Prokaryotic Gene Expression (Learning Objectives)

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1 Prokaryotic Gene Expression (Learning Objectives) 1. Learn how bacteria respond to changes of metabolites in their environment: short-term and longer-term. 2. Compare and contrast transcriptional control of bacterial biosynthetic and breakdown pathways and their control mechanisms. 3. Explain the organization of bacterial genomes and the molecular components of their regulation: operon, structural genes, promoter, operator, repressor and corepressor, inducer, activator, and activator binding site. 4. Compare and contrast the molecular mechanism (on/off switches) controlling expression of repressible and inducible operons.

2 Review of Basic concepts! True of False Living cells are efficient systems that do not waste energy or resources Biochemical pathways are either biosynthetic or breakdown pathways Cellular feedback inhibition enables cells/organisms to response to their environment A protein is produced in a cell if its gene is present and expressed (i.e. transcribed and translated) A transcription unit (gene) consist of coding region and a promoter region Conformation of an allosteric protein can be modified and this affects its interaction with other biological molecules

3 Metabolism and Metabolites Metabolism Set of chemical reactions that synthesize or breakdown molecules Metabolites Substances produced or take part in cellular metabolism

4 Examples of Metabolites Tryptophan Amino acid Essential in humans Non-essential in bacteria Produced by a Biosynthetic (anabolic) pathway Lactose Sugar in milk (Disaccharide) Source of carbon and energy Broken down by a catabolic pathway only when glucose is absent. Galactose Glucose

5 Tryptophan: a precursor of imp biological substances Serotonin: a neurotransmitter regulates mood, emotions, aggression, sleep, appetite, anxiety, memory, perceptions, and body temperature. Released during sunny weather, eating chocolate or by Ecstasy. Melatonin regulation of the circadian rhythm- sleep Plants hormones Human Connections Serotonin Melatonin

6 Control mechanisms of biosynthetic In response to Environmental changes Short-term Pathways in Bacteria Adjust the activity of enzymes of existing pathways (feedback inhibition). Longer-term Regulate gene transcription.

7 Control of Tryptophan Biosynthesis in Bacteria Feedback inhibition Turning off specific gene expression (transcription)

8 Transcriptional Control of Bacterial Operons Operons of biosynthetic (anabolic ) pathways Operons of degradation (catabolic ) pathways Normally turned on but can be turned off or shut down Normally turned off but can be turned on Repressible operons (trp operon) Inducible operons (lac operon)

9 Organization of Bacterial Genomes Operons are gene clusters that share the same promoter region DNA Operator Structural Genes Polycistronic mrna TrpE TrpD TrpC TrpB TrpA Separate Proteins

10 Genetic Components of Bacterial Operons and their molecular control - Structural genes transcribed as one long mrna molecule (polycistronic mrna) - Promoter region-rna polymerase binding - Operator region on-off switch between the promotor and the first gene - Regulatory gene coding for a specific allosteric repressor protein

11 On-Off switch of Transcription The operator DNA region- binding site for the repressor protein Operons have different operator sequences Switch is off - repressor can bind to the operator sequence (active repressor) Switch is on - repressor cannot bind to the operator (inactive repressor)

12 Both repressible and inducible operons demonstrate negative control because active repressors can only have negative effects on transcription.

13 Single Switch Repressor-mediated control Repressible Operon trp operon State in absence of tryptophan Promoter RNA Polymerase Operator Repressor Inducible Operon: lac operon State in absence of lactose Promoter RNA Polymerase Operator Repressor

14 Single Switch Activator-mediated control Expression induced by an active activatorexample, Arabinose operon Activator protein Activator-Binding site RNA Polymerase Promoter

15 Molecular Controls of Prokaryotic Gene Expression A. Single Switch 1. Repressor-mediated control 2. Activator-mediated control B. Combination Switch Activator & repressor-mediated control Activator protein Activator-Binding site Activator protein Activator-Binding site Promoter RNA Polymerase Promoter RNA Polymerase Promoter RNA Polymerase Operator Repressor Operator Repressor

16 Tryptophan operon: An example of a repressible operon

17 Role of Tryptophan as a co-repressor Impact of level of tryptophan on - conformation of repressor - state of trp operon expression

18 Human Connections Lactose Lactose intolerance Lactase deficiency Fermentation of bacteria inhabiting intestines

19 lac Operon Inducible operon - presence of lactose Has a double combination switchonly when glucose is absent

20 Dual control Expression induced by an active activator and an inactive repressor Example. lac operon Activator protein Activator-Binding site DNA Polymerase Promoter Operator Combination Switch Activator & repressormediated control Repressor

21 lac Operon: An Inducible Operon Usually on or off? Repressor is in active or inactive conformation? Repressor bound or unbound to lac operator?

22 Role of Allolactose as inducer of lac operon expression or its active transcription Virtual Cell Animation Collections

23 Combination Switches of the lac operon Complex control of lac operon Bacteria prefer to use glucose Other sugars are used only when glucose is absent An additional molecular mechanism is necessary Combination of Switches ons.html#

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