REGULATION OF GENE EXPRESSION. Bacterial Genetics Lac and Trp Operon

Transcription

1 REGULATION OF GENE EXPRESSION Bacterial Genetics Lac and Trp Operon

2 Levels of Metabolic Control The amount of cellular products can be controlled by regulating: Enzyme activity: alters protein function Transcription: affects whether a gene is turned on or off

3 Example: E. coli in human intestine No tryptophan present, needs to synthesize Tryptophan present, does not need to synthesize Feedback inhibition on the production of trp

4 Levels of Metabolic Control Protein function level: adjust activity of enzymes already present Short term, quick onset regulation

5 Levels of Metabolic Control Transcription level: control the expression of genes which affects number of enzymes produced Long term, slow onset regulation

6 Gene Expression Types of expression Constitutive (unregulated) Inducible (regulated) Repressible (regulated) Definition Genes are always ON Genes are only turned ON as needed Genes are only turned OFF as needed Example Housekeeping genes (regulatory genes) Structural genes Structural genes

7 Gene Expression Why would catabolic pathways have genes that are inducible or repressible?

8 Catabolic Pathway: Inducible Catabolism: breakdown of complex molecules into simpler units It would be a waste of energy for bacteria to make enzymes for the breakdown of a substance if that substance does not exist do not need to catabolize molecules that are not present in its cellular environment

9 Gene Expression Why would anabolic pathways have genes that are inducible or repressible?

10 Anabolic Pathway: Repressible Anabolism: synthesis of molecules from simpler subunits If an essential nutrient is not present in the environment, bacteria must be able to synthesize the nutrient on its own However, bacteria do not need to produce the essential nutrient if it is already present in the environment

11 Examples Classify the metabolic pathway: catabolic or anabolic Lac Operon Polypeptides that make up enzymes for lactose hydrolysis and uptake Trp Operon

12 Operon A group of genes that share a single promoter Components: Promoter Operator Structural genes Results in the production of proteins that have related functions.

13 Example: trp operon Components Related function of the trp operon: enzymatic proteins that catalyze reactions in the synthesis of tryptophan trp operon Promoter Structural Genes Final product is tryptophan Operator Fig a

14 trp operon Structural Genes 5 genes making 5 polypeptides that combine to make 3 enzymes Enzymes participate in a sequence of steps to make tryptophan (trp) an amino acid Fig a

15 Operon Components Promoter Region on DNA where RNAP binds to start transcription acts as the on/off switch for genes Operator region on promoter that controls access of RNAP to the genes Can activate or repress transcription Structural genes Genes to be transcribed by RNAP

16 Operon Regulation: trp operon trp operon Promoter Structural Genes Regulatory Gene Operator Final product is tryptophan Fig a

17 Operator Constitutive expression Regulatory protein Allosteric effect

18 Operon Regulation Regulatory gene region on DNA that codes for the production of the regulatory protein upstream of the operon constitutive expression: transcribed continuously Regulatory protein Binds to operator and blocks attachment of RNAP Allosteric: alternates between active and inactive forms

19 trp operon Regulation Regulatory gene: trpr Allosteric regulatory protein Active or inactive? has 2 forms: active or inactive Fig a

20 trp operon Regulation Inducer or Repressor? Active or inactive? *Regulatory protein is repressor itself when its active Which form is the regulatory protein made as, active or inactive? Recall: the active regulatory protein will bind to the operator which blocks RNAP transcription Hint: trp operon produces tryptophan, an amino acid essential to its survival Fig a

21 Regulatory Gene: trpr Codes for an trp repressor protein Constitutively expressed in its inactive form

22 trp operon Regulation Inactive How do you switch from the inactive form of the regulatory protein to the active form? How do you turn on the regulatory protein? Fig a

23 Effector any molecule that can regulate the activity of a protein Corepressor: activates the repressor causing regulatory protein to bind to the operator Turn off gene expression Inducer: inactivates the repressor prevents regulatory protein from binding to the operator Turn on gene expression

