Melvin Cohn

Sur la biosynthese de la β-galactosidase (lactase) chez Escherichia coli. La specificite de l'induction

1. 1. The formation of β-galactosidase by E. coli is induced exclusively by substances possessing an intact galactosidic radical. Various inversions or substitutions at one or more positions as well as the suppression of the carbon 6 result in the disappearance of the inductive property. 2. 2. The substances which have the inductive property are not necessarily substrates of the enzyme. Thus certain α-galactosides (melibiose) are inductors yet they are not hydrolysed by β-galactosidase. 3. 3. The inductivity is, in general, independent of the affinity for the affinity for the enzyme. Certain substances (phenyl-β-thiogalactoside) which have a high affinity for β-galactosidase in vitro as in vivo are deprived of the inductive property. 4. 4. The formation of β-galactosidase is inhibited by phenyl-β-D-thiogalactoside, but the inhibition is not competitive, while it should be if it was due to the inhibition of the enzyme. 5. 5. These observations are incompatible with all hypotheses which imply that the induction is connected, either with the activity of the enzyme, or with the formation of a specific complex between the enzyme and the inductor.

Studies on the induced synthesis of β-galactosidase in Escherichia coli: The kinetics and mechanism of sulfur incorporation

Abstract A study of the kinetics of sulfur incorporation into the molecule of β-galactosidase during the induced synthesis of this enzyme in E. coli brings proof that the enzyme-protein is synthesized entirely de novo without any appreciable participation of materials coming from other cellular proteins. Furthermore, there is no measurable renewal of β-galactosidase sulfur in growing cells whether or not the enzyme is being synthesized. The induced synthesis of β-galactosidase appears as a virtually irreversible process. The bulk of the other cellular proteins in E. coli are equally stable and do not undergo any appreciable degradation and resynthesis during growth. The apparent contradiction between these results and the generally accepted concepts regarding the dynamic state of intracellular proteins is discussed.

Purification et propriétés de la β-galactosidase (lactase) d'escherichia coli☆

1. 1. A method for the purification of the adaptive enzyme of E. coli, β-galactosidase (lactase) has been described. 2. 2. Lactose and o-nitrophenyl-β-d-galactopyranoside are quantitatively hydrolysed by this enzyme preparation which, however, is inactive on sugars not possessing the β-galactosidic configuration. 3. 3. The following results (4, 5, and 6) indicate that the hydrolysis of lactose and o-nitrophenyl-β-d-galactopyranoside i is catalyzed by the same enzyme. 4. 4. The speed of thermal inactivation is the same whether one determines the activity with lactose or o-nitrophenyl-β-d-galactopyranoside as substrate. 5. 5. The electrophoretic analysis of this purified preparation reveals the presence of at least two constituents. Each of these electrophoretically distinct constituents is active on lactose and on o-nitrophenyl-β-d-galactopyranoside, and the ratio of these activities is the same for both fractions. 6. 6. A precipitating anti-lactase serum has been obtained by immunization of rabbits with the purified enzyme preparation. A quantitative study of the precipitin reaction indicates that the enzymatic activity, whether it be determined on lactose or o-nitrophenyl-β-d-galactopyranoside, is associated with a single antigenic species. There has also been demonstrated the existence of a protein which possesses no enzymatic activity but which markedly crossreacts with the β-galactosidase. 7. 7. The action of various ions on the enzymatic hydrolysis of lactose has been investigated. The effects of monovalent cations are particularly marked and complex. Depending upon the conditions (the presence of certain other cations), a given ion (sodium) is able to behave as either an inhibitor or an activator. 8. 8. These effects are the result of the competetive relationship between the monovalent cations. In order to account for the above phenomena, a hypothesis is presented, bringing into play only two characteristic constants: the affinity (inverse of the apparent dissociation-constant of ion-enzyme complex) and the “activance” (activity of the enzyme when saturated by the ion). 9. 9. For the hydrolysis of lactose, potassium shows the maximum “activance”, while the sodium is only weakly active. On the other hand, with o-nitrophenol-β-d-galactopyranoside as substrate, the “activance” of sodium is higher than that of the potassium. This remarkable inversion of the effects of ions demonstrates that the activance is linked to the chemical structure of the substrate.

NON-INDUCIBLE MUTANTS OF THE REGULATOR GENE IN THE "LACTOSE" SYSTEM OF ESCHERICHIA COLI.

Two independent mutants of Escherichia coli K12 have been isolated which have lost the capacity to be induced by β -galactosides for the production of the proteins governed by the lactose Operon in E. coli . These mutations are located in the regulator ( i ) gene of the Lac region. In heterogenotes, the mutated i s allele is dominant over the wild i + allele. Lac + revertants are obtained as a result of a secondary mutation either in the i gene ( i − ) or in the operator ( o c ). The i s mutations appear to result in an alteration of the lactose repressor which becomes unable to respond to β -galactosides.

Rôle du lactose et de ses produits métaboliques dans l'induction de l'opéron lactose chez Escherichia coli

Abstract Role of lactose and its metabolic products in the induction of the lactose operon in Escherichia coli It is shown that lactose is not an inducer of the “Lac” operon in Escherichia coli K-12. Induction of this operon in the presence of lactose results from transgalactosidation reactions for which β-galactosidase (EC 3.2.1.23) itself is responsible. Several compounds obtained by the action of galactosidase on a mixture of lactose and various alcohols have been isolated and shown to be inducers of the system.

The relationships in biosynthesis of the β-galactosidase- and Pz-proteins in Escherichia coli☆

E. coli cells possess a protein, Pz, which appears to be synthesized under all conditions of growth. In the presence of a galactoside, the cells also synthesize another protein, Gz, endowed with β-galactosidase properties, and very closely similar to Pz in antigenic structure as well as in solubility properties. The synthesis of Gz interferes very significantly with the net rate of Pz synthesis. Only those species of Enterobacteriaceae which possess the Pz protein are competent to synthesize β-galactosidase.