Induction of gene expression

The synthesis of many enzymes required for the catabolism (breakdown) of a substrate is regulated by enzyme induction. If the substrate molecule is not present in the environment, there is little point in synthesising the enzyme needed to break it down. In terms of conservation of cellular energy, it makes more sense for such enzymes to be produced only when they are needed, that is, when the appropriate substrate molecule is present. The substrate itself therefore acts as the inducer of the enzyme's synthesis. An example of this that has been studied in great depth concerns the enzyme j-galactosidase, used by E. coli to convert the disaccharide lactose into its constituent sugars:

Enzymes whose production can be switched on and off are called inducible enzymes. This distinguishes them from constitutive enzymes, which are always produced regardless of prevailing conditions.

Lactose

ß-galactosidase

Glucose + Galactose j-galactosidase has been intensively studied as part of the lac operon of E. coli. This is made up of three structural genes designated Z, Y and A, which are clustered together and share a common promoter and terminator (Figure 11.14). The genes code for, respectively, j-galactosidase, a permease and a transacetylase. The permease is necessary for the transport of lactose into the cell, while the role of the transacetylase is not entirely clear, although it is essential for the metabolism of lactose. Grouping the three genes together in this way ensures an 'all-or-nothing' expression of the three proteins. Transcription of these structural genes into their respective mRNAs is initiated by the enzyme RNA polymerase binding to the promoter sequence. However, this is only possible in the presence of lactose; in its absence, a repressor protein binds to an operator site, adjacent to the promoter, preventing RNA polymerase binding to the promoter, and therefore preventing mRNA

A group of functionally related genes involved in the regulation of enzyme synthesis and positioned together at the same locus is called an operon. It contains both structural and regulatory genes.

A repressor protein regulates the transcription of a gene by binding to its operator sequence. It is encoded by a regulatory gene.

Figure 11.14 The lac operon comprises three structural genes under the control of a single promoter and operator sequence. The role of the regulatory gene I is described in Figure 11.15

production. Production of the repressor protein is encoded by a regulator gene (I), situated slightly upstream from the operon (Figure 11.15a).

How then, does the presence of lactose overcome this regulatory mechanism? Allo-lactose, an isomer of lactose and an intermediate in its breakdown, attaches to a site on the lac repressor, thereby reducing the latter's affinity for the operator, and neutralising its blocking effect (Figure 11.15b). The structural genes are then transcribed into mRNA, which is subsequently translated into the three proteins described above, and the lactose is broken down. In the absence of lactose, there are only trace amounts of j-galactosidase present in an E. coli cell; this increases some 1000-fold in its presence.

Figure 11.15 The lac operon is inducible. (a) In the absence of the substrate lactose, the lac operon is 'switched off', due to a repressor protein encoded by the regulatory gene i. The repressor binds to the operator site, preventing the binding of RNA polymerase to the promoter and therefore blocking transcription. (b) Allolactose acts as an inducer by binding the repressor protein and preventing it from blocking the promoter site. Transcription of the three structural genes is able to proceed unhindered

Figure 11.15 The lac operon is inducible. (a) In the absence of the substrate lactose, the lac operon is 'switched off', due to a repressor protein encoded by the regulatory gene i. The repressor binds to the operator site, preventing the binding of RNA polymerase to the promoter and therefore blocking transcription. (b) Allolactose acts as an inducer by binding the repressor protein and preventing it from blocking the promoter site. Transcription of the three structural genes is able to proceed unhindered

When all the lactose has been consumed, the repressor protein is free to block the operator gene once more, and the needless synthesis of further j-galactosidase ceases.

The lac operon can also be induced by isopropyl j-thiogalactoside (IPTG); E. coli is not able to break this down, so the genes remain permanently switched on. IPTG is utilised as an inducer in cloning systems involving the expression of the lacZ gene on pUC plasmids (Chapter 12).

The lac operon is subject to control by positive as well as negative regulator proteins. Transcription of the operon only occurs if another regulatory protein called catabolite activator protein (CAP) is bound to the promoter sequence (see Box 11.5). This is dependent on a relatively high concentration of the nucleotide cAMP which only occurs when glucose is scarce. The activation of the lac operon thus occurs only if lactose is present and glucose is (almost) absent.

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