Publications

• Kinetic and structural characterization of NUDT15 and NUDT18 as catalysts of isoprene pyrophosphate hydrolysis.

  Scaletti E.R., Unterlass J.E., Almlof I., Koolmeister T., Vallin K.S., Kapsitidou D., Tsuber V., Helleday T., Stenmark P., Jemth A.S.

  Isoprene pyrophosphates play a crucial role in the synthesis of a diverse array of essential nonsterol and sterol biomolecules and serve as substrates for posttranslational isoprenylation of proteins, enabling specific anchoring to cellular membranes. Hydrolysis of isoprene pyrophosphates would be ... >> More

  Isoprene pyrophosphates play a crucial role in the synthesis of a diverse array of essential nonsterol and sterol biomolecules and serve as substrates for posttranslational isoprenylation of proteins, enabling specific anchoring to cellular membranes. Hydrolysis of isoprene pyrophosphates would be a means to modulate their levels, downstream products, and protein isoprenylation. While NUDIX hydrolases from plants have been described to catalyze the hydrolysis of isoprene pyrophosphates, homologous enzymes with this function in animals have not yet been reported. In this study, we screened an extensive panel of human NUDIX hydrolases for activity in hydrolyzing isoprene pyrophosphates. We found that human nucleotide triphosphate diphosphatase NUDT15 and 8-oxo-dGDP phosphatase NUDT18 efficiently catalyze the hydrolysis of several physiologically relevant isoprene pyrophosphates. Notably, we demonstrate that geranyl pyrophosphate is an excellent substrate for NUDT18, with a catalytic efficiency of 2.1 × 10⁵ m^-1·s^-1, thus making it the best substrate identified for NUDT18 to date. Similarly, geranyl pyrophosphate proved to be the best isoprene pyrophosphate substrate for NUDT15, with a catalytic efficiency of 4.0 × 10⁴ M^-1·s^-1. LC-MS analysis of NUDT15 and NUDT18 catalyzed isoprene pyrophosphate hydrolysis revealed the generation of the corresponding monophosphates and inorganic phosphate. Furthermore, we solved the crystal structure of NUDT15 in complex with the hydrolysis product geranyl phosphate at a resolution of 1.70 Å. This structure revealed that the active site nicely accommodates the hydrophobic isoprenoid moiety and helped identify key binding residues. Our findings imply that isoprene pyrophosphates are endogenous substrates of NUDT15 and NUDT18, suggesting they are involved in animal isoprene pyrophosphate metabolism. << Less

  FEBS J 291:4301-4322(2024) [PubMed] [EuropePMC]

  This publication is cited by 7 other entries.

• Isopentenyl diphosphate/dimethylallyl diphosphate-specific Nudix hydrolase from the methanogenic archaeon Methanosarcina mazei.

  Ishibashi Y., Matsushima N., Ito T., Hemmi H.

  Nudix hydrolases typically catalyze the hydrolysis of nucleoside diphosphate linked to moiety X and yield nucleoside monophosphate and X-phosphate, while some of them hydrolyze a terminal diphosphate group of non-nucleosidic compounds and convert it into a phosphate group. Although the number of N ... >> More

  Nudix hydrolases typically catalyze the hydrolysis of nucleoside diphosphate linked to moiety X and yield nucleoside monophosphate and X-phosphate, while some of them hydrolyze a terminal diphosphate group of non-nucleosidic compounds and convert it into a phosphate group. Although the number of Nudix hydrolases is usually limited in archaea comparing with those in bacteria and eukaryotes, the physiological functions of most archaeal Nudix hydrolases remain unknown. In this study, a Nudix hydrolase family protein, MM_2582, from the methanogenic archaeon Methanosarcina mazei was recombinantly expressed in Escherichia coli, purified, and characterized. This recombinant protein shows higher hydrolase activity toward isopentenyl diphosphate and short-chain prenyl diphosphates than that toward nucleosidic compounds. Kinetic studies demonstrated that the archaeal enzyme prefers isopentenyl diphosphate and dimethylallyl diphosphate, which suggests its role in the biosynthesis of prenylated flavin mononucleotide, a recently discovered coenzyme that is required, for example, in the archaea-specific modified mevalonate pathway. << Less

  Biosci. Biotechnol. Biochem. 86:246-253(2022) [PubMed] [EuropePMC]

  This publication is cited by 3 other entries.

