Question

Catabolite Repression \(E\). coli cells are growing in a medium that contains lactose but no glucose. Indicate whether each of the following changes or conditions would increase, decrease, or not change the expression of the lac operon. It may be helpful to draw a model depicting what is happening in each situation. a. Addition of a high concentration of glucose b. A mutation that prevents dissociation of the Lac repressor from the operator c. A mutation that completely inactivates \(\beta\) galactosidase d. A mutation that completely inactivates galactoside permease e. A mutation that prevents binding of CRP to its binding site near the lac promoter

Short answer

a: Decrease, b: Decrease, c: Decrease, d: Decrease, e: Decrease.

Step-by-step solution

Analyze the addition of glucose

When glucose is added to the medium, it triggers catabolite repression, where cAMP levels decrease. Low cAMP affects the CRP-cAMP complex formation, reducing its binding to the promoter, resulting in decreased transcription of the lac operon. Therefore, adding glucose decreases lac operon expression.

Examine the mutation preventing Lac repressor dissociation

If there's a mutation where the Lac repressor cannot dissociate from the operator, it will continuously block RNA polymerase from transcribing the lac operon. Hence, this mutation leads to a decrease in lac operon expression.

Consider the inactivation of β-galactosidase

The inactivation of β-galactosidase means lactose cannot be broken down into allolactose, the inducer that detaches the Lac repressor from the operator. Since the repressor remains bound, transcription does not occur, leading to decreased expression of the lac operon.

Evaluate the impact of inactivating galactoside permease

Galactoside permease is responsible for transporting lactose into the cell. Its inactivation reduces lactose entry, which in turn reduces allolactose concentration, maintaining the repressor bound to the operator and thus decreasing expression of the lac operon.

Assess the effect of disabling CRP binding

CRP must bind to its site near the lac promoter to assist RNA polymerase binding and initiate transcription effectively. A mutation preventing CRP binding impairs this process, leading to decreased expression of the lac operon.

Key concepts

Catabolite Repression

Catabolite repression is a priority mechanism that helps bacteria such as \( E. coli \) efficiently manage energy resources. When \( E. coli \) encounters a situation where both glucose and lactose are available, it will prefer to consume glucose due to its simpler and more immediate energy release capacity. In this scenario, the presence of glucose decreases cAMP levels, inhibiting the formation of the CRP-cAMP complex. As a result, the lac operon, crucial for lactose metabolism, is suppressed. The cells thus demonstrate a classic example of catabolite repression, focusing resources on metabolizing the "easier" sugar first.

CRP-cAMP Complex

The CRP-cAMP complex plays a crucial role in the regulation of the lac operon. CRP stands for cAMP Receptor Protein, and this complex is formed when cAMP, a secondary messenger, binds to CRP. In the absence of glucose, cAMP levels are high, facilitating the formation of the CRP-cAMP complex, which then binds to a specific site near the lac promoter.

This binding enhances the association of RNA polymerase with the promoter, stimulating transcription of the lac operon. In situations where glucose is present, decreased cAMP levels lead to fewer CRP-cAMP complexes, hence less activation of the lac operon.

Lac Repressor

The Lac repressor is a protein that regulates the expression of the lac operon by binding to the operator, a specific DNA sequence. This binding prevents RNA polymerase from initiating transcription. In the absence of lactose, the repressor remains bound to the operator, keeping the operon

"off." For transcription to occur, the Lac repressor must dissociate from the operator. This release happens when lactose is present, as it gets converted to allolactose, which then binds to the repressor, altering its shape and reducing its affinity for the operator, thereby allowing transcription initiation.

β-galactosidase Inactivation

β-galactosidase is an essential enzyme encoded by the lac operon, responsible for catalyzing the conversion of lactose into glucose and galactose. It also assists in forming allolactose, which is significant in the regulatory feedback loop of the lac operon. If β-galactosidase is inactivated by mutation or other factors, lactose cannot be adequately processed, and allolactose levels diminish.

Without allolactose, the Lac repressor remains bound to the operator, preventing RNA polymerase from conducting transcription. This inactivation of β-galactosidase results in decreased expression of the lac operon.

Galactoside Permease Inactivation

Galactoside permease is responsible for transporting lactose into the \( E. coli \) cell. This enzyme is another product of the lac operon and is vital for lactose uptake. Inactivating galactoside permease limits the cellular entry of lactose, which subsequently leads to a lower internal concentration of allolactose.

Allolactose serves as the inducer that detaches the Lac repressor from the operator. A lack in allolactose keeps the repressor bound, blocking RNA polymerase, and decreasing the lac operon expression. Thus, the inactivation of galactoside permease severely affects the cell's ability to utilize lactose as an energy source.