Prokaryo'c Operon Model Ac'vity

Transcription

1 Prokaryo'c Operon Model Ac'vity

2 Differen'al Expression of Genes Prokaryotes and eukaryotes precisely regulate gene expression in response to environmental condi6ons In mul6cellular eukaryotes, gene expression regulates development and is responsible for differences in cell types molecules play many roles in regula6ng gene expression in eukaryotes

3 Prokaryotes coolio! Cyanobacteria release extracellular vesicles containing food parcels and DNA. Extracellular vesicle. what s that all about?

4 Bacteria oaen respond to environmental change by regulacng transcripcon Natural selec6on has favored bacteria that produce only the products needed by that cell Cells can regulate the produc6on of enzymes by feedback inhibi6on or by gene regula6on One mechanism for control of gene expression in bacteria is the operon model

5 Precursor Regulation of a metabolic pathway Feedback inhibition Enzyme 1 Enzyme 2 trpe trpd trpc Regulation of gene expression trpb Enzyme 3 trpa Tryptophan (a) Regulation of enzyme activity (b) Regulation of enzyme production

6 A schemacc representacon of a Generalized Feedback Loop (two types) Nega've Feedback Loop = the effect causes a reversal of the inical condicon. scmulatory- inhibitory response Posi've Feedback Loop = the effect intensifies inical condicon. scmulatory- scmulatory response

7 Operons: The Basic Concept A cluster of func6onally related genes can be coordinately controlled by a single on- off switch The switch is a segment of DNA called an operator usually posi6oned within the promoter An operon is the en6re stretch of DNA that includes the operator, the promoter, and the genes that they control

8 The operon can be switched off by a protein The prevents gene transcrip'on by binding to the operator and blocking polymerase The is the product of a separate regulatory gene

9 The can be in an ac've or inac've form, depending on the presence of other molecules A co- is a molecule that cooperates with a protein to switch an operon off For example, E. coli can synthesize the amino acid tryptophan when it has insufficient tryptophan

10 The trp operon in E. coli: regulated synthesis of repressible enzymes DNA Promoter m 5ʹ trpr Regulatory gene 3ʹ polymerase trp operon Promoter Genes of operon trpe trpd trpc trpb trpa Operator Start codon Stop codon m 5ʹ Protein Inactive (a) Tryptophan absent, inactive, operon on E D C B A Polypeptide subunits that make up enzymes for tryptophan synthesis DNA m 5ʹ trpr 3ʹ trpe No made Protein Tryptophan (co) Active (b) Tryptophan present, active, operon off

11 By default the trp operon is on and the genes for tryptophan synthesis are transcribed When tryptophan is present, it binds to the trp protein, which turns the operon off The is accve only in the presence of its co- tryptophan; thus the trp operon is turned off (repressed) if tryptophan levels are high

12 The trp operon in E. coli: regulated synthesis of repressible enzymes DNA Promoter m 5ʹ trpr Regulatory gene 3ʹ polymerase trp operon Promoter Genes of operon trpe trpd trpc trpb trpa Operator Start codon Stop codon m 5ʹ Protein Inactive (a) Tryptophan absent, inactive, operon on E D C B A Polypeptide subunits that make up enzymes for tryptophan synthesis DNA m 5ʹ trpr 3ʹ trpe No made Protein Tryptophan (co) Active (b) Tryptophan present, active, operon off

13 The trp operon in E. coli: regulated synthesis of repressible enzymes (part 1: tryptophan absent) DNA trp operon Promoter m 5 trpr Regulatory gene 3 polymerase Promoter Operator Start codon m 5 Genes of operon trpe trpd trpc trpb trpa Stop codon Protein Inactive (a) Tryptophan absent, inactive, operon on E D C B A Polypeptide subunits that make up enzymes for tryptophan synthesis

14 The trp operon in E. coli: regulated synthesis of repressible enzymes (part 2: tryptophan present) DNA m 5 Protein trpr 3 Tryptophan (co) trpe Active No made (b) Tryptophan present, active, operon off

