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Circulation Jun 2013
Topics: Animals; Diphosphates; Male; Progeria; Vascular Calcification
PubMed: 23690467
DOI: 10.1161/CIRCULATIONAHA.113.003341 -
Molecules (Basel, Switzerland) Oct 2020Inositol pyrophosphates (PP-InsPs) comprise an important group of intracellular, diffusible cellular signals that a wide range of biological processes throughout the... (Review)
Review
Inositol pyrophosphates (PP-InsPs) comprise an important group of intracellular, diffusible cellular signals that a wide range of biological processes throughout the yeast, plant, and animal kingdoms. It has been difficult to gain a molecular-level mechanistic understanding of the actions of these molecules, due to their highly phosphorylated nature, their low levels, and their rapid metabolic turnover. More recently, these obstacles to success are being surmounted by the chemical synthesis of a number of insightful PP-InsP analogs. This review will describe these analogs and will indicate the important chemical and biological information gained by using them.
Topics: Animals; Diphosphates; Humans; Inositol; Metabolic Networks and Pathways; Phosphorylation; Protein Domains; Protein Stability
PubMed: 33023101
DOI: 10.3390/molecules25194515 -
MBio Oct 2022Prenyldiphosphate synthases catalyze the reaction of allylic diphosphates with one or more isopentenyl diphosphate molecules to form compounds such as farnesyl...
Prenyldiphosphate synthases catalyze the reaction of allylic diphosphates with one or more isopentenyl diphosphate molecules to form compounds such as farnesyl diphosphate, used in, e.g., sterol biosynthesis and protein prenylation, as well as longer "polyprenyl" diphosphates, used in ubiquinone and menaquinone biosynthesis. Quinones play an essential role in electron transport and are associated with the inner mitochondrial membrane due to the presence of the polyprenyl group. In this work, we investigated the synthesis of the polyprenyl diphosphate that alkylates the ubiquinone ring precursor in Toxoplasma gondii, an opportunistic pathogen that causes serious disease in immunocompromised patients and the unborn fetus. The enzyme that catalyzes this early step of the ubiquinone synthesis is Coq1 (TgCoq1), and we show that it produces the C35 species heptaprenyl diphosphate. TgCoq1 localizes to the mitochondrion and is essential for T. gondii growth. We demonstrate that the growth defect of a T. gondii TgCoq1 mutant is rescued by complementation with a homologous gene or with a (C45) solanesyl diphosphate synthase from Trypanosoma cruzi (TcSPPS). We find that a lipophilic bisphosphonate (BPH-1218) inhibits T. gondii growth at low-nanomolar concentrations, while overexpression of the TgCoq1 enzyme dramatically reduced growth inhibition by the bisphosphonate. Both the severe growth defect of the mutant and the inhibition by BPH-1218 were rescued by supplementation with a long-chain (C30) ubiquinone (UQ). Importantly, BPH-1218 also protected mice against a lethal T. gondii infection. TgCoq1 thus represents a potential drug target that could be exploited for improved chemotherapy of toxoplasmosis. Millions of people are infected with Toxoplasma gondii, and the available treatment for toxoplasmosis is not ideal. Most of the drugs currently used are only effective for the acute infection, and treatment can trigger serious side effects requiring changes in the therapeutic approach. There is, therefore, a compelling need for safe and effective treatments for toxoplasmosis. In this work, we characterize an enzyme of the mitochondrion of T. gondii that can be inhibited by an isoprenoid pathway inhibitor. We present evidence that demonstrates that inhibition of the enzyme is linked to parasite death. In addition, the inhibitor can protect mice against a lethal dose of T. gondii. Our results thus reveal a promising chemotherapeutic target for the development of new medicines for toxoplasmosis.
Topics: Animals; Mice; Diphosphates; Diphosphonates; Sterols; Toxoplasma; Toxoplasmosis; Ubiquinone; Vitamin K 2
PubMed: 36129297
DOI: 10.1128/mbio.01966-22 -
Plant Molecular Biology Dec 2011Norway spruce (Picea abies) defends itself against herbivores and pathogens by formation of traumatic resin ducts filled with terpenoid-based oleoresin. An important...
