotide exchange factor (GEF) that replaces GDP on EF-Tu with GTPEF-G: G protein that promotes translocation of mRNAEF-Tu: G protein that binds aminoacyl-tRNA and delivers it to the A siteEF-Ts: Guanine-nucleotide exchange factor (GEF) that replaces GDP on EF-Tu with GTPEF-G: G protein that promotes translocation of mRNA11EF-Tu brings charged tRNA to appropriate codonEF-Tu brings charged tRNA to appropriate codon1270S initiation complex70S initiation complexEF-Tu brings charged tRNA to appropriate codonEF-Tu brings charged tRNA to appropriate codonGTPGTPEF-Tu-GTPGDPEF-Tu-GDPCharged tRNA13GTPGTPEF-Tu-GTPGDPEF-Tu-GDPCharged tRNACharged tRNATo ribosomeGTPGTPEF-Tu-GTPEF-TEF-TGDPGTPGTPResetting EF-Tu by EF-TsResetting EF-Tu by EF-TsEF-TsIs the GTPExchange FactorEF-TsIs the GTPExchange Factor143’3’2’2’Covalent linkage of amino acid to cognate tRNACovalent linkage of amino acid to cognate tRNA5’5’3’3’15Peptide bond formationPeptide bond formation16Polypeptide ChainPolypeptide ChainN’N’ C’ C’a.a. 2a.a. 2a.a. 3a.a. 3a.a. 4a.a. 4a.a. 5a.a. 5NH3+a.a. 1a.a. 1COO-17Peptide Bond Synthesis is Catalyzed by the 23S RNA not by a protein enzyme – the Ribosome is a Ribozyme (peptidyl trnasferase)!Peptide Bond Synthesis is Catalyzed by the 23S RNA not by a protein enzyme – the Ribosome is a Ribozyme (peptidyl trnasferase)!The 50S subunit. 3’ end of P-site tRNA is red, 3’ end of A site tRNA is green. There is no protein near, only RNA.18Peptide bond formation by peptidyl transferasePeptide bond formation by peptidyl transferaseEP50S50S30S30SEPEP12 3codonscodonsN’-Decoding CenterDecoding CenterPeptidyl transferase CenterPeptidyl transferase Center19Translocation by EF-GTranslocation by EF-GEPE PEPEP20EF-G Mimics EF-Tu-tRNAEF-G Mimics EF-Tu-tRNAEF-Tu-tRNA EF-GProtein21An Active RibosomeAn Active RibosomeA= aminoacylP = peptidylE= exitA= aminoacylP = peptidylE= exitDecoding CenterDecoding CenterPeptidyl transferase CenterPeptidyl transferase Center22Polypeptide-chain growth Polypeptide-chain growthN’N’N’C’tRNA1tRNA2235’GCU AUC GGG UUG AAG CUCUUCaaCGACCCaaAACaaRibosome MovementUAGE P AfMet-aa-aa-aa-50S50STranslation ElongationTranslation Elongation30S245’GCU AUC GGG UUG AAG CUCUUCaaCCCAACaaRibosome MovementUAGE P AfMet-aa-aa-aa-aa-50STranslation ElongationTranslation Elongation30S25E P A5’GCU AUC GGG UUG AAG CUCUUCaaCCC AACaaRibosome MovementfMet-aa-aa-aa-aa-50STranslation ElongationTranslation Elongation30S26E P A5’GCU AUC GGG UUG AAG CUCGAGaaAACaaRibosome MovementfMet-aa-aa-aa-aa-UUCaaCCCTranslation ElongationTranslation Elongation30S50S27Ribosome MovementE P A5’GCU AUC GGG UUG AAG CUCGAGaaCCCAACfMet-aa-aa-aa-aa-aa-UUCaaTranslation ElongationTranslation Elongation30S50S28Ribosome MovementE P A5’GCU AUC GGG UUG AAG CUCGAGaaCCCAACfMet-aa-aa-aa-aa-aa-UUCaaTranslation ElongationTranslation Elongation30S50S29Ribosome MovementE P A5’GCU AUC GGG UUG AAG CUCGAGaaAACfMet-aa-aa-aa-aa-aa-UUCaaTranslation ElongationTranslation Elongation30S50S30Ribosome MovementE P A5’GCU AUC GGG UUG AAG CUCGAGaaAACfMet-aa-aa-aa-aa-aa-aa-UUC30S50S31Ribosome MovementE P A5’GCU AUC GGG UUG AAG CUC
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