Lac operon

The lac operon consists of three adjacent genes required for the transport and of lactose (milk sugar) in the Escherichia coli (E. coli) and some other bacteria. The term operon is used when genes (in this case lacZYA) are co-transcribed into a single messenger RNA. The lac operon is regulated by several factors, one of which is the availability of lactose as an energy source. Control of the lac genes was the first genetic regulatory mechanism to be elucidated, one reason for this is that it is one of the simplest, at least in outline, consisting of simple negative (lac repressor) and positive (CAP) regulatory elements. The lac operon has been considered the canonical example of prokaryotic gene regulation.

Multimeric nature of repressor and the complex operator

Lac repressor is a tetramer of identical subunits. Each subunit contains a helix-turn-helix (HTH) motif capable of binding to DNA. The operator site where repressor binds is a DNA sequence with inverted repeat symmetry. The two DNA half-sites of the operator together bind to two of the subunits of the tetrameric repressor. Although the other two subunits of repressor are not doing anything in this model, this property was not understood for many years.

~ ~ ~ ~ ~ ~ ~ ~ ~ ~

Eventually it was discovered that two additional (minor) operators are involved in lac regulation. One (O3) lies in the end of the lacI gene and the other (O2) is about 400 bp downstream in the early part of lacZ. These two sites were not found in the early work because they have redundant functions and individual mutations do not affect repression very much. Single mutations to either O2 or O3 have only 2 to 3-fold effects. However, their importance is demonstrated by the fact that a double mutant defective in both O2 and O3 is dramatically de-repressed (by about 70-fold).

~ ~ ~ ~ ~ ~ ~ ~ ~ ~

In the current model, repressor is bound simultaneously to both the main operator O1 and to either O2 or O3. The intervening DNA loops out from the complex. The redundant nature of the two minor operators suggests that it is not a specific looped complex that is important. One idea is that the system works through tethering. If bound repressor releases from O1 momentarily, binding to a minor operator keeps it in the vicinity, so that it may rebind quickly. This would increase the affinity of repressor for O1.

~ ~ ~ ~ ~ ~ ~ ~ ~ ~

Mechanism of induction

The repressor is an allosteric protein, i.e. it can assume either one of two slightly different shapes, which are in equilibrium with each other. In one form repressor is capable of binding to the operator DNA, and in the other form it cannot bind to the operator. According to the classical model of induction, binding of the inducer, either allolactose or IPTG, to the repressor affects the distribution of repressor between the two shapes. Thus, repressor with inducer bound is stabilized in the non-DNA-binding conformation. However, this simple model cannot be the whole story, because repressor is bound quite stably to DNA, yet it is released rapidly by addition of inducer. Therefore it seems clear that repressor can also bind inducer while still bound to DNA.

~ ~ ~ ~ ~ ~ ~ ~ ~ ~