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Publication Abstract from PubMed

Diacylglycerol kinases (DGKs) are multi-domain lipid kinases that phosphorylate diacylglycerol into phosphatidic acid, modulating the levels of these key signaling lipids. Recently, increasing attention has been paid to DGKalpha isozyme as a potential target for cancer immunotherapy. We have previously shown that DGKalpha is positively regulated by Ca(2+) binding to its N-terminal EF-hand domains (DGKalpha-EF). However, little progress has been made for the structural biology of mammalian DGKs and the molecular mechanism underlying the Ca(2+) -triggered activation remains unclear. Here we report the first crystal structure of Ca(2+) -bound DGKalpha-EF and analyze the structural changes upon binding to Ca(2+) . DGKalpha-EF adopts a canonical EF-hand fold, but unexpectedly, has an additional alpha-helix (often called a ligand mimic [LM] helix), which is packed into the hydrophobic core. Biophysical and biochemical analyses reveal that DGKalpha-EF adopts a protease-susceptible "open" conformation without Ca(2+) that tends to form a dimer. Cooperative binding of two Ca(2+) ions dissociates the dimer into a well-folded monomer, which resists to proteolysis. Taken together, our results provide experimental evidence that Ca(2+) binding induces substantial conformational changes in DGKalpha-EF, which likely regulates intra-molecular interactions responsible for the activation of DGKalpha and suggest a possible role of the LM helix for the Ca(2+) -induced conformational changes. SIGNIFICANCE STATEMENT: Diacylglycerol kinases (DGKs), which modulates the levels of two lipid second messengers, diacylglycerol and phosphatidic acid, is still structurally enigmatic enzymes since its first identification in 1959. We here present the first crystal structure of EF-hand domains of diacylglycerol kinase alpha in its Ca(2+) bound form and characterize Ca(2+) -induced conformational changes, which likely regulates intra-molecular interactions. Our study paves the way for future studies to understand the structural basis of DGK isozymes.

Crystal structure and calcium-induced conformational changes of diacylglycerol kinase alpha EF-hand domains.,Takahashi D, Suzuki K, Sakamoto T, Iwamoto T, Murata T, Sakane F Protein Sci. 2019 Jan 17. doi: 10.1002/pro.3572. PMID:30653270^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.