1、Copyright 2005 Pearson Education,Inc.publishing as Benjamin CummingsAnticipatory Questions 1.What might happen if an organism had its cells expressing all genes within the genome all the time?2.At what levels can control of cellular activities/pathways be controlled?3.Based on our discussions up to
2、this point,what do you think the term“negative feedback”means?4.What steps are involved in the initiation of prokaryotic transcription?Copyright 2005 Pearson Education,Inc.publishing as Benjamin CummingsLearning Objectivesunderstand that regulation of gene expression is a means by which to control t
3、iming and rate of generation regarding functional gene product(either RNA or polypeptide/protein).explain the concept of an operon in terms of components functions(promoter,operator,repressor,co-repressor,inducer,gene cluster,polycistronic transcript).compare and contrast repressible and inducible o
4、peron systems/pare and contrast negative versus positive regulation of operons apply the operon concept to gene expression as it relates to genetic engineering(specifically,our cloning and expression of the“tomato”gene).Copyright 2005 Pearson Education,Inc.publishing as Benjamin CummingsIndividual b
5、acteria respond to environmental change by regulating their gene expression A bacterium can tune its metabolism to the changing environment and food sources This metabolic control occurs on two levels:Adjusting activity of metabolic enzymes Regulating genes that encode metabolic enzymesLE 18-20Regul
6、ation of enzymeactivityRegulation of enzymeproductionEnzyme 1Regulation of gene expressionEnzyme 2Enzyme 3Enzyme 4Enzyme 5Gene 2Gene 1Gene 3Gene 4Gene 5TryptophanPrecursorFeedbackinhibitionCopyright 2005 Pearson Education,Inc.publishing as Benjamin CummingsOperons:The Basic Concept In bacteria,genes
7、 are often clustered into operons,composed of An operator,an“on-off”switch A promoter Genes for metabolic enzymes An operon can be switched off by a protein called a repressor A corepressor is a small molecule that cooperates with a repressor to switch an operon offLE 18-21aPromoterPromoterDNAtrpRRe
8、gulatorygeneRNApolymerasemRNA3 5 ProteinInactiverepressorTryptophan absent,repressor inactive,operon onPolycistronic*mRNAtrpEtrpDtrpCtrpBtrpAOperatorStart codonStop codontrp operonGenes of operonEPolypeptides that make upenzymes for tryptophan synthesisDCBA5*=mRNA carries the information of several
9、genes,which are translated into several proteinsLE 18-21b_1DNAProteinTryptophan(corepressor)Tryptophan present,repressor active,operon offmRNAActiverepressorLE 18-21b_2DNAProteinTryptophan(corepressor)Tryptophan present,repressor active,operon offmRNAActiverepressorNo RNA madeCopyright 2005 Pearson
10、Education,Inc.publishing as Benjamin CummingsTrp Operon Animation http:/ 2005 Pearson Education,Inc.publishing as Benjamin CummingsRepressible and Inducible Operons:Two Types of Negative Gene Regulation A repressible operon is one that is usually on;binding of a repressor to the operator shuts off t
11、ranscription The trp operon is a repressible operon An inducible operon is one that is usually off;a molecule called an inducer inactivates the repressor and turns on transcription The classic example of an inducible operon is the lac operon,which contains genes coding for enzymes in hydrolysis and
12、metabolism of lactoseLE 18-22aDNAlaclRegulatorygenemRNA5 3 RNApolymeraseProteinActiverepressorNoRNAmadelacZPromoterOperatorLactose absent,repressor active,operon offLE 18-22bDNAlaclmRNA5 3 lac operonLactose present,repressor inactive,operon onlacZlacYlacARNApolymerasePolycistronic mRNAProteinAllolac
13、tose(inducer)Inactiverepressor-GalactosidasePermeaseTransacetylase5 Copyright 2005 Pearson Education,Inc.publishing as Benjamin CummingsLac Operon Animationhttp:/ 2005 Pearson Education,Inc.publishing as Benjamin Cummings Inducible enzymes usually function in catabolic pathways Repressible enzymes u
14、sually function in anabolic pathways Regulation of the trp and lac operons involves negative control of genes because operons are switched off by the active form of the repressorCopyright 2005 Pearson Education,Inc.publishing as Benjamin CummingsPositive Gene Regulation Some operons are also subject
15、 to positive control through a stimulatory activator protein,such as catabolite activator protein(CAP)When glucose(a preferred food source of E.coli)is scarce,the lac operon is activated by the binding of CAP When glucose levels increase,CAP detaches from the lac operon,turning it offLE 18-23aDNAcAM
16、PlaclCAP-binding sitePromoterActiveCAPInactiveCAPRNApolymerasecan bindand transcribeOperatorlacZInactive lacrepressorLactose present,glucose scarce(cAMP level high):abundant lacmRNA synthesizedLE 18-23bDNAlaclCAP-binding sitePromoterRNApolymerase cant bind efficientlyOperatorlacZInactive lacrepresso
17、rInactiveCAPLactose present,glucose present(cAMP level low):little lacmRNA synthesizedCopyright 2005 Pearson Education,Inc.publishing as Benjamin CummingsCatabolite Activator Protein Mechanism http:/highered.mcgraw- on“combination of switches-the lac operon”Copyright 2005 Pearson Education,Inc.publi
18、shing as Benjamin CummingsThe Arabinose Operon-A Composite of Negative&Positive Regulationa)In the presence of arabinose:CAP-cAMP complex and araC-arabinose complex bind to initiator region this allows RNA polymerase to bind to the promoter transcription beginsb)In the absence of arabinose:araC prot
19、ein assumes a different conformationacts as a repressor binds to araI and a second operator region araO forms a loop this loop prevents transcriptionCopyright 2005 Pearson Education,Inc.publishing as Benjamin CummingsRegulatory geneOperator (part of the promoter)Promoter for the cluster of genesB,A,
20、and DApplication of Operons:Copyright 2005 Pearson Education,Inc.publishing as Benjamin CummingsaraC regulatory generepressorInducer(arabinose)Gene DArabinose operon with in-frame foreign DNA inserted:Tomato genestartstartstartstartstopstopstopstopProtein BProtein AProtein DRed Fluorescent Protein (RFP)Gene BGene AGene DTomato gene53Polycistronic mRNAtranscriptiontranslationtranslationtranslationtranslation