Norway spruce (Picea abies) defends itself against herbivores and pathogens by formation of traumatic resin ducts filled with terpenoid-based oleoresin. An important group of enzymes in terpenoid biosynthesis are the short-chain isoprenyl diphosphate synthases which produce geranyl diphosphate (C(10)), farnesyl diphosphate (C(15)), and geranylgeranyl diphosphate (C(20)) as precursors of monoterpenes, sesquiterpenes, and diterpene resin acids, respectively. After treatment with methyl jasmonate (MJ) we investigated the expression of all isoprenyl diphosphate synthase genes characterized to date from Norway spruce and correlated this with formation of traumatic resin ducts and terpene accumulation. Formation of traumatic resin ducts correlated with higher amounts of monoterpenes, sesquiterpenes and diterpene resin acids and an upregulation of isoprenyl diphosphate synthase genes producing geranyl diphosphate or geranylgeranyl diphosphate. Among defense hormones, jasmonate and jasmonate-isoleucine conjugate accumulated to higher levels in trees with extensive traumatic resin duct formation, whereas salicylate did not. Jasmonate and ethylene are likely to both be involved in formation of traumatic resin ducts based on elevated transcripts of genes encoding lipoxygenase and 1-aminocyclopropane-1-carboxylic acid oxidase associated with resin duct formation. Other genes involved in defense signalling in other systems, mitogen-activated protein kinase3 and nonexpressor of pathogenesis-related gene1, were also associated with traumatic resin duct formation. These responses were detected not only at the site of MJ treatment, but also systemically up to 60 cm above the site of treatment on the trunk.
Topics: Acetates; Alkyl and Aryl Transferases; Cyclopentanes; Diphosphates; Diterpenes; Oxylipins; Picea; Plant Growth Regulators; Polyisoprenyl Phosphates; Sesquiterpenes; Terpenes
PubMed: 22002747
DOI: 10.1007/s11103-011-9832-7 -
Archives of Biochemistry and Biophysics Dec 2007The tightly coupled nature of the reaction sequence catalyzed by monoterpene synthases has prevented direct observation of the topologically required isomerization step...
The tightly coupled nature of the reaction sequence catalyzed by monoterpene synthases has prevented direct observation of the topologically required isomerization step leading from geranyl diphosphate to the enzyme-bound, tertiary allylic intermediate linalyl diphosphate, which then cyclizes to the various monoterpene skeletons. X-ray crystal structures of these enzymes complexed with suitable analogues of the substrate and intermediate could provide a clearer view of this universal, but cryptic, step of monoterpenoid cyclase catalysis. Toward this end, the functionally inert analogues 2-fluorogeranyl diphosphate, (+/-)-2-fluorolinalyl diphosphate, and (3R)- and (3S)-homolinalyl diphosphates (2,6-dimethyl-2-vinyl-5-heptenyl diphosphates) were prepared, and compared to the previously described substrate analogue 3-azageranyl diphosphate (3-aza-2,3-dihydrogeranyl diphosphate) as inhibitors and potential crystallization aids with two representative monoterpenoid cyclases, (-)-limonene synthase and (+)-bornyl diphosphate synthase. Although these enantioselective synthases readily distinguished between (3R)- and (3S)-homolinalyl diphosphates, both of which were more effective inhibitors than was 3-azageranyl diphosphate, the fluorinated analogues proved to be the most potent competitive inhibitors and have recently yielded informative liganded structures with limonene synthase.
Topics: Acyclic Monoterpenes; Diphosphates; Diterpenes; Enzyme Activation; Enzyme Inhibitors; Enzyme Stability; Intramolecular Lyases; Monoterpenes; Polyisoprenyl Phosphates
PubMed: 17949678
DOI: 10.1016/j.abb.2007.09.008 -
Bioscience, Biotechnology, and... Jul 2007To determine the substrate specificities of wild and mutated types of farnesyl diphosphate (FPP) synthases from Bacillus stearothermophilus, we examined the reactivities...
