D-Alanine: membrane acceptor ligase from Lactobacillus casei.

The enzymatic incorporation of D-alanine into membranes of Lacfobacillus casei (ATCC 7469) is described. The activity is dependent on ATP, supernatant fraction and membrane fragments; it is enhanced by the addition of Mg2+. The incorporation is insensitive to ribonuclease. The product of this reaction is characterized as a hydroxylamine (1 M, pH 7.0, 37”) labile ester that is not extracted into lipophilic solvents. The lability of the incorporated D-alanine in hydroxylamine is similar to that observed for the D-alanine ester residues in the glycerol teichoic acid from this organism. The chromatographic properties of the product are not consistent with alanyl phosphatidylglycerol. The K, for D-alanine and D-ol-NH2-n-butyric acid is 18 pM and 850 pM, respectively. L-[r*C]Alanine is not incorporated. The following analogues are effective inhibitors of D-alanine incorporation: (a) D-(r-NH2-n-butyric acid; (b) DL-alanine hydroxamate; (c) DL-oc-amino-n-butyric acid hydroxamate; (d) DL-alanine amide; and (e) D-serine. Although the specificity profile is similar to that described for the D-alanineactivating enzyme (BADDILEY, J., AND NEUHAUS, F. C., Bio-