• PLANT VOLATILES. Biosynthesis of monoterpene scent compounds in roses.

  Magnard J.L., Roccia A., Caissard J.C., Vergne P., Sun P., Hecquet R., Dubois A., Hibrand-Saint Oyant L., Jullien F., Nicole F., Raymond O., Huguet S., Baltenweck R., Meyer S., Claudel P., Jeauffre J., Rohmer M., Foucher F., Hugueney P., Bendahmane M., Baudino S.

  The scent of roses (Rosa x hybrida) is composed of hundreds of volatile molecules. Monoterpenes represent up to 70% percent of the scent content in some cultivars, such as the Papa Meilland rose. Monoterpene biosynthesis in plants relies on plastid-localized terpene synthases. Combining transcript ... >> More

  The scent of roses (Rosa x hybrida) is composed of hundreds of volatile molecules. Monoterpenes represent up to 70% percent of the scent content in some cultivars, such as the Papa Meilland rose. Monoterpene biosynthesis in plants relies on plastid-localized terpene synthases. Combining transcriptomic and genetic approaches, we show that the Nudix hydrolase RhNUDX1, localized in the cytoplasm, is part of a pathway for the biosynthesis of free monoterpene alcohols that contribute to fragrance in roses. The RhNUDX1 protein shows geranyl diphosphate diphosphohydrolase activity in vitro and supports geraniol biosynthesis in planta. << Less

  Science 349:81-83(2015) [PubMed] [EuropePMC]

• Functional characterization of the atypical integral membrane lipid phosphatase PDP1/PPAPDC2 identifies a pathway for interconversion of isoprenols and isoprenoid phosphates in mammalian cells.

  Miriyala S., Subramanian T., Panchatcharam M., Ren H., McDermott M.I., Sunkara M., Drennan T., Smyth S.S., Spielmann H.P., Morris A.J.

  The polyisoprenoid diphosphates farnesyl diphosphate (FPP) and geranylgeranyl diphosphate (GGPP) are intermediates in the synthesis of cholesterol and related sterols by the mevalonate pathway and precursors for the addition of isoprenyl anchors to many membrane proteins. We developed tandem mass ... >> More

  The polyisoprenoid diphosphates farnesyl diphosphate (FPP) and geranylgeranyl diphosphate (GGPP) are intermediates in the synthesis of cholesterol and related sterols by the mevalonate pathway and precursors for the addition of isoprenyl anchors to many membrane proteins. We developed tandem mass spectrometry assays to evaluate polyisoprenoid diphosphate phosphatase activity of an unusual integral membrane lipid enzyme: type 1 polyisoprenoid diphosphate phosphatase encoded by the PPAPDC2 gene (PDP1/PPAPDC2). In vitro, recombinant PDP1/PPAPDC2 preferentially hydrolyzed polyisoprenoid diphosphates, including FPP and GGPP over a variety of glycerol- and sphingo-phospholipid substrates. Overexpression of mammalian PDP1/PPAPDC2 in budding yeast depletes cellular pools of FPP leading to growth defects and sterol auxotrophy. In mammalian cells, PDP1/PPAPDC2 localizes to the endoplasmic reticulum and nuclear envelope and, unlike the structurally related lipid phosphate phosphatases, is predicted to be oriented with key residues of its catalytic domain facing the cytoplasmic face of the membrane. Studies using synthetic isoprenols with chemical properties that facilitate detection by mass spectrometry identify a pathway for interconversion of isoprenols and isoprenoid diphosphates in intact mammalian cells and demonstrate a role for PDP1/PPAPDC2 in this process. Overexpression of PDP1/PPAPDC2 in mammalian cells substantially decreases protein isoprenylation and results in defects in cell growth and cytoskeletal organization that are associated with dysregulation of Rho family GTPases. Taken together, these results focus attention on integral membrane lipid phosphatases as regulators of isoprenoid phosphate metabolism and suggest that PDP1/PPAPDC2 is a functional isoprenoid diphosphate phosphatase. << Less

  J. Biol. Chem. 285:13918-13929(2010) [PubMed] [EuropePMC]

  This publication is cited by 7 other entries.