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Lecture Material for Test 3.docx

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Lovaye Kajiura

Lecture Material for Test 3 October 13 Template-directed synthesis -similar to DNA but don’t need primer -where you start, you pick -diff component of holoenzyme -sigma locates what you’re interested in -holoenzyme has to move along, binds loosely to DNA -sigma factor helps located promoter -promoter sequence similar to concensus sequence -same holoenzyme with diff sigma factors locate diff genes -follow direction of TATAAT -in virus DNA, genes are overlapped -change of one base can cause many changes in diff proteins -in cell DNA, has a lot of DNA available -change in one base (third base) will not affect much since DNA is redundant -form open complex, let go of sigma factor -core enzyme binds very tightly to DNA -becomes a very processive enzyme -initiation, elongation, termination -2 types of termination -one type forms hairpin, one single strand DNA -has ternary complex will be disrupted by any formation of bulky structure -other type: Rho factor -pulls RNA off complex -transcription in eukaryotes -has 3 RNAP for diff kinds of genes -RNAP II – polymerase that is responsible for making all mRNA -responsible gene -want to be able to control -RNAPs equally complicated -inhibitors -if there are any molecules that go in and bind to complex -don’t need to know too much about this -actinomycin D – not specific, just intercalates and bind tightly to DNA -polymerase won’t be able to find promoter -crystallography gives electron density -gives where atoms are position -need good resolution to see actual atoms -drug designs based on very fundamental diff btw diff processes -antibiotics look for ribosomes of bacteria -inhibit translation in bacteria -since bacterial translation diff enough the human translation -specificity important in terms of drug therapy, toxicity – how unspecific a drug is -therapy for cancer is unspecific -eukaryotes have diff types of promoter -usually related to TATA but not all genes have TATA box -some genes have BRE sequence -at diff regions, has diff concensus sequence -instead of having one single sigma factor, we have a large number of protein called general transcription factors -sigma factor uses in prokaryote system to help identify gene that you want since e.coli doesn’t have a lot of genes -in eukaryotes, have many genes and many diff cell types, need many more combinations how to search through genes -TBP (TATA binding protein) – transcription factor for polymerase II TFII contains 2 proteins: TBP and TAF -binds to minor groove (nonspecific for bases) -eukaryotic system involves many transcription factors Postranscriptional Processing -after making RNA molecules (primary transcript) -needs to be processed before it can be delivered to fxtional molecule -no processing prokaryotes -add on 5’ caps and poly A tails at 3’ in eukaryotes -lengthens lifetime of mRNA -lot of nucleases in cell that digest single strands easily -want to protect both ends -cap for other machinery to look for -mRNA from eukaryotic has many intervening sequence – introns -exons expressed -splice introns out -consensus sequence at exon-intro junction -tranesterification -RNA less stable b/c has 2’ hydroxide -energy of moving almost the same -same gene can give many diff isoforms -diff splicing rxn gives diff molecules from same gene -by removing diff portion of gene (treat parts of intron) get diff products -alternative splicing – variation in processing of RNA molecule -intro can splice itself out from RNA -RNA can form very stable tertiary structures -DNA molecule always has complementary strand; always in duplex -RNA does not have complementary strand, has sequences that are complementary to itself and form many diff unique structures -tRNA when it gets transcribed, a lot of regions removed, some added on -some bases are modified -forms mature RNA after mods -many genes, genes needed are transcribed -locate gene, decided which one to be trancsirbed -initiate, transcription factor comes off -termination sequence (can depend or not on Rho factor) -in eukaryote, no termination sequence or Rho factor -RNA molecule keeps being transcribed until some nuclease cuts it off -don’t need to kno all the splicing processes since they are still being researched October 15 -need telomerase to add to ends of DNA so it does not die upon replication -if telomerase is triggered in every cell, cell would live longer but not healthier -theres a reason they’re not expressed in certain cells -mRNA used to make many copies of proteins according to w.e instruction you have -mRNA starts translation before transcription finishes -occurs in same compartment in prokaryotic cells -in eukaryotic cells, mRNA made in nucleus and translation occurs in cytosol -want to express protein when needed -transcribed DNA when needed by sending out sigma factors -mRNA has to be there only for the needed amt of time -control -the more secondary structures you have on mRNA, the more stable; less susceptible to nucleases -TATAAT on mRNA sequence -where do you start making mRNA -approx 10 base upstream -need to kno amino acids -adaptor molecule: tRNA -folds into clover leaf structure -unique 3d structure -one end recognizes codon, other an holds aa -if we have 2 bases, would have only 16 aa (4 ) -one insertion/deletion will cause frameshift -changing bases not as detrimental -b/c there are 64 codons and only 20 aa - redundancy -64 codon -anticodons pairs
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