15 Repressible and Inducible Operons: Two Types of NegaCve Gene RegulaCon A repressible operon is one that is usually on ; binding of a to the operator shuts off transcripcon The trp operon is a repressible operon it can be shut off DNA trp operon Promoter Regulatory gene Promoter Genes of operon trpr trpe trpd trpc trpb trpa m 5 3 polymerase Operator Start codon m 5 Stop codon Protein Inactive (a) Tryptophan absent, inactive, operon on E D C B A Polypeptide subunits that make up enzymes for tryptophan synthesis

16 An inducible operon is one that is usually off ; a molecule called an inducer inaccvates the and turns on transcripcon The lac operon is an inducible operon and contains genes that code for enzymes used in the hydrolysis and metabolism of lactose By itself, the lac is ac6ve and switches the lac operon off A molecule called an inducer inac6vates the to turn the lac operon on

17 The lac operon in E. coli: regulated synthesis of inducible enzymes DNA m 5 Protein Regulatory gene lac I 3 Promoter polymerase Active Operator (a) Lactose absent, active, operon off IacZ No made DNA lac I lac operon lacz lacy laca m 5 3ʹ 3 polymerase m 5 Start codon Stop codon Protein β-galactosidase Permease Transacetylase Allolactose (inducer) Inactive (b) Lactose present, inactive, operon on

18 The lac operon in E. coli: regulated synthesis of inducible enzymes (part 1: lactose absent) DNA Regulatory gene Promoter Operator lac I IacZ m 5 3 polymerase No made Protein Active (a) Lactose absent, active, operon off

19 The lac operon in E. coli: regulated synthesis of inducible enzymes (part 2: lactose present) DNA lac operon lac I lacz lacy laca m 3 polymerase Start codon Stop codon 5 m 5 Protein β-galactosidase Permease Transacetylase Inactive Allolactose (inducer) (b) Lactose present, inactive, operon on

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21 GROUP CHALLENGE As a team of three, use an operon model as a prop while prac6cing explaining how the model works to other people in the group. You should use the space provided on the back of the ac6vity sheet to take notes on how the genes are regulated by your operon. When you feel that everyone in the group has mastered the process of their operon model, pair up with another group that has the opposite type of operon model. Take turns between the groups explaining how their model regulates gene expression. Be sure to take notes on both models, for example, similari6es and differences between repressibile and inducible operons. Complete the ProkaryoCc Operon Model AcCvity sheet and submit!

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23 Inducible enzymes usually funccon in catabolic pathways; their synthesis is induced by a chemical signal Repressible enzymes usually funccon in anabolic pathways; their synthesis is repressed by high levels of the end product Regula7on of the trp and lac operons involves nega've control of genes because operons are switched off by the ac6ve form of the

24 PosiCve Gene RegulaCon Some operons are also subject to posi6ve control through a s6mulatory protein, such as catabolite ac6vator protein (CAP), an accvator of transcrip6on When glucose (a preferred food source of E. coli) is scarce, CAP is ac6vated by binding with cyclic AMP (camp) Ac6vated CAP ataches to the promoter of the lac operon and increases the affinity of polymerase, thus accelera6ng transcrip6on

25 Positive control of the lac operon by catabolite activator protein (CAP) (part 1: glucose scarce) Promoter Operator DNA lac I lacz CAP-binding site camp Active CAP Inactive CAP Allolactose polymerase binds and transcribes Inactive lac (a) Lactose present, glucose scarce (camp level high): abundant lac m synthesized

26 When glucose levels increase, CAP detaches from the lac operon, and transcrip6on returns to a normal rate volume control CAP helps regulate other operons that encode enzymes used in catabolic pathways

27 Positive control of the lac operon by catabolite activator protein (CAP) (part 2: glucose present) DNA Promoter CAP-binding site Inactive CAP lac I lacz Operator polymerase less likely to bind Inactive lac (b) Lactose present, glucose present (camp level low): little lac m synthesized

28 DNA Promoter Operator Positive control of the lac operon by catabolite activator protein (CAP) lac I CAP-binding site camp Active CAP polymerase binds and transcribes lacz Inactive CAP Allolactose Inactive lac (a) Lactose present, glucose scarce (camp level high): abundant lac m synthesized DNA Promoter lac I lacz CAP-binding site Inactive CAP Operator polymerase less likely to bind Inactive lac (b) Lactose present, glucose present (camp level low): little lac m synthesized