Operon Concept Lecture PDF

# Operon Concept ## Discovery of the Operon - In 1961, Jacob and Monod studied the metabolism of lactose by *E. coli*. - Three enzyme activities/three genes were induced together by galactosides. - Based on their observation they proposed operon concept. - Nobel prize in the year 1965. ## Introd...

# Operon Concept ## Discovery of the Operon - In 1961, Jacob and Monod studied the metabolism of lactose by *E. coli*. - Three enzyme activities/three genes were induced together by galactosides. - Based on their observation they proposed operon concept. - Nobel prize in the year 1965. ## Introduction - An operon is a collection of prokaryotic genes transcribed together on a single mRNA transcript (polycistronic) to serve a single purpose. - Composed of: - An operator, an "on-off" switch. - A promoter. - Genes for metabolic enzymes. - Can be switched off by a repressor protein. - A corepressor is a small molecule that binds to a repressor to switch an operon off. ## Regulatory proteins control operon transcription: ### Positive Regulation - An activator protein binds to the promoter and activates transcription. ### Negative Regulation - A repressor protein binds to the operator and inhibits transcription. ## Operons are either REPRESSIBLE or INDUCIBLE | Type | Description | |:---|:---| | REPRESSIBLE | Involves binding a co-repressor (metabolite/end-product) to repress/turn OFF operon transcription | | INDUCIBLE | Involves binding an inducer (metabolite/starting substrate) to induce/turn ON operon transcription | ## 3 combinations found in nature: | Type | REPRESSIBLE | INDUCIBLE | |:---|:---|:---| | NEGATIVE | This operon uses a repressor that binds a co-repressor to repress transcription. | This operon uses a repressor that binds an inducer to induce transcription. | | POSITIVE | Not found. | This operon uses an activator that binds an inducer to induce transcription. | ## Lac Operon - The *lac* operon was the first operon discovered. - It contains 3 genes coding for *E. coli* proteins that permit the bacteria to use the sugar lactose. - **LacZ encodes Beta-galactosidase** - Breaks up lactose into glucose and galactose (galactose also converted to glucose for metabolism). - Isomerizes lactose into allolactose inducer (presence of lactose means presence of allolactose). - **Lacy encodes permease** - For lactose transport across the cell membrane. - **LacA encodes transacetylase** - Poorly understood function. ## The Lac operon has 2 control circuits: ### NEGATIVE INDUCIBLE - Uses a repressor that binds an inducer (allolactose) to induce operon transcription. - Requires presence of lactose. ### POSITIVE INDUCIBLE - Uses an activator (CAP) that binds an inducer (cyclic AMP) to induce operon transcription. - Requires absence of (preferred) glucose. ## LAC OPERON NEGATIVE INDUCIBLE CONTROL CIRCUIT: - In the absence of lactose, an active repressor protein binds to the operator and blocks transcription by RNA Polymerase: - A diagram of this is shown in (a) Lactose absent, repressor active, operon off. - When lactose is present in the cell, allolactose, an isomer of lactose, binds to the repressor. - This inactivates the repressor, because it can no longer bind the operator. - Now RNA Polymerase can transcribe the *Lac* operon: - A diagram of this is shown in (b) Lactose present, repressor inactive, operon on. - RNA polymerase have low affinity towards the lac operon. - So even the operon is on, transcription will occur poorly. - There comes the role of positive inducible control ## LAC OPERON POSITIVE INDUCIBLE CONTROL CIRCUIT: - Attachment of RNA polymerase to the promoter requires the presence of a Catabolite gene Activator Protein (CAP) that is bound to the cAMP. - Presence of glucose lowers the cAMP by inactivating adenylyl cyclase responsible for the synthesis of cAMP. - Due to lesser cAMP formation of CAP-cAMP is low. - So RNA polymerase attachment and transcription will be negligible in the presence of glucose. - As CAP-cAMP is essential for transcription of the lac operon it is considered as a positive regulator. - A diagram of this is shown in (a) Lactose present, glucose scarce (cAMP level high): abundant lac mRNA synthesized. ## Structure of IPTG - A diagram of the chemical structure of Isopropyl-ß-D-thiogalactoside (IPTG) is shown. ## Trp Operon - TrpE gene product - TrpD gene product - TrpC gene product - TrpB gene product - TrpA gene product ## The Trp operon has 2 control mechanisms: ### NEGATIVE REPRESSIBLE OPERON - Uses a repressor that binds a co-repressor (end product Trp) to repress operon transcription by 70-fold. - Requires presence of Trp. ### ATTENUATION - Involves premature transcription termination - Requires high Trp levels. ## TRP OPERON NEGATIVE REPRESSIBLE CONTROL CIRCUIT: - By itself, the operon is on. RNA polymerase can bind to the promotor and moves freely through the operator to transcribe the genes: - A diagram of this is shown in (a) Tryptophan absent, repressor inactive, operon on. - When co-repressor (end-product) Trp is present, it binds to the repressor. This activates the repressor, causing it to bind the operator to block Trp operon transcription. - A diagram of this is shown in (b) Tryptophan present, repressor active, operon off.