Regulation of gene expression of Lac Operon in E. coli

• Regulation of gene expression involves two steps; transcription and translation to form a complete protein or enzyme.
• Some enzymes are required under certain conditions but not all times. Synthesis of such enzymes continuously all the time, is wastage of energy.
• In order to minimize this energy loss, some control mechanisms are required to control transcription and translation.
• These mechanisms allow transcription and translation to occur only when the enzymes are required and block the same when the enzymes are not required.
• Such control over transcription and translation as per the requirement of certain protein is called regulation of gene expression.
• One best example of regulation of gene expression is Lac Operon in E. coli.

Lac Operon in E. coli:

• An operon is a functional unit of DNA that contains a cluster of genes which are controlled by a single promoter. Such genes present in the operon are either expressed together for a particular protein or not at all.
• ‘Lac’ in Lac Operon stands for lactose. Lac Operon is an example of control of transcription.
• It controls the transcription of genes that code for the enzymes required for the transport and metabolism of lactose in many enteric bacteria including coli.
• Lac Operon consists of two types of genes; structural genes and regulatory genes.
• Structural genes of Lac Operon are lac Z, Lac Y and Lac A which encode for respective enzymes required for the transport and metabolism of lactose.
  • lac Z: It encodes the enzyme β-galactosidase which hydrolyzes lactose to form glucose and galactose and also converts lactose into its isomeric form, allolactose.
  • lac Y: It encodes the enzyme β-galactoside permease which is a transporter protein and facilitates the entry of lactose into the cell.
  • lac A: It encodes the enzyme β-galactoside transacetylase. This enzyme transfers an acetyl group from acetyl-CoA to toxic β-galactosides, glucosides and lactosides and removes them out of the cell.
• Regulatory genes include a repressor gene (Lac I) along with Promoter (P) and Operator (O) genes.
• Regulatory genes regulate the transcription of structural genes.
  • lac I: It encodes repressor protein that binds to the operator sequence.
  • Promoter gene: It serves as the binding site for RNA polymerase.
  • Operator gene: It serves as the binding site for repressor protein.

Mechanism of regulation of Lac Operon:

• Lac Operon is regulated by the following three mechanisms:
  • Repression of Lac mRNA transcription
  • Induction of Lac mRNA transcription
  • Catabolic repression: (In the presence of glucose and lactose)

Repression of Lac mRNA transcription (In the absence of lactose):

• RNA polymerase must bind to promoter gene in order to transcribe structural genes.
• Repressor gene (lac I) encodes repressor protein that binds to operator gene.
• There are three regions in operator gene. O1, O2 and O3.
• Repressor protein generally binds to region O1 of Operator but sometimes can bind to secondary region O2 which is near lac Z and region O3 which is upstream to promoter.
• When repressor protein binds to operator, RNA polymerase cannot bind to promoter. It is because binding of repressor to operator overlaps promoter and prevents RNA polymerase from binding to promoter.
• No binding of RNA polymerase to promoter, stops the transcription of structural genes. This occurs in the absence of lactose in the cell.
• Some coli even in their repressed state have minimal level of enzymes, β- galactosidase and β-galacotoside permease which are essential for operon regulation in the immediate supply or lactose afterwards.

Induction of Lac mRNA transcription (In the presence of lactose):

• When lactose is present in the cell, it binds to repressor protein causing its conformational change. This change dissociates repressor protein form the operator.
• Lactose appears to serve as an inducer here but actually it is its isomeric form, allolactose, which acts as a real inducer.
• β-galactosidase converts lactose into allolactose that binds to repressor protein and inactivates it.
• Inactivation of repressor protein reduces its affinity towards the operator. This leads to the binding of RNA polymerase to the promoter region.
• Finally, transcription of structural genes occurs and the enzymes β-galactosidase, β-galactoside permease and β-galactoside transacetylase are synthesized responsible for the metabolism of lactose.

Catabolite repression or positive control (In the presence of glucose and lactose):

• When glucose is present in the cell, metabolism of lactose doesn’t occur as glucose suppresses the transcription of lactose metabolizing enzymes.
• It is regulated by catabolite activator protein (CAP) encoded by CAP gene located elsewhere in the chromosome.
• The binding of CAP near promoter upstream to Lac I facilitates the binding of RNA polymerase to promoter more tightly. This tight binding accelerates the rate of transcription by 20 folds.
• CAP initially is inactive which requires cyclic adenosine monophosphate (cAMP) for activation. cAMP, being a secondary messenger, binds to CAP only in the absence of glucose.
• When glucose level increases in the cell, it inhibits the synthesis of cAMP. High glucose level also transports cAMP out of the cell decreasing its concentration in the cell.
• Hence, when glucose level increases, cAMP level decreases and CAP is not activated, finally inhibiting transcription of Lac genes for lactose metabolism.

Feedback regulation of lac operon:

1. Active transport of lactose (inducer) into the cell by β-galactoside permease increases its concentration thus resulting in increased induction of lac operon.
2. Induction of lac operon is enhanced by transgalactosylation of lactose by β-galactosidase producing allolactose.
3. Increased concentration of glucose due to the hydrolysis of lactose and allolactose by β -galactosidase results in catabolite repression of lac operon.

Regulation of gene expression of Lac Operon in E. coli