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Chapter 16 Control of Gene Expression Topics to discuss • DNA binding proteins • Prokaryotic gene regulation – Lac operon – Trp operon • Eukaryotic gene regulation Today’s lecture 6 groups of DNA-binding regulatory proteins have been identified Prokaryotic Gene Regulation Nutritional enviroment and a bug’s growth! • Operons • Operons provide coordinate expression A model operon Now let’s look at 2 groups of operons Negative inducible operons Negative repressible operons Negative inducible operon INDUCIBLE Turned on repressor Now let’s look at 2 specific operons • The Lactose operon • The Tryptophan operon Francois Jacob and Jacques Monod 1961 • Studied lactose metabolism in prokayotes J. Monod Won the Nobel Prize in 1965 Sources of prokaryotic energy • 1. Disaccharide • Lactose • 2. Monosaccharide • Glucose Building the lac operon Lac Operon and Proposed Arrangement with control I gene I P O Z Not technically part of the “operon” Y A Negative inducible operon Conditions: No glucose but lactose is present Lactose Metabolism Requires Coordination of all these genes! Laboratory Exercise: Mutational Analysis of the lac operon Try out your ability to reason through the questions. • Predict if you expect to obtain the proteins from the lacZ and lacY genes IF certain mutations are present in the operon. The next three slides are the “key” to the mutations. Mutational Key: • • • Any + means wildtype. Any – means mutant. Two other mutants: “Oc and Is” • I- cells synthesize full levels in the presence or absence of inducerTranscription Turned ON. Inhibitor cannot bind to DNA • P- the DNA promoter mutation : cannot bind RNA polymerase Transcription OFF • Is super-suppressors can bind DNA but not inducer. DNA and Transcription Turned OFF. • Oc is an DNA mutation: repressor cannot bind. Transcription ON. Examples I s=the Super-repressor Allolactose CANNOT BIND VERSUS I - = DNA binding site mutated, prevents binding, allows transcription DNA binding site Next: 3 important terms of DNA control. • Constitutive activity • Trans acting • Cis acting Β-galactosidase No Lactose 1 I+P+O+Z+Y+ 2 I-P+O+Z+Y+ 3 I+P-O+Z+Y+ 4 I+P+OcZ+Y+ 5 I+P+O+Z-Y-/ I-P+O+Z+Y+ 6 I+P+OcZ+Y-/ I+P+O+ZY+ 7 I+P+OcZ-Y+/ I-P+O+Z+Y- 8 ISP+O+Z+Y-/ I-P+O+Z-Y+ 9 I+P-OcZ+Y+/ I+P+O+Z-Y- - Lactose + Permease No lactose - Lactose + Negative inducible operon Question: what happens when both lactose and glucose are present together in the same cell? To answer this question you first need to understand this reaction ATP Adenylyl cyclase Cyclic AMP + catabolite activator protein (helix-turn-helix) Glucose may inhibit this enzyme • cAMP and glucose levels are inversely proportional. The promoter needs to be in an ideal conformation for RNA polymerase. How CAP and cAMP affects the promoter END of PART I Trypthophan Operon Trypthophan Operon • 5 genes involved in the synthesis of the amino acid trypthophan Negative repressible operon 2 shapes to the mRNA attenuation Eukaryotic Gene Regulation Do eukaryotes show coordinate gene regulation? Yes, the same response element may be found in related genes. Changes in Chromatin structure and eukaryotic gene regulation • Chromatin Structure – DNase hypersensitivity ( • Histone acetylation • DNA methylation