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The Journal of Biological Chemistry Feb 2022Ecto-nucleotide pyrophosphatase/phosphodiesterase (ENPP) family members (ENPP1-7) have been implicated in key biological and pathophysiological processes, including... (Review)
Review
Ecto-nucleotide pyrophosphatase/phosphodiesterase (ENPP) family members (ENPP1-7) have been implicated in key biological and pathophysiological processes, including nucleotide and phospholipid signaling, bone mineralization, fibrotic diseases, and tumor-associated immune cell infiltration. ENPPs are single-pass transmembrane ecto-enzymes, with notable exceptions of ENPP2 (Autotaxin) and ENNP6, which are secreted and glycosylphosphatidylinositol (GPI)-anchored, respectively. ENNP1 and ENNP2 are the best characterized and functionally the most interesting members. Here, we review the structural features of ENPP1-7 to understand how they evolved to accommodate specific substrates and mediate different biological activities. ENPPs are defined by a conserved phosphodiesterase (PDE) domain. In ENPP1-3, the PDE domain is flanked by two N-terminal somatomedin B-like domains and a C-terminal inactive nuclease domain that confers structural stability, whereas ENPP4-7 only possess the PDE domain. Structural differences in the substrate-binding site endow each protein with unique characteristics. Thus, ENPP1, ENPP3, ENPP4, and ENPP5 hydrolyze nucleotides, whereas ENPP2, ENPP6, and ENNP7 evolved as phospholipases through adaptions in the catalytic domain. These adaptations explain the different biological and pathophysiological functions of individual members. Understanding the ENPP members as a whole advances our insights into common mechanisms, highlights their functional diversity, and helps to explore new biological roles.
Topics: Catalytic Domain; Nucleotides; Phosphoric Diester Hydrolases; Pyrophosphatases; Signal Transduction; Structure-Activity Relationship
PubMed: 34958798
DOI: 10.1016/j.jbc.2021.101526 -
Cell Feb 2023Although many prokaryotes have glycolysis alternatives, it's considered as the only energy-generating glucose catabolic pathway in eukaryotes. Here, we managed to create...
Although many prokaryotes have glycolysis alternatives, it's considered as the only energy-generating glucose catabolic pathway in eukaryotes. Here, we managed to create a hybrid-glycolysis yeast. Subsequently, we identified an inositol pyrophosphatase encoded by OCA5 that could regulate glycolysis and respiration by adjusting 5-diphosphoinositol 1,2,3,4,6-pentakisphosphate (5-InsP) levels. 5-InsP levels could regulate the expression of genes involved in glycolysis and respiration, representing a global mechanism that could sense ATP levels and regulate central carbon metabolism. The hybrid-glycolysis yeast did not produce ethanol during growth under excess glucose and could produce 2.68 g/L free fatty acids, which is the highest reported production in shake flask of Saccharomyces cerevisiae. This study demonstrated the significance of hybrid-glycolysis yeast and determined Oca5 as an inositol pyrophosphatase controlling the balance between glycolysis and respiration, which may shed light on the role of inositol pyrophosphates in regulating eukaryotic metabolism.
Topics: Saccharomyces cerevisiae; Diphosphates; Saccharomyces cerevisiae Proteins; Inositol Phosphates; Glycolysis; Respiration; Pyrophosphatases; Glucose
PubMed: 36758548
DOI: 10.1016/j.cell.2023.01.014 -
Proceedings of the National Academy of... May 2022The metazoan innate immune second messenger 2′3′-cGAMP is present both inside and outside cells. However, only extracellular cGAMP can be negatively regulated by the...
