Tuesday 9 13 prokaryotes Rna folds up into these stems and loops Small rnas are involved in splicing and regulation Transcription making an rna molecule from the dna template made in 5 to 3 direction means template is running 3 to 5 The nontemplate strand has the same sequence as the rna Rna polymerase causes the promoter to start synthesizing Terminal genes stop the rna polymerase so it doesn t go all the way around the genome Rna polymerases don t need a primer they can start on a bare strand of dna Rna polymerase can recognize the gene and will start transcribing Can alrign a bunch of genes and form a consensus sequence c g they use when they are 50 50 First base transcribed is labeled as 1 The closer the gene matches the consensus the more highly the gene expression is going to be Core rna polymerase 2 alpha and 2 beta subunits the sigma factor is an additional protein that recognizes the consensus sequence and increasing the binding to those particular sites Gives specificity to consensus sequence To stop transcription one mechianism is rho independent mechanism Relies ona particular sequence at the end of the gene a pair of inverted repeats that form a step loop structure in the transcript AU bonds less tightly bound to make rna not very well attached to the template Other mechanism rho dependent requires a rho protein Rho protein chases the polymerase and dislodges it Rna pauses from the hair pin loop and gives time for the rho protein to catch up Have 4 polymerases to transcribe different classes of genes Promoters are bigger and more complex Core promotor has a 25 box Upstream also have recognizable consenuses sequences Show if a gene is going to be highly expressed Transcription complex tata binding protein binds 2 core promotor together with tfiid are the first to bind and recognize core promotor Must recrut all the other elements of rna polymerase II Footprinting experiments piece of dna mix some of this dna with proteins of choice dnaseI chews up all of the dna that is not protected by a protein wrapped around it leaves over a set of fragments bands indicate where dnaseI cut the bases 0 with slash through it As we increase the amount of protein there are no cuts showing that protein is protecting a part of the sequence Seeing footprints of where protein is nteracting dna and protecting it from degredation Termination not as tightly regulated as prokaryotes mrna as its being transcribed downstream will have a consensus sequence that will initate protein cleavage to release mrna but rna polymerase is still running along Eukaryotes The rat1 endonuclease chases the polymerase down and will dislode it so it doesn t continue transcribing to the next gene Chapter 14 RNA molecules and rna processing Basic prok messanger rna start of transcription usually first part isn t translated into protein then a consensus sequence called shine dalgarno sequence which is where the ribosome recognizes where it shoud start translating Start and stop codon Then after there is also an uncoded region Euk mrnas have 3 different modifications 5 cap a guanine that is put on backwards 5 to 5 linkage After the 5 is added the bases get metyl groups The 3 end also gets modified with a poly a tail A long stream of A s that protects the 3 end and is important in regulation of translation Central dogma isn t true in euks Introns intervening regions Mrna gets chopped down to eliminate introns Called splicing Splicing the exons together and in the process removing the introns Gt ag rule first 2 and last 2 bases in an intron Look for that structure to find the boundary of the introns 90 of ur genes at alternitavely splices 9 15 lecture protein Transfer RNA there are some introns that can splice out themselves first evidence of rna acting as an enzyme Gives some insight into the early stages of life of the RNA world could be the entire genetic system all by themselves Cleaving is when you cut a piece on the end off splicing is cutting out something in the middle and putting the pieces back together Substitution editing substitutes one base for another to produce a different Transfer rnas have a clover leaf structure 4 arms one is an acceptor arm at 3 end ACC is the amino acid acceptor Anticodon arm interacts with the mrna to help get the right bases Trna is transcribed as a regular rna but modifers come in to change some of the bases Tnra has a precursor that has to be cleaved off and an intron that has to be spliced out brings you to the basic trna then thers there addition of the ACC added to the end of the 3 and some modification of the bases Bacterical rubisomes have 2 subunits to make a 70s ribosome that includes rna components and protein components rRna Start with double stranded rna that gets processed and associated with a protein that somehow inhibits the message in your body miRNAs are important to development and gene expression rna transcribed in nucleus processed mostly in the nucleus 3 regions found in all mrna 5 untranslated region 3 untranslated region start codon stop codon Be able to draw a prokaryotic mrna and eukaryotic one Intron in between region with no genetic info Modificatiosn on an mrna Chapter 15 Genetic Code and Translation Series of arguments and experiments that led to the genetic code 1 Logic have 4 nucleotides and need 20 amino acids Must be a triplet code 64 different codes which is more than enough to code for 20 different amino acids 2 Insertional Mutagens a set of experiments drugs proflavin which are mutagins and insert themselves into the helix acts as a nucleotide causes intsertion mutations and deletions THE BIG DOG ATE THE FAT CAT experminets that an be associated with a certain amino acid triplet found that whenever they had an insertion of one base it was a mutant changed the entire code if they combined a 1 mutiation with a 1 mutation then it becomes normal wild type 3 or 3 were also wild type Code is not overlapping 3 Homopolymers and cell free translation system they know the structure of the code but they don t know what letters code for which amino acid 4 Random heteropolymers 5 Ordered heteropolymers and triplet binding assays Technology enabled them to synthesize ordered triplets triplet binding assay takes charged trnas that already are loaded with amino acids and radio label just one of those amino acids triplets ribosomes Ribosome has the triplet inside and one trna recognizes that triplet and latches onto th ribosome The tna that isbinding on the