Lecture 13 Infectious disease antibiotics is not profitable not enough people with infectious diseases Primase lays down an RNA primer then DNA polymerase extends the primer 5 to 3 o 3 HYDROXYL end o 5 phosphate end o When the DNA polymerase abuts the next primer than ligase fixes the nicks comes in and binds the two parts together o Each time you add a base you basically burn 1 ATP each time you fix a nick burn one ATP o Not going to be asked names of most of the enzymes in the DNA synthesis Bacteria tend to have one origin of replication per circular chromosome at terminal ending the daughter DNA molecule separates from the original Eukaryotic chromosomes have many replication origins Nucleic acid polymerases DNA or RNA add to the 3 OH of the growing chain 3 OH is used as a nucleophile nucleotide triphosphates comes in and diphosphate is broken off and the phosphate is added to the chain The central dogma o DNA RNA Protein o Direction of information travel in a cell o DNA top nucleic acids have 5 and 3 ends by convention the 5 end is always on the left side This line is the genotype where all the genes are o RNA line o Protein bottom line Proteins always have an amino terminal N and a carboxyl terminal C N side is on the left o The PHENOTYPE comes AFTER the proteins the protein is synthesized it acts on some type of substrate or catalyze some type of reaction and then its phenotype is expressed o DNA to RNA transcription o RNA to Protein translation o DNA to RNA Nucleic acid deoxyribose nucleic acid to nucleic acid ribose nucleic acid The difference being the hydroxyl at the 2 position o Promoter T s have been replaced by U s in the mRNA Picture of the different lines notice the o Found at the 5 end of the genes upstream of genes o Initiation of transcription o RNAP RNA polymerase multi subunit enzyme recognizes DNA sequence at promoters and forms hydrogen bonds o Sigma factors required for initiation used to move the RNAP past the promoter and start synthesizing the RNA catalysis of RNA polymerization o The promoter has a 35 sequence and a 10 sequence TATA of also called the Pribnow box Usually occur 10 an 35 away from where transcription starts o Promoters can move to the left or to the right however the template strand will switch depending on which way it is moving always at to the 3 end Terminator where the RNA polymerase is given signal to stop during transcription Translation o RBS the ribosomal binding site on the mRNA upstream of where the translation start site is signal for the ribosome to actually bind o AUG universal start codon for almost every gene codes for methionine amino acid o Stop codons UGA UAA UAG to not code for any amino acids o Multiple ribosomes can work on one mRNA at the same time o Ribosomes will start binding and synthesizing peptides BEFORE the RNAP is even done synthesizing the RNA because this is happening in the cytosol in the bacteria Question which of these can be found on mRNA o 5 end 3 end ribosome binding sites translation start translation stop Question which can be found on DNA o 5 end 3 end promoters transcription stop sites origin of replication Question which can be found on the protein o N terminal and C terminal ends EXAM Q be able to label the three lines be able to label promoters start sites stop sites 5 end 3 end amino and carboxyl terminals LOOK OVER THE 3 LINE DIAGRAM PICTUES ALL OF THEM Bacteria has a way of synthesizing multiple proteins from 1 RNA transcript does this through operons o Operons have one promoter for multiple genes o For each potential protein on the RNA there is an RBS start codon stop codon o This is how the lac operon works Eukaryotes do it completely differently no operons you find introns non coding regions cut out before the RNA is translated into proteins o This all happens in the nucleus mature RNA then comes out of the nucleus o Leaving introns in a proteins can result in a mal formed protein can often cause Allows for simultaneous transcription and translation because in cytosol diseases can become cancerous Compartmentalization o Prokaryotes no compartmentalization o Eukaryotes Compartmentalization Regulation of gene expression In nucleus DNA replication Transcription In cytoplasm translation Principles of gene regulation o Allows organism to adapt to changing environments o Allows organism to conserve energy o Allows organism to switch developmental growth processes o Ex if your living in an environment without lactose you don t want to waste energy making B galactosidase but you want to have the gene available to be synthesized when you run into an environment that does have lactose present o Ex 2 sporulation highly regulated do not want to make spores under high energy conditions wait until nutrients are low and toxic by products are building up then you start to express the gene involved in sporulation o Transcription regulate how much mRNA of a gene you make Constitutive genes that are ON all the time unregulated Inducible only get turned on when environmental conditions warrant them being turned on Repressible gene can be turned off when it is not needed anymore o Translation not as commonly used as a method of regulation Problem with controlling at translation is that you ve already made the mRNA so you ve already wasted energy Eukaryotes do this more often than prokaryotes prokaryotes do it even less because they do not have compartmentalization so if they already started making the mRNA they probably already started making the protein o Allosteric sties another form of regulation You have chemicals that work on enzymes and make it so the enzyme cannot catalyze a reaction this is obviously after translation Often used in metabolic pathways where the end product can inhibit previous steps stop enzyme from making more product o Figures of the different types of controls o Bacteria usually regulate which enzymes are available in the cell by transcriptional control Ex in the lac operon there is a lac operon repressor when this enzyme is bound there is no transcription of the genes found downstream of the RNA promoter stoping it from moving downward towards the gene Inducer for example in this case could be lactose when the repressor is induced it comes off the gene and the genes will be transcribed Glucose lactose graph see figure slide 50 o First a cell will use glucose then when it uses the glucose in the cell up it will use lactose o The line is not exactly smooth the small plateau in the