The metazoan innate immune second messenger 2′3′-cGAMP is present both inside and outside cells. However, only extracellular cGAMP can be negatively regulated by the extracellular hydrolase ENPP1. Here, we determine whether ENPP1’s regulation of extracellular cGAMP is a ubiquitous mechanism of attenuating stimulator of interferon genes (STING) signaling. We identified ENPP1H362A, a point mutation that cannot degrade the 2′-5′ linkage in cGAMP while maintaining otherwise normal function. The selectivity of this histidine is conserved down to bacterial nucleotide pyrophosphatase/phosphodiesterase (NPP), allowing structural analysis and suggesting an unexplored ancient history of 2′-5′ cyclic dinucleotides. Enpp1H362A mice demonstrated that extracellular cGAMP is not responsible for the devastating phenotype in ENPP1-null humans and mice but is responsible for antiviral immunity and systemic inflammation. Our data define extracellular cGAMP as a pivotal STING activator, identify an evolutionarily critical role for ENPP1 in regulating inflammation, and suggest a therapeutic strategy for viral and inflammatory conditions by manipulating ENPP1 activity.
Topics: Animals; Humans; Immunity, Innate; Inflammation; Membrane Proteins; Mice; Nucleotides, Cyclic; Phosphoric Diester Hydrolases; Pyrophosphatases; Signal Transduction
PubMed: 35588451
DOI: 10.1073/pnas.2119189119 -
Proceedings of the National Academy of... Dec 2023Ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1) expression correlates with poor prognosis in many cancers, and we previously discovered that ENPP1 is the...
Ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1) expression correlates with poor prognosis in many cancers, and we previously discovered that ENPP1 is the dominant hydrolase of extracellular cGAMP: a cancer-cell-produced immunotransmitter that activates the anticancer stimulator of interferon genes (STING) pathway. However, ENPP1 has other catalytic activities and the molecular and cellular mechanisms contributing to its tumorigenic effects remain unclear. Here, using single-cell RNA-seq, we show that ENPP1 in both cancer and normal tissues drives primary breast tumor growth and metastasis by dampening extracellular 2'3'-cyclic-GMP-AMP (cGAMP)-STING-mediated antitumoral immunity. ENPP1 loss-of-function in both cancer cells and normal tissues slowed primary tumor growth and abolished metastasis. Selectively abolishing the cGAMP hydrolysis activity of ENPP1 phenocopied ENPP1 knockout in a STING-dependent manner, demonstrating that restoration of paracrine cGAMP-STING signaling is the dominant anti-cancer mechanism of ENPP1 inhibition. Finally, expression in breast tumors deterministically predicated whether patients would remain free of distant metastasis after pembrolizumab (anti-PD-1) treatment followed by surgery. Altogether, ENPP1 blockade represents a strategy to exploit cancer-produced extracellular cGAMP for controlled local activation of STING and is therefore a promising therapeutic approach against breast cancer.
Topics: Female; Humans; Breast Neoplasms; Immunity, Innate; Membrane Proteins; Phosphoric Diester Hydrolases; Pyrophosphatases
PubMed: 38117852
DOI: 10.1073/pnas.2313693120 -
Cellular and Molecular Life Sciences :... Apr 2021Extracellular NAD represents a key signaling molecule in different physiological and pathological conditions. It exerts such function both directly, through the... (Review)
Review
Extracellular NAD represents a key signaling molecule in different physiological and pathological conditions. It exerts such function both directly, through the activation of specific purinergic receptors, or indirectly, serving as substrate of ectoenzymes, such as CD73, nucleotide pyrophosphatase/phosphodiesterase 1, CD38 and its paralog CD157, and ecto ADP ribosyltransferases. By hydrolyzing NAD, these enzymes dictate extracellular NAD availability, thus regulating its direct signaling role. In addition, they can generate from NAD smaller signaling molecules, like the immunomodulator adenosine, or they can use NAD to ADP-ribosylate various extracellular proteins and membrane receptors, with significant impact on the control of immunity, inflammatory response, tumorigenesis, and other diseases. Besides, they release from NAD several pyridine metabolites that can be taken up by the cell for the intracellular regeneration of NAD itself. The extracellular environment also hosts nicotinamide phosphoribosyltransferase and nicotinic acid phosphoribosyltransferase, which inside the cell catalyze key reactions in NAD salvaging pathways. The extracellular forms of these enzymes behave as cytokines, with pro-inflammatory functions. This review summarizes the current knowledge on the extracellular NAD metabolome and describes the major biochemical properties of the enzymes involved in extracellular NAD metabolism, focusing on the contribution of their catalytic activities to the biological function. By uncovering the controversies and gaps in their characterization, further research directions are suggested, also to better exploit the great potential of these enzymes as therapeutic targets in various human diseases.
