PowerPoint PresentationSlide 2rRNA is made by Pol I in the NucleolusSlide 4What does “S” mean? “Svedberg Units”Sedimentation CoefficientSlide 6Slide 7tRNA contains modified bases.Slide 9Eukaryotic mRNASlide 11Slide 12Slide 135’ mRNA CappingSlide 15Poly(A) tailsSlide 17Slide 18Alternative SplicingSlide 20Slide 21The Splicing ReactionSmall nuclear ribonucleoproteins (snRNPs)Slide 24Slide 25Slide 26Slide 27Slide 28Fully Processed mRNA is Specifically Transported to the CytoplasmSlide 30The Genetic CodeSlide 32Characteristics of the Genetic CodemRNA is “read out” in 3 nucleotide units – codons. Each codon specifies either a particular amino acid or a translational stop.Evidence for the CodeSlide 36Slide 37Slide 38Slide 39Slide 40Slide 41The Code is DegenerateWobble HypothesisSlide 44Modified Bases in tRNAs Include Inosine in the Wobble PositionSlide 46Slide 47Slide 48Slide 49Slide 50Slide 51Slide 52The Second Genetic CodeA Class I Aminoacyl-tRNA SynthetaseSlide 55Distinguishing Amino AcidsSlide 57Slide 58Slide 59Slide 60Slide 61Lecture 24 - Chapters 38 & 39Eukaryotic RNA processing and the genetic codeLecture 24 - Chapters 38 & 39Eukaryotic RNA processing and the genetic code“Now that we have all this useful information, it would be nice to do something with it.”From the Unix Programmer’s ManualThe Rosetta Stone, inscribed in 196 B.C.OutlineOutline•Ribosomal RNA processing•tRNA processing•mRNA processing•RNA binding proteins•The genetic code•Wobble hypothesis•tRNA structure•tRNA anticodon binding to codon•Aminoacyl-tRNA synthetase•Ribosomal RNA processing•tRNA processing•mRNA processing•RNA binding proteins•The genetic code•Wobble hypothesis•tRNA structure•tRNA anticodon binding to codon•Aminoacyl-tRNA synthetaserRNA is made by Pol I in the NucleolusrRNA is made by Pol I in the Nucleolus•rRNA genes are arranged as tandem repeats.•In humans, there are approximately 400 rRNA genes in 5 clusters (on different chromsomes).•Localized transcription and processing take place in the nucleolus.rRNA GenerRNA GenerRNA GeneTranscribed SpacerTranscribedSpacerEtcEtcThe processing of eukaryotic pre-rRNAThe processing of eukaryotic pre-rRNArRNA Processingis NOT SplicingWhat does “S” mean? “Svedberg Units”Sedimentation CoefficientWhat does “S” mean? “Svedberg Units”Sedimentation CoefficientThe functional ribosome is the 70S which is composed of one 30S subunit and one 50S subunit.The functional ribosome is the 70S which is composed of one 30S subunit and one 50S subunit.40S 60S 80S In eukaryotesActive formEukaryotic Ribosomal RNA (rRNA)Eukaryotic Ribosomal RNA (rRNA)4.22 x 106Dribosome30S30S50S50S70S70Sprokaryotesprokaryotes/16S/16S/23S + 5S/23S + 5S/21/21/34/34Transfer RNA precursor processingTransfer RNA precursor processingTrailertRNA contains modified bases.tRNA contains modified bases.Two modified U’s found in tRNA. There are many other modified bases (over 100 known). Modifications are introduced after the RNA is transcribed (post-transcriptionally).UDEukaryotic mRNA ProcessingEukaryotic mRNA ProcessingPre-mRNAPre-mRNAmRNAmRNAEukaryotic mRNAEukaryotic mRNAInitiation codonInitiation codonMonocistronicMonocistronicCoding RegionPoly(A) tail5’Cap5’ Untranslated Region5’ Untranslated Region3’ Untranslated Region3’ Untranslated Region7-methylguanylate(m7G) 7-methylguanylate(m7G)HHCaps 0Caps 0Capping the 5 endCapping the 5 endCapping the 5 endCapping the 5 end 7-methylguanylate(m7G) 7-methylguanylate(m7G)HCaps 1Caps 1Capping the 5 endCapping the 5 end 7-methylguanylate(m7G) 7-methylguanylate(m7G)Caps 2Caps 25’ mRNA Capping5’ mRNA Capping•Occurs immediately after the 5’ end is made – before transcription is completed.•Functions–Important for splicing–Protection from RNases – Increased stability–Enhances efficiency of Translation – Cap Binding Protein–Required for efficient transport from the nucleusPolyadenylation of a primary transcript.Polyadenylation of a primary transcript.PABP = polyA-binding proteinPABP = polyA-binding proteinPoly(A) tailsPoly(A) tailsMost eukaryotic mature mRNAs have ~100-200 adenine residues attached at their 3’- end, called the 3ʹ-poly A tail•Functions•Nuclear export•Translation•Stability of mRNA.Prokaryotic mRNAProkaryotic mRNAPolycistronic mRNAPolycistronic mRNADNADNAProtein AProtein AProtein BProtein BProtein cProtein cRBSRBSRBSRBSRBSRBSBiochemistry: The Molecular Basis of Life, 5/e Alternate Splicing of Monocistronic pre- mRNAsAlternate Splicing of Monocistronic pre- mRNAsMonocistronic: only one coding regionMonocistronic: only one coding regionReleased exon not translated but degradedReleased exon not translated but degradedReleased exon not translated but degradedReleased exon not translated but degraded++Protein A’Protein A’’mRNA A’mRNA A’’Alternative SplicingAlternative Splicing•The human genome contains only about 20,000 genes.•The average gene has 7 introns.•It is estimated that 95-100% of these genes produce 2 or more different mRNAs as a result of alternative splicing.•Some genes have enough introns that they could theoretically produce over 30,000 different proteins. That’s more proteins from 1 gene than there are genes.Splice sitesSplice sitesInvariant in eukaryotesPolypyrimidine tract5’--3’Phosphoester bond being attackedPhosphoester bond being attackedNew Phosphoester bondNew Phosphoester bondalcoholalcoholesteresterThe Splicing ReactionThe Splicing ReactionExon 2Exon 2Exon 1Exon 11st transesterification1st transesterification2nd transesterification2nd transesterificationSmall nuclear ribonucleoproteins (snRNPs)Small nuclear ribonucleoproteins (snRNPs)Splicing – part 1Splicing – part 1Splicing – part 2: Splicing – part 2: +U2 and U6 snRNAs form the catalytic center of splicesomes U2 and U6 snRNAs form the catalytic center of splicesomes 5’ 2’ phosphodiester bond5’ 2’ phosphodiester bondTranscription and processing of mRNA are coupledTranscription and processing of mRNA are coupled1234RNA editingRNA editingliverliverSmall intestineSmall intestinehttp://www.sciencephoto.com/media/210444/enlargehttp://www.sciencephoto.com/media/210444/enlargeRibosomal subunitsRibosomal subunitscytoplasmcytoplasmnucleusnucleusNuclear pore channelsNuclear pore channelsNucleocytoplasmic transport of ribosomal subunitsNucleocytoplasmic transport of ribosomal subunitsA set of bound
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