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Part II of Molecular Bio Review A review of Laneuville's section of Molecular Biology, pertaining to Prokaryotic transcription, translation and protein synthesis. It includes lecture notes, objectives and their answers.

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University of Ottawa
Jos�e Coutu

MOLECULAR BIO REVIEW II TRANSCRIPTION IN PROKARYOTES LectureTranscription Reaction is DNA to RNA Double stranded DNA is the information archive which is transcribed into RNA molecules which are expendable information carriersonly one of the 2 DNA strands is transcribed the antisense strand35the sequence of the DNA sense strand 53 corresponds to the transcribed RNA sequence Ts instead of UsCOMPONENT AND KEY TERMS FOR TRANSCRIPTION DefinitionsTranscription unit DNA segment that is transcribedIts transcription begins when RNA polymerase binds to thepromoter of the gene at the region of the 5 end of the coding strand 5 end of the gene promoter is close to the first nt transcribed forming the RNA approx 60 bp TSS transcription start site location where transcription startsGene Region of the genome transcribed into a mRNA or rRNA or tRNAMessenger RNA contains information for proteins sequences NOT changed in prokaryotes but it is in eukaryotesGenes and RNAeach organism has a different size of genome and a different number of genes therefore gene density varies as the complexity of the organism increases so does the size of the genome but not quite the same linear relationship as with number of genesOrganism Size of genome Number of genes 6Prokaryotes 47 x 10 4 000 7Yeast 15 x 10 5 000 to 10 000 8Drosophila 14 x 10 20 000 9Human 33 x 10 20 000 to 25 000 the majority of the RNA in the cell is rRNA then tRNAmRNA accounts for 3 to 5of the total RNA in the cellhowever 32 of the cells RNA capacity is toward mRNA this is because mRNA is unstable while rRNA and tRNA are more stable Typeat eqb Capacity of synthesis rRNA 83 58 tRNA 14 10 mRNA 35 32 snRNA 1 1 snoRNA 1 1 Other RNA 1 1RNA Polymerasecatalyzes the transcription reactionIn prokaryotes only one kind of RNA polymerase catalyzes the synthesis of RNA whereas in eukaryotes there are three types of polymeraseshowever the same biochemical reaction is catalyzed in both prokaryotes and eukaryotes they also have similar 3D structureHowever regulation of transcription is more complex in eukaryotes Eukaryotes RNA polymerases Type Localisation Cellular Transcripts I Nucleolus rRNA 58s rRNA 18S rRNA 28S II Nucleoplasm mRNA snRNA III Nucleoplasm tRNA rRNA 5S RNA polymerase in prokaryotes is 465 Da and are about 7 000 molecules of RNA polymerase in one cellThe core enzyme of RNA polymerase has two alpha subunits one beta subunit and one beta prime subunit The holoenzyme has this plus a sigma factor 22Subunit Size Gene Function Alpha x2 40 kDA rpoAEnzyme assemblyPromoter recognitionBinding to activators Beta 155 kDa rpoB Catalytic site creates phosphodiester bondsBeta 160 kDa rpoCCatalytic site creates phosphodiester bonds Sigma 3290 kDa rpoDPromoter specificityCTDC terminal domainHTHhelix turn helix motif of sigma which binds to the nucleic acidSome AntibioticsRifampicin tuberculosis inhibits the initial step of RNA synthesis by interfering with the formation of the firstphosphodiester link of the RNA chainThis means elongation is blockedHowever if elongation hasalready started rifampicin does not affect the system because the DNARNA hybrid prevents binding ofantibioticActinomycin D binds DNA and prevents the movement of RNA polymerase affects elongationTHE TRANSCRIPTION REACTION 1 Promoter Recognition RNA polymerase searches and binds to the promoterthe strength of a promoter is defined by the frequency at which RNA polymerase initiates transcription the sigma factor helps identify the promoter most commonly strong promoters have the sequence TTGACA at 35 position 35 nt upstream fromposition 1 and TATAAT at 10 note that there are different sequences identified by different sigmafactorsSIGMA reduces the affinity of RNA polymerase for sequences that are not promoters increases the affinity of RNA pol for promoter sequences the alpha helix of sigma protein plays role in binding RNA pol to TATAAT sequence
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