Gene regulation in prokaryotes is one of the method of conservation of cell resources by turning OFF and ON of genes transcribing. Controlling of gene expression leads to regulation of metabolism of cell. Regulations of prokaryotic genes are done in units called as Operons. Operons are genes grouped together translated into single mRNA molecule containing coding sequences of more than one gene. Operons can be inducible or repressible and in both ways, it allows the coordinated control of genes required for metabolism.
Two scientists Jacob and Monod proposed the operon model in 1961 for the co-ordinate regulation of transcription of genes involved in specific metabolic pathways. The operon is a unit of gene expression and regulation which typically includes structural genes for enzymes involved in a specific biosynthetic pathway whose expression is coordinately controlled. Control elements such as an operator sequence, which is a DNA sequence that regulates transcription of the structural genes and regulatory gene whose products recognize the control elements, for example a repressor which binds to and regulates an operator sequence.
Lac Operon (Inducible Operon)
In this type of regulation, a very small molecule such as lactose triggers the production of specific protein and this process is called induction and the molecule that triggers the process is called as inducer. Control of lactose metabolism involves three types of structural genes that code for the structure of enzymes involved in lactose uptake and metabolism. Lac operon includes the genes for β galactosidase (Z), galactoside permease (Y) and thiogalactoside transacetylase (A). The three structural genes are encoded in a single transcription unit lacZYA, which has a single promoter. There is one repressor gene also which plays role in regulation-lacI gene. Lac repressor is a protein that is a tetramer of identical monomers. Lac operon has two secondary binding sites for the lac repressor, which are referred to as pseudooperators, one is O2 other is O3 along with the main operator site-O1 to which it binds tightly. In the presence of lactose the lac operon is induced.
Lac Repressor It is encoded by lacI gene, which is active as a tetramer of identical subunits. It has very strong affinity for both lac operator-binding site and DNA. The lac operator site consists of 28 bp, which is palindromic and has symmetry with that of lac repressor. In the absence of lactose, the repressor occupies the operator-binding site. Both the lac repressor and the RNA polymerase can bind simultaneously to the lac promoter and operator sites.
Properties of Repressor In lac-operon, Lac repressor inhibits the expression of genes whereas repression of purines biosynthesis is done via Pur repressor. These two proteins have 31% identical sequences and have similar three-dimensional structures. The gene regulatory properties of these proteins differ in relation to the binding of small molecules to the repressor and presence of recognition sites on the genome.
Induction In the presence of lactose an inducer molecule like Allolactose, which is an isomer of lactose, binds to a specific site on the lac repressor causing a conformational change which leads to dissociation of repressor from the operator. β galactosidase molecule in E. coli converts lactose into allolactose. Another inducer of lac operon that is often used in experiments is IPTG which is non-metabolizable molecule. The addition of inducer such as IPTG very rapidly stimulates transcription of the lactose operon structural genes and the removal of the inducer leads to immediate inhibition of this induced transcription.
Positive Regulation: Catabolite Repression In this type of regulation, expression of genes for catabolism of lactose, arabinose and other sugars in presence of glucose is prevented. As lac promoters are not very strong so they require a specific activator protein like cAMP receptor protein (CRP) for high level of transcription. Effect of glucose is regulated by cAMP and a protein called CRP (cAMP receptor protein). Since glucose is catalyzed preferentially to lactose, both glucose and lactose act in concert on lac operon. CRP has a binding site for cAMP and it binds to DNA when cAMP levels are high. In the presence of glucose E. coli do not require any alternate carbon source and to conserve energy, bacteria shut down the genes that control lactose consumption when glucose is around. This is achieved with the help of a ―repressor protein‖ that sits on lactose genes. When glucose concentration is low the synthesis of cAMP is increased and so it levels increases. This leads to binding of cAMP to CRP/CAP. CAP-cAMP complex binds to specific site near lac promoter and stimulates transcription. CRP binding induces a bend in DNA and this is believed to enhance RNA polymerase binding to the promoter enhancing transcription by 50-fold. Binding of CAP to RNA polymerase by its λ subunit is positive regulation.