lac-Operon

The lactose operon, short lac operon is an operon that is both during transport and in the degradation of lactose in bacteria, for example Escherichia coli, plays an important role. The lac operon is one of the classic paradigmatic model system for gene regulation; an extracellular signal ( the availability of certain sugar) is translated by the switching on or off of the structural genes of the operon in an energetically favorable adjustment of the metabolism of the cell.

Structure of the operon

, The lac operon is composed of a promoter (P ), three operators ( O1, O2, O3) and three structural genes:

• The lacZ gene encodes the enzyme β -galactosidase ( LacZ, EC 3.2.1.23 ). They hydrolyzed, i.e. splits lactose into glucose and galactose, but can also include lactose into allolactose, a isomer of lactose converted.
• The lacY gene encoding a transport protein known as β - galactoside permease ( LacY ), which allows the inclusion of lactose into the cell.
• The lacA gene coding for the enzyme β - galactoside transacetylase ( EC 2.3.1.18 ). It is not necessary for the lactose decomposition, and its function is not finally resolved. However, there are indications that the acetylation nichtabbaubarer β - galactosides by the enzyme has a detoxification function for the cell.

Regulation of the operon

These three proteins of the lac operon are only expressed ( produced ) when lactose is present in the ambient medium and there is no more favorable for the cell energy source. Such a preferable energy source is, for example, glucose. A system of negative and positive regulation controls the degradation of the most efficient energy source.

The lac operon is regulated by both negative by a repressor, as well as positive by an activator. In addition, the mechanism of inducer exclusion ( inducer exclusion) controls the activity of the Lac permease. The effect of these three regulations that lactose can be only when there are no more efficient alternative metabolized.

Negative regulation of

The negative regulation of the lac operon is at the transcriptional level by a so-called repressor, the lacI protein. This is a homotetrameric protein, which in each case two of the operators at the same time can bind O1 and O2 or O1 and O3 of the advantage of the negative regulation is that, as long as no lactose must be metabolized, no enzymes for their degradation must be provided. The actual repression occurs via the binding of the repressor to O1. The attachment to the same two operators with the result that the DNA located between the two operators forms a loop shape. This has an effect on the strength of repression: If the repressor can only bind to O1 ( in mutants lacking O2 and O3 are missing), the repression of drops about 1000 -fold to about 20-fold. If he does either of O1 and O2 ( in mutants lacking O3 ) and to O1 and O3 ( in mutants lacking O2) binds the same time, the genes of the operon by factors of 700 and 440 are repressed.

The reason for the decrease in the rate of expression is that the necessary RNA polymerase can not efficiently bind to the DNA. Thus, the reading process of the gene, transcription prevented.

The repressor in turn is controlled by a regulatory gene, the lacI gene encoding. This gene is located separately in front of the lac operon (see picture above ) and is expressed by its own constitutive promoter. Since the operator O3 overlaps with the 5 'end of the lacI gene, it may, if this operator is bound to a DNA loop with a high affinity O1 of Lac repressor, lead to a hindrance of the RNA polymerase. It can not reach to the end of the gene and is incident on the DNA. Thus, the mRNA of the lacI gene is incomplete; lacking, for example, the stop codon. Incomplete RNA is translated resulting in the formation of the stopped ribosome ( stop codon is missing). In the bacterial cell they are freed by tmRNA (trans- translation) and it marks the incomplete LacI protein for degradation. This process of trans - translation is an important process in the regulation of the lac operon; without tmRNA incur incomplete lacI repressors, which would still be partially active.

Positive regulation

Is responsible for the positive regulation of the lac operon an activator protein CAP ( catabolite activator protein ), the best studied effector of catabolite repression. The CAP activity is directly dependent on the concentration of cAMP. The addition of cAMP to CAP causes a conformational change in the regulatory protein, which greatly increases its specific DNA binding. The CAP- cAMP complex binds to a binding site of the DNA and interacts directly from there with the RNA polymerase. Wherein the affinity of the RNA polymerase to the promoter is significantly increased. Thus, there are three elements for the positive regulation of the lac promoter necessary: the CAP protein, cAMP and a CAP- binding site in the lac - promoter. Mutations that lead to either the absence of the CAP protein or to the absence of cAMP or to the inactivation of cap- binding site, have the same effect on the lac operon. Decreases its expression about 50 -fold.

Influence of lactose

Is lactose as an energy source is the most efficient substrate in the cell's environment, it is spent by the β - Galactosidpermease into the cell. There it is partially converted by β - galactosidase in allolactose. This means that the Gal - β -1 ,4- Glc- bond is converted to a Gal - β Glc -1 ,6- bond. In this form, an addition to the repressor LacI is now possible.

With this addition, the conformation of the repressor changed and it releases from the operator. Now, the RNA polymerase to begin transcription. Through the subsequent translation further molecules permease and β -galactosidase are provided. So lactose can be permanently used as a substrate until it's depleted or a better energy source is available.

Effect of glucose

For the cell it is advantageous, preferred glucose instead of lactose as a substrate. Consequently, glucose must be removed before lactose. For this to happen, glucose needs to slow down the breakdown of lactose. However, this does not happen directly by the glucose itself during transport of glucose into the cell, the phosphotransferase system is utilized. Herein, the phosphate residue of the PEP on the transport protein IIA E is transmitted to the glucose. The unphosphorylated IIA now E inhibits the lactose permease, whereby no lactose is transported into the cell and the lac operon is inactivated. This process is called inducer. So it happens even in the presence of lactose hardly for gene expression, and the glucose is preferentially degraded.

Many textbooks cause a decrease in cAMP concentration as the reason for the effect of glucose on the expression of the lac operon. This assumption, however, is now obsolete. It was found that the cAMP concentration in the presence of lactose is approximately the same as in the presence of glucose sugar or both at the same time.

History

1961 was developed on the basis of the lac operon of Escherichia coli by the French scientists François Jacob and Jacques Monod operon model of gene regulation. For this work she received the 1965 Nobel Prize in Physiology or Medicine.

Special features of the lac operon

Lactose generally absorbed in the small intestine of humans, but E. coli can be found in particular in the colon. Consequently, lactose is available as a source of nutrition typically not, or only in small amounts for E. coli are available in the large intestine. In fact, the lactose operon is likely to degrade glyceryl- galactosides. These are generally produced by the degradation of fats of animal cells and in this case, if the cells are rejected lining the large intestine and disintegrate. Glyceryl - galactosides are here both as an inducer of the Lac I protein ( repressor ) and as a substrate for β -galactosidase, which cleaves glyceryl galactosides in glycerol and galactose.

Furthermore, the Lac operon can not ( enterobacteria ) which are used are relatively close to E. coli, Salmonella and detected in a variety of other enteric bacteria. It is likely that this segment is seen evolutionarily relatively new in the E. coli genome and it was originally outside the enterobacteria.