24 trp operon Regulation Inactive Corepressor or inducer? What effect will the effector have on the regulatory protein? What effect will the effector have on gene expression of the trp operon? Is the effector a corepressor or inducer? Fig a

25 Effector: Corepressor When would you want the trp operon to be turned OFF? What molecule do you think is a logical choice to act as a corepressor in the trp operon? Hint: negative feedback Tryptophan (Corepressor) Fig b

26 trp operon Regulation Effector molecule: tryptophan (trp) Corepressor Regulatory protein: trp repressor Made inactive, RNAP can transcribe, trp made Activated by trp (corepressor) Active trp repressor binds to operator Prevents RNAP from transcribing genes Stops trp production

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28 Repressible operon Gene is normally on Transcription is inhibited when a molecule binds allosterically to a regulatory protein Negative feedback / feedback inhibition Anabolic pathways Example: Trp operon is turned off when tryptophan (corepressor) binds to repressor

29 trp operon animation Tutorial chp13/ html Animation hill.com/olc/dl/120080/ bio26.swf TrpAttenuation.mov (ignore information on attenuation)

30 Models of Negative Regulation Repressible Operon (e.g. trp) Repressor made in inactive form à activated by corepressor n Turns gene off Inducible Operon (e.g. lac) Repressor made in active form à inactivated by inducer n Turns gene on

31 lac operon Recall: Lactose = glucose + galactose

32 lac operon Components lac operon Promoter Structural Genes Regulatory Gene Operator Please Offer Zach Your Apple Hydrolyze lactose into glucose and galactose Lactose transporter (into cell) Adds acetyl group to galactose Fig

33 lac operon Structural Genes codes for 3 enzymes β- galactosidase (lacz gene): lactose hydrolysis into glucose and galactose permease (lacy gene): membrane protein that transports lactose into the cell transacetylase (laca gene): adds acetyl group to galactose (significance in lactose metabolism unclear)

34 lac operon Regulation lac operon Promoter Structural Genes Regulatory Gene Operator Please Offer Zach Your Apple Fig

35 lac operon Regulation Transcription Translation Repressor Active or inactive? Regulatory gene: LacI Allosteric regulatory protein has 2 forms: active or inactive Fig a

36 lac operon Regulation Transcription Translation Repressor Active or inactive? Which form is the regulatory protein made as, active or inactive? Recall: the active regulatory protein will bind to the operator which blocks RNAP transcription Hint: lac operon produces enzymes that breakdown lactose Fig a

37 Regulatory Gene: LacI Codes for an active lac repressor protein which binds to the operator Constitutive expression of the active form

38 lac operon Regulation Transcription RNAP Translation Repressor Active What effect will the effector have on the on gene expression of the lac operon? What type of effector do you need to turn off the regulatory protein: corepressor or inducer? Fig a

39 lac operon Regulation Inducer Inactivated repressor by inducer Active transcription Transcription RNAP Repressor Inducer Translation Inducer Molecule A molecule that can bind to the repressor to prevent the repressor from binding to the operator. Fig b

40 lac operon Regulation Transcription RNAP Repressor Translation What molecule would be an appropriate inducer for the lac operon? Fig a

41 lac operon Regulation Effector molecule: allolactose An isomer of lactose An inducer allolactose binds to regulatory protein (lac repressor) Inactivates lac repressor preventing it from binding to the operator Allows RNA polymerase to transcribe genes Produces enzyme β- galactosidase which hydrolyzes lactose

42 lac operon Regulation E. Coli cells mainly use glucose as a source of energy When living in a system with lactose, bacteria can hydrolyze lactose to produce glucose The lac operon is only turned on when lactose is present

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44 Inducible operon Gene normally off Transcription is stimulated when a molecule binds allosterically to a regulatory protein Proteins made on demand Catabolic pathway (breakdown substances) Example: Lac operon turned on when allolactose (inducer) binds to repressor

45 lac operon animation Animation movie- flash.htm LacOperon.html (a little low tech) Tutorial chp13/ html animations/content/lacoperon.html (looks the same as above)