To determine the substrate specificities of wild and mutated types of farnesyl diphosphate (FPP) synthases from Bacillus stearothermophilus, we examined the reactivities of 8-hydroxygeranyl diphosphate (HOGPP) and 8-methoxygeranyl diphosphate (CH(3)OGPP) as allylic substrate homologs. The wild-type FPP synthase reaction of HOGPP (and CH(3)OGPP) with isopentenyl diphosphate (IPP) gave hydroxyfarnesyl- (and methoxyfarnesyl-) diphosphates that stopped at the first stage of condensation. On the other hand, with mutated type FPP synthase (Y81S), the former gave hydroxygeranylgeranyl diphosphate as the main double-condensation product together with hydroxyfarnesyl diphosphate as a single-condensation product and a small amount of hydroxygeranylfarnesyl diphosphate as a triple-condensation product. Moreover, the latter gave a double-condensation product, methoxygeranylgeranyl diphosphate, as the main product and only a trace of methoxyfarnesyl diphosphate was obtained.
Topics: Amino Acid Substitution; Diphosphates; Diterpenes; Geobacillus stearothermophilus; Geranyltranstransferase; Substrate Specificity
PubMed: 17617711
DOI: 10.1271/bbb.70067 -
Nature Communications May 2023Many proteins involved in eukaryotic phosphate homeostasis are regulated by SPX domains. In yeast, the vacuolar transporter chaperone (VTC) complex contains two such...
Many proteins involved in eukaryotic phosphate homeostasis are regulated by SPX domains. In yeast, the vacuolar transporter chaperone (VTC) complex contains two such domains, but mechanistic details of its regulation are not well understood. Here, we show at the atomic level how inositol pyrophosphates interact with SPX domains of subunits Vtc2 and Vtc3 to control the activity of the VTC complex. Vtc2 inhibits the catalytically active VTC subunit Vtc4 by homotypic SPX-SPX interactions via the conserved helix α1 and the previously undescribed helix α7. Binding of inositol pyrophosphates to Vtc2 abrogates this interaction, thus activating the VTC complex. Accordingly, VTC activation is also achieved by site-specific point mutations that disrupt the SPX-SPX interface. Structural data suggest that ligand binding induces reorientation of helix α1 and exposes the modifiable helix α7, which might facilitate its post-translational modification in vivo. The variable composition of these regions within the SPX domain family might contribute to the diversified SPX functions in eukaryotic phosphate homeostasis.
Topics: Saccharomyces cerevisiae; Diphosphates; Biological Transport; Homeostasis; Inositol Phosphates
PubMed: 37156835
DOI: 10.1038/s41467-023-38315-w -
Seminars in Arthritis and Rheumatism Jun 2021To explore the lived experience of people with calcium pyrophosphate deposition (CPPD) disease and the impact of this condition on their daily lives.
OBJECTIVE
To explore the lived experience of people with calcium pyrophosphate deposition (CPPD) disease and the impact of this condition on their daily lives.
METHODS
Patients with CPPD and their caregivers were invited to take part in a one-to-one (patient only) or paired (patient and caregiver) semi-structured interview. Interviews covered patients' diagnosis and treatment experiences, and the impact of CPPD on their daily lives. Transcribed interviews were analysed using inductive thematic analysis.
RESULTS
28 patient interviews, six of which included a caregiver, were conducted across five countries. Acute CPP crystal arthritis flares resulted in temporary but profound disability for most patients, disrupting their ability to go about day-to-day activities, and they sought immediate medical attention. CPPD+OA and chronic CPP crystal inflammatory arthritis presented patients with longer term limitations in daily lives. Patients and their caregivers described these disruptions and limitations, which included a reduced ability or inability to complete household and self-care tasks, exercise, socialise, work and drive. They also described how arthritis pain and resulting limitations adversely impacted upon patients' psychological wellbeing. Delays in referral to specialists and diagnostic uncertainty were described by many. Lack of appropriate treatment or access to treatments only upon worsening of symptoms impacted upon the length of time some patients spent in pain and with functional limitations.
