Diglyceride Kinase from Escherichia coli

Abstract

The diglyceride kinase activity of membranes from Escherichia coli was extracted into acidic butan‐1‐ol. The enzyme was purified in organic solvent by precipitation at − 20°C, chromatography on DEAE‐cellulose and repeated chromatography on Sephadex LH‐60. The final 1460‐fold purified enzyme preparation gave a single protein band upon isoelectric focusing in the presence of Triton X‐100 (p I , 4.0) and upon polyacrylamide‐gel electrophoresis in the presence of sodium dodecylsulphate. The latter method as well as gel chromatography on Sephadex LH‐60 indicated a molecular weight of about 15400. The purified enzyme was devoid of lipid, and it required re‐addition of lipid for activity. sn ‐1,2‐Dipalmitate and ceramide were phosphorylated, whereas the C 55 ‐isoprenoid alcohol, ficaprenol, did not serve as a substrate under the same conditions. Conversely, the butanol‐soluble C 55 ‐isoprenoid‐alcohol kinase from Staphylococcus aureus did not phosphorylate sn ‐1,2‐dipalmitate.