Structural Gene For Alkaline Phosphatase Research Articles

Enzyme synthesis following conjugation and recombination in Escherichia coli

The structural gene for alkaline phosphatase, when injected by an Hfr into a constitutive recipient, does not function extensively. After an initial spurt, phosphatase synthesis suffers an eclipse and is not resumed until the P + gene is integrated into the recipient's chromosome. The time-course of genetic recombination was investigated by following the synthesis of alkaline phosphatase in crosses between allelic phosphatase-negative mutants. The appearance of P P + recombinant nuclei in such crosses is paralleled by an increase in the differential rate of enzyme synthesis. Functional recombinants arise 30 to 40 minutes after the entry of the Hfr P gene and multiply exponentially from the time they originate. The kinetics of recombination are unrelated to the generation time of the F −. In a recipient carrying a thermosensitive mutation which blocks DNA replication at 42 °C, recombination occurs in the absence of DNA replication. Interference with DNA replication, in fact, greatly enhances the rate of recombination.

Ultraviolet-sensitive targets in the enzyme-synthesizing apparatus of Escherichia coli.

Inhibition by ultraviolet light of beta-galactosidase and alkaline phosphatase synthesis was investigated in both ultraviolet (UV)-sensitive and UV-resistant (wild-type) Escherichia coli, with the objective of determining the sensitivity of various targets. Kinetics of enzyme formation by unmated bacteria and in mating systems, in which the donor provided the specific genetic material and the recipient the cytoplasm, permit the following conclusions regarding the sensitivity of various targets. Catabolite repression resulting from UV damage causes most of the inhibition of beta-galactosidase formation. When it is largely eliminated by a step-down in nutrition, the principal target in UV-sensitive bacteria appears to be the structural gene (lacZ(+)), but damage to the cytoplasm is also important. Transitory inhibition by inactivation of messenger ribonucleic acid is also observed. In wild-type bacteria, repair reduces the importance of lesions in deoxyribonucleic acid sufficiently that cytoplasmic damage appears to be at least as important. Repair occurs within 10 min, as shown by recovery of enzyme-synthesizing ability. Caffeine and proflavine prevent recovery. Newly mated bacteria respond to irradiation very differently than do unmated bacteria. The beta-galactosidase or alkaline phosphatase structural gene (lacZ(+) or phoP(+)) is much more inhibited after it is transferred than it is in unmated bacteria. This sensitivity seems to depend on a sensitive state of the injected material, rather than on a different physiological condition of the entire zygote. Irradiation of recipient uvr(+) bacteria much more strongly inhibited expression of injected genes than if the F(-) was uvr(s). Studies on mating systems are not very useful for learning about the function of unmated bacteria.