A Mutationally Altered Alkaline Phosphatase from Escherichia coli

An alkaline phosphatase-negative (P-) mutant of Escherichia coli K-12 that produces a protein antigenically related to the wild type enzyme has been found to accumulate a zinc-deficient dimeric form of this metalloenzyme. The mutationally altered alkaline phosphatase could be activated by incubation at pH values between 5 and 6 in the presence of Zn(II) at 37°. The kinetics of the activation process was examined with purified preparations of the apoenzyme. Activation appeared to consist of multiple stages of structural rearrangements induced by zinc binding. The pH sensitivity of the activation of the isolated protein could be correlated with a pH conditionality of the P- phenotype. Thus, the mutant acted as a P- strain when grown at pH 7.4, but was P+ when grown at pH 6.0. Although unable to grow effectively in a medium containing an organic phosphate as the sole phosphate source at pH 7.4, the P- strain survived much better than its P+ parent in a medium totally lacking phosphorus.