Transcript Document
Announcements 1. Reading Ch. 15: skim btm 425-426 2. Look over problems Ch. 15: 5, 6, 7. Review of Last Lecture I. RNA processing in eukaryotes II. Translation of mRNA into protein - tRNA and ribosomes III. Three steps of translation IV. First evidence that proteins are important to heredity Outline of Lecture 26 Regulating gene expression in prokaryotes In E. coli, some proteins are found at 5-10 copies/cell; others are found at 100,00 copies. How does a cell regulate the levels or amounts of different proteins at different times - in response to the environment? I. Lactose metabolism - the lac operon, an inducible system II. Trp operon, a repressible system Background on gene organization •Operon: a series of gene coding regions (usually products with related functions) under the control of a single gene regulatory unit. •Allows rapid response to changing environmental conditions. 3 structural genes lac operon Bacterial Strategy • If glucose is present, – then use glucose as a carbon source. • If glucose is not present, and if lactose is present, – then use lactose (indirectly) as carbon source. Levels of enzymes needed to use lactose as carbon source increase dramatically when lactose is present; enzymes are inducible and lactose is the inducer. The -galactosidase Enzyme lacY gene -> Permease (entry of lactose into cell) lacZ gene-> lacA -> transacetylase lac operon, a polycistronic mRNA Could the cell make varying amounts of lacZ and lacA RNA? Genes involved in Lactose Metabolism Repressor Regulator Protein Enzyme lactose ?- removes transporter toxic byproducts Normally binds to Operator when lactose is absent, but when lactose is present, comes off. The Operon Model - components pol R L Lac operon when no lactose is present pol R R pol pol R R pol pol R R L L pol pol R pol R pol Learning Check pol R L Will transcription and translation of Z, Y,and A enzymes occur? What would happen if a wild-type copy of I was added? What happens when cell has both lactose and glucose? glucose lactose b-galactosidase + - - + + - - - - - + + How does cell prevent breaking lactose down into glucose and galactose? Catabolite Activating Protein (CAP) Catabolite Repression of lac Operon -/+ Glucose Therefore, for maximal transcription, repressor must be bound by repressor and CAP must bind CAP-binding site Goal: efficiency, don’t waste energy converting lactose, when glucose available Glucose Inhibits Formation of cAMP from ATP glucose lac operon regulatory binding sites CAP site Glu cAMP lactose + - - + - + promoter lac rep CAP-cAMP - + operator lac coding lac rep CAP-cAMP - + + RNA polymerase mRNA Polymerase binds here CAP DNA Repressor 3-D structure of Repressor bound to Operator and CAP bound to Promoter Lewis et al. (1996) Science 271:1247 glucose absent lactose absent II. Tryptophan Operon It makes sense for E. coli to synthesize the enzymes to make the amino acid tryptophan ONLY when: 1. The tryptophan concentration is low AND 2. The tryptophan-charged tRNA is low Tryptophan Operon - Repressor Binds when tryptophan is present mRNA leader sequence involved in Attenuation - the cast of characters Translation Event controls Transcription Event !?!? High TryptophantRNA -->> Terminates Transcription Low TryptophantRNA -->> Transcription Continues