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Structure, regulation, interfacial enzymology, and functions of lipid metabolizing enzymes involved in signal transduction cascades
A major effort in our laboratory is focused on understanding the biochemistry and chemistry underlying the molecular aspects involved in regulating lipid metabolizing signaling enzymes and the physiological roles of this regulation. Control of lipid metabolizing enzymes involves the modulation of two key parameters; their sub-cellular distribution and their intrinsic enzymatic activity. Our studies have concentrated on three families of lipid-metabolizing signaling enzymes: diacylglycerol kinases, phospholipases D, and phospholipases C. Specific Areas of Interest Interfacial Enzymology of Lipid Metabolizing Signaling Enzymes: We are particularly interested in identifying the critical modulating proteins, lipids, and post-translational modifications that alter the localization and/or activity of lipid metabolizing enzymes. In these studies we consider the fact that these enzymes act as interfacial enzymes and their regulation includes a number of interfacial-dependent parameters. Our recent studies have identified some of the diacylglycerol metabolizing enzyme DGK-? (diacylglycerol kinase-theta) interfacial parameters that are altered upon neuronal depolarization. Further, our studies demonstrated that activation of DGK-? requires a protein that contains a polybasic region. We have recently obtained evidence that identifies at least one, if not only, activator binding domain on DGK-?. Enzyme Structure/Function Studies: We are also interested in the structural components of these enzymes that are critical for their distribution/re-distribution to specific sub-cellular compartments. Additionally, and to compliment the enzymology studies, we are interested in elucidating the catalytic mechanism(s) of these enzymes. These studies will be conducted partly in collaboration with Dr. Mario Amzel. Our long-term goal is to understand the biochemistry and chemistry of these enzymes and determine how changes in their sub-cellular localization and/or enzymatic activity affect their signaling functions. Physiological Functions of DGKs in Neurons: There is growing evidence that DGKs play physiological roles in mammalian neurons. This evidence includes cellular localization of specific isoforms, and the observations that likely modulate (a) susceptibility to epileptic seizures (DGK-?), (b) neuronal spine density (DGK-? and DGK-?), and (c) pre-synaptic glutamate release during DHPG (3,5-dihydroxyphenylglycine)-induced long-term potentiation (DGK-?). We are currently examining the role of DGK-? in glutamatergic neurons. These studies have initially focused on identifying the physiologic regulator of DGK-?, and test the hypothesis that this enzyme modulates induced glutamate release in these mammalian neurons. We discovered that DGK-? modulates glutamate release from cortical and hippocampal neurons in part by modulating synaptic vesicle cycling. These studies are conducted in collaboration with Dr. Rick Huganier’s laboratory.
Tu-Sekine, B., Goldschmidt, H, Raben, D.M. (2015) Diacylglycerol, Phosphatidic Acid, and their Metabolizing Enzymes in Synaptic Vesicle Recycling. Adv. Biol. Reg. Jan;57:147-52.
Petro E, and Raben DM. (2013) Bacterial expression strategies for several Sus scrofa diacylglycerol kinase alpha constructs: solubility challenges. Scientific Reports 2013;3:160.
Ueda S, Tu-Sekine B, Yamanoue M, Raben DM, and Shirai Y. (2013) The expression of diacylglycerol kinase theta during the organogenesis of mouse embryos. BMC Developmental Biology 2013, 13:35.
Bolduc D, Rahdar M, Tu-Sekine B, Sivakumaren SC, Raben DM, Amzel LM, Devreotes P, Gabelli SB, and Cole P. (2013) Phosphorylation-mediated PTEN conformational closure and deactivation revealed with protein semisynthesis. Elife 2013 Jul 9;2:e00691.
Tu-Sekine B, Goldschmidt H, Petro E, and Raben DM. (2013) Diacylglycerol Kinase Theta: Regulation and Stability. Adv. Biol. Reg. Jan;53(1):118-26.
Tu-Sekine B, and Raben DM. (2012) Dual Regulation of DGK-?: Polybasic Proteins Promote Activation by Phospholipids and Increase Substrate Affinity. J. Biol. Chem. 287(50):41619-41627.
Tu-Sekin, B, and Raben DM. (2011) Regulation and Roles of Neuronal Diacylglycerol Kinases: a Lipid Perspective. Crit. Rev. Biochem. Mol. Biol. Oct;46(5):353-64.
Mohan S, Tse CM, Gabelli SB, Sarker R, Cha B, Fahie K, Nadella M, Zachos NC, Tu-Sekine B, Raben DM, Amzel LM, Donowitz M. (2010) NHE3 Activity Is Dependent on Direct Phosphoinositide Binding at the N Terminus of Its Intracellular Cytosolic Region. J. Biol. Chem. 285(45): 34566-78.
Tu-Sekine, B. and Raben DM. (2010) Characterization of Cellular DGK?. Advances in Enzyme Reg. 50:81-94.
Link TM, Park U, Vonakis BM, Raben DM, Soloski M.J., Caterina MJ. (2010) TRPV2 plays a pivotal role in macrophage particle binding and phagocytosis. Nature Immunology Mar;11(3):232-9. Epub 2010 Jan 31.
Tu-Sekine and Raben DM. (2009) Regulation of DGK-? J. Cell Physiol. 220(3):548-52.
Raben DM and Wattenberg BW. (2009) Signaling at the Membrane Interface by the DGK/SK Enzyme Family. J Lipid Res 50th Anniversary Edition: J. Lipid Res. April Supplement: S35-S39.
Raben DM and Tu-Sekine B. (2008) Nuclear Localization Of Diacylglycerol Kinases: Regulation And Roles. Frontiers in Bioscience 13:590-597.
Wattenberg BW and Raben DM. (2007) Diacylglycerol Kinases Put the Brakes on Immune Function. Science STKE (398) pe43.
Tu-Sekine B, Ostroski M, and Raben DM. (2007) Modulation of DGK? Activity by ?-Thrombin and Phospholipids. Biochemistry, 46(3): 924 -932.