CONCLUSION
This study is the first to demonstrate the wide-ranging impact of CPPD, and highlights the need for improved diagnosis, physician training, as well as greater emphasis upon finding targeted therapies to specifically treat CPPD.
Topics: Calcinosis; Calcium Pyrophosphate; Caregivers; Chondrocalcinosis; Diphosphates; Humans
PubMed: 33941385
DOI: 10.1016/j.semarthrit.2021.04.010 -
Nucleic Acids Research Jan 2024Enzymatic methods to quantify deoxyribonucleoside triphosphates have existed for decades. In contrast, no general enzymatic method to quantify ribonucleoside...
Enzymatic methods to quantify deoxyribonucleoside triphosphates have existed for decades. In contrast, no general enzymatic method to quantify ribonucleoside triphosphates (rNTPs), which drive almost all cellular processes and serve as precursors of RNA, exists to date. ATP can be measured with an enzymatic luminometric method employing firefly luciferase, but the quantification of other ribonucleoside mono-, di-, and triphosphates is still a challenge for a non-specialized laboratory and practically impossible without chromatography equipment. To allow feasible quantification of ribonucleoside phosphates in any laboratory with typical molecular biology and biochemistry tools, we developed a robust microplate assay based on real-time detection of the Broccoli RNA aptamer during in vitro transcription. The assay employs the bacteriophage T7 and SP6 RNA polymerases, two oligonucleotide templates encoding the 49-nucleotide Broccoli aptamer, and a high-affinity fluorogenic aptamer-binding dye to quantify each of the four canonical rNTPs. The inclusion of nucleoside mono- and diphosphate kinases in the assay reactions enabled the quantification of the mono- and diphosphate counterparts. The assay is inherently specific and tolerates concentrated tissue and cell extracts. In summary, we describe the first chromatography-free method to quantify ATP, ADP, AMP, GTP, GDP, GMP, UTP, UDP, UMP, CTP, CDP and CMP in biological samples.
Topics: Diphosphates; Nucleotides; Ribonucleotides; Biochemistry
PubMed: 38008466
DOI: 10.1093/nar/gkad1091 -
Journal of Bone and Mineral Research :... Feb 2017Extensive or persistent calcium phosphate deposition within soft tissues after severe traumatic injury or major orthopedic surgery can result in pain and loss of joint...
Extensive or persistent calcium phosphate deposition within soft tissues after severe traumatic injury or major orthopedic surgery can result in pain and loss of joint function. The pathophysiology of soft tissue calcification, including dystrophic calcification and heterotopic ossification (HO), is poorly understood; consequently, current treatments are suboptimal. Here, we show that plasmin protease activity prevents dystrophic calcification within injured skeletal muscle independent of its canonical fibrinolytic function. After muscle injury, dystrophic calcifications either can be resorbed during the process of tissue healing, persist, or become organized into mature bone (HO). Without sufficient plasmin activity, dystrophic calcifications persist after muscle injury and are sufficient to induce HO. Downregulating the primary inhibitor of plasmin (α2-antiplasmin) or treating with pyrophosphate analogues prevents dystrophic calcification and subsequent HO in vivo. Because plasmin also supports bone homeostasis and fracture repair, increasing plasmin activity represents the first pharmacologic strategy to prevent soft tissue calcification without adversely affecting systemic bone physiology or concurrent muscle and bone regeneration. © 2016 American Society for Bone and Mineral Research.
Topics: Animals; Calcinosis; Cardiotoxins; Diphosphates; Fibrinolysin; Fibrinolysis; Genetic Predisposition to Disease; Mice, Inbred C57BL; Muscle, Skeletal; Ossification, Heterotopic; Regeneration
PubMed: 27530373
DOI: 10.1002/jbmr.2973