Publications

• Characterization of the yeast DGK1-encoded CTP-dependent diacylglycerol kinase.

  Han G.-S., O'Hara L., Siniossoglou S., Carman G.M.

  The Saccharomyces cerevisiae DGK1 gene encodes a diacylglycerol kinase enzyme that catalyzes the formation of phosphatidate from diacylglycerol. Unlike the diacylglycerol kinases from bacteria, plants, and animals, the yeast enzyme utilizes CTP, instead of ATP, as the phosphate donor in the reacti ... >> More

  The Saccharomyces cerevisiae DGK1 gene encodes a diacylglycerol kinase enzyme that catalyzes the formation of phosphatidate from diacylglycerol. Unlike the diacylglycerol kinases from bacteria, plants, and animals, the yeast enzyme utilizes CTP, instead of ATP, as the phosphate donor in the reaction. Dgk1p contains a CTP transferase domain that is present in the SEC59-encoded dolichol kinase and CDS1-encoded CDP-diacylglycerol synthase enzymes. Deletion analysis showed that the CTP transferase domain was sufficient for diacylglycerol kinase activity. Point mutations (R76A, K77A, D177A, and G184A) of conserved residues within the CTP transferase domain caused a loss of diacylglycerol kinase activity. Analysis of DGK1 alleles showed that the in vivo functions of Dgk1p were specifically due to its diacylglycerol kinase activity. The DGK1-encoded enzyme had a pH optimum at 7.0-7.5, required Ca(2+) or Mg(2+) ions for activity, was potently inhibited by N-ethylmaleimide, and was labile at temperatures above 40 degrees C. The enzyme exhibited positive cooperative (Hill number = 2.5) kinetics with respect to diacylglycerol (apparent K(m) = 6.5 mol %) and saturation kinetics with respect to CTP (apparent K(m) = 0.3 mm). dCTP was both a substrate (apparent K(m) = 0.4 mm) and competitive inhibitor (apparent K(i) = 0.4 mm) of the enzyme. Diacylglycerol kinase activity was stimulated by major membrane phospholipids and was inhibited by CDP-diacylglycerol and sphingoid bases. << Less

  J. Biol. Chem. 283:20443-20453(2008) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.

• An unconventional diacylglycerol kinase that regulates phospholipid synthesis and nuclear membrane growth.

  Han G.-S., O'Hara L., Carman G.M., Siniossoglou S.

  Changes in nuclear size and shape during the cell cycle or during development require coordinated nuclear membrane remodeling, but the underlying molecular events are largely unknown. We have shown previously that the activity of the conserved phosphatidate phosphatase Pah1p/Smp2p regulates nuclea ... >> More

  Changes in nuclear size and shape during the cell cycle or during development require coordinated nuclear membrane remodeling, but the underlying molecular events are largely unknown. We have shown previously that the activity of the conserved phosphatidate phosphatase Pah1p/Smp2p regulates nuclear structure in yeast by controlling phospholipid synthesis and membrane biogenesis at the nuclear envelope. Two screens for novel regulators of phosphatidate led to the identification of DGK1. We show that Dgk1p is a unique diacylglycerol kinase that uses CTP, instead of ATP, to generate phosphatidate. DGK1 counteracts the activity of PAH1 at the nuclear envelope by controlling phosphatidate levels. Overexpression of DGK1 causes the appearance of phosphatidate-enriched membranes around the nucleus and leads to its expansion, without proliferating the cortical endoplasmic reticulum membrane. Mutations that decrease phosphatidate levels decrease nuclear membrane growth in pah1Delta cells. We propose that phosphatidate metabolism is a critical factor determining nuclear structure by regulating nuclear membrane biogenesis. << Less

  J. Biol. Chem. 283:20433-20442(2008) [PubMed] [EuropePMC]