The Role of Metal Ions in the Peptidase Activity of Escherichia coli K-12

To obtain information concerning the factors that control the ability of Escherichia coli strain K-12 to hydrolyze glycyl- l -leucine, glycyl- l -phenylalanine, l -leucylglycine, and l -phenylalanylglycine, a study was made of the intracellular localization of the specific dipeptidases and of the effect of changes in cultural conditions on the dipeptidase content of the bacteria. Upon fractionation of cell extracts, significant activity only toward Leu-Gly is found in the ribosomal portion, while at least three peptidases can be shown to be present in the soluble portion: one, which is relatively insensitive to ethylenediaminetetraacetate, is activated by Co 2+ and catalyzes the hydrolysis of Gly-Leu and Gly-Phe; the other two, both of which are EDTA-sensitive, differ in their response to Co 2+ and to Mn 2+ , and in their action on Leu-Gly and Phe-Gly. All the dipeptidases are present in cells grown in peptone-containing media, and the specific activity values of each subcellular fraction, as well as the distribution of peptidase activity between the two fractions, are essentially constant whether the cells are harvested from exponential phase or stationary phase cultures. The dipeptidases are also found in extracts of cells grown in synthetic media devoid of peptone, and the fact that the specific enzymic activity toward each dipeptide is the same regardless of the composition of the growth medium and of the age of the culture indicates that the dipeptidases may be classified as constitutive enzymes. In intact bacteria, the dipeptidases exist in a partially latent condition, and the demonstrable activity of whole cells toward the dipeptides is subject to control by environmental factors, among which are the metal ion content and pH of the culture medium. Evidence is presented in support of the hypothesis that exposure of bacteria to a mixture of metallic salts (either during culture growth or after the attainment of the stationary phase) leads to the replacement of the metal component of active peptidases by a metal that is inhibitory to enzyme action in vivo ; upon disruption of the bacteria, the inactive metal-peptidase complexes dissociate and the peptidases can regain activity. In cells that can form such metal-enzyme complexes, the peptidases are protected from inactivation by exogenous H + , while acid stability is lacking in cells grown in a metal-deficient medium. Among the metals usually present in culture media, Zn 2+ appears to be the most impo