Regulation of DGK? Activity and Substrate Acyl Chain Specificity by Negatively Charged Phospholipids.

ABSTRACT: Diacylglycerol kinase ? (DGK?) is a membrane-bound enzyme that catalyzes the ATP-dependent phosphorylation of diacylglycerol to form phosphatidic acid (PA) in the phosphatidylinositol cycle. DGK? lacks a putative regulatory domain and has recently been reported to be regulated by highly curved membranes. To further study the effect of other membrane properties as a regulatory mechanism of DGK?, our work reports the effect of negatively charged phospholipids on DGK? activity and substrate acyl chain specificity. These studies were conducted using purified DGK? and detergent-free phospholipid aggregates, which present a more suitable model system to access the impact of membrane physical properties on membrane-active enzymes. The structural properties of the different model membranes were studied by means of differential scanning calorimetry and ³¹P-NMR. It is shown that the enzyme is inhibited by a variety of negatively charged phospholipids. However, PA, which is a negatively charged phospholipid and the product of DGK? catalyzed reaction, showed a varied regulatory effect on the enzyme from being an activator to an inhibitor. The type of feedback regulation of DGK? by PA depends on the particular PA molecular species as well as the physical properties of the membrane that the enzyme binds to. In the presence of highly packed PA-rich domains, the enzyme is activated. However, its acyl chain specificity is only observed in liposomes containing 1,2-dioleoyl PA in the presence of Ca²⁺. It is proposed that to endow the enzyme with its substrate acyl chain specificity, a highly dehydrated (hydrophobic) membrane interface is needed. The presence of an overlap of mechanisms to regulate DGK? ensures proper phosphatidylinositol cycle function regardless of the trigged stimulus and represents a sophisticated and specialized manner of membrane-enzyme regulation.

SUBMITTER: Bozelli JC 

PROVIDER: S-EPMC7036746 | biostudies-literature | 2020 Feb

REPOSITORIES: biostudies-literature