15] Choline/ethanolamine kinase from rat liver

Abstract

Choline kinase is the initial enzyme in the CDPcholine pathway for phosphatidylcholine biosynthesis in mammalian cells. It catalyzes the Mg 2+ -ATP-dependent phosphorylation of choline. Choline kinase is rate limiting for phosphatidylcholine formation in several cells and tissues. The enzyme exists as at least two isoelectric forms in all tissues examined; a third form is inducible with polycyclic aromatic hydrocarbons and hepatoxic agents in liver. Both constitutive isoforms in liver have ethanolamine kinase activity, and kinetic and immunological data show that both kinase activities are catalyzed by the same enzymes. These results on the regulation and enzymatic activities of choline kinase indicate that the enzyme plays an important role in the long-term regulation of the CDPcholine pathway and may be involved in the coordinate regulation of phosphatidylethanolamine biosynthesis. To study choline and ethanolamine phosphorylation with a purified kinase preparation and to investigate the nature of the isoforms, a purification protocol for rat liver choline-ethanolamine kinase is developed. With the use of size-exclusion chromatography as the final step, a preparation of very high specific activity can be obtained. To obtain homogeneous, though less active, enzyme, chromatofocusing is used as the final step.