Topics: ADP Ribose Transferases; Animals; Disease; Humans; Metabolome; NAD; Pentosyltransferases; Pyrophosphatases; Signal Transduction
PubMed: 33755743
DOI: 10.1007/s00018-020-03742-1 -
Biomolecules Jan 2022Most free-living organisms encode for a deoxyuridine triphosphate nucleotidohydrolase (dUTPase; EC 3.6.1.23). dUTPases represent a family of metalloenzymes that catalyze... (Review)
Review
Most free-living organisms encode for a deoxyuridine triphosphate nucleotidohydrolase (dUTPase; EC 3.6.1.23). dUTPases represent a family of metalloenzymes that catalyze the hydrolysis of dUTP to dUMP and pyrophosphate, preventing dUTP from being incorporated into DNA by DNA polymerases, maintaining a low dUTP/dTTP pool ratio and providing a necessary precursor for dTTP biosynthesis. Thus, dUTPases are involved in maintaining genomic integrity by preventing the uracilation of DNA. Many DNA-containing viruses, which infect mammals also encode for a dUTPase. This review will summarize studies demonstrating that, in addition to their classical enzymatic activity, some dUTPases possess novel functions that modulate the host innate immune response.
Topics: Animals; DNA; Immunity, Innate; Mammals; Pyrophosphatases
PubMed: 35204728
DOI: 10.3390/biom12020227 -
Frontiers in Immunology 2023Ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1) is a type II transmembrane glycoprotein expressed in many tissues. High expression levels of ENPP1 have been...
Ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1) is a type II transmembrane glycoprotein expressed in many tissues. High expression levels of ENPP1 have been observed in many cancer types such as lung cancer, ovarian cancer, and breast cancer. Such overexpression has been associated with poor prognosis in these diseases. Hence, ENPP1 is a potential target for immunotherapy across multiple cancers. Here, we isolated and characterized two high-affinity and specific anti-ENPP1 Fab antibody candidates, 17 and 3G12, from large phage-displayed human Fab libraries. After conversion to IgG1, the binding of both antibodies increased significantly due to avidity effects. Based on these antibodies, we generated antibody-drug conjugates (ADCs), IgG-based bispecific T-cell engagers (IbTEs), and CAR T-cells which all exhibited potent killing of ENPP1-expressing cells. Thus, these various antibody-derived modalities are promising therapeutic candidates for cancers expressing human ENPP1.
Topics: Humans; Female; Phosphoric Diester Hydrolases; Immunoconjugates; Breast Neoplasms; Immunoglobulin G; Pyrophosphatases
PubMed: 36761762
DOI: 10.3389/fimmu.2023.1070492 -
Microbiology (Reading, England) Dec 2020Nudix proteins catalyse hydrolysis of pyrophosphate bonds in a variety of substrates and are ubiquitous in all domains of life. Their widespread presence and broad... (Review)
Review
Nudix proteins catalyse hydrolysis of pyrophosphate bonds in a variety of substrates and are ubiquitous in all domains of life. Their widespread presence and broad substrate specificity suggest that they have important cellular functions. In this review, we summarize the state of knowledge on microbial Nudix proteins involved in pathogenesis.
Topics: Amino Acid Sequence; Bacterial Proteins; Diphosphates; Pyrophosphatases; Sequence Alignment; Viral Proteins; Virulence; Virulence Factors; Nudix Hydrolases
PubMed: 33253082
DOI: 10.1099/mic.0.000993 -
Molecules (Basel, Switzerland) Apr 2021Inorganic pyrophosphatase (PPase) is a ubiquitous enzyme that converts pyrophosphate (PP) to phosphate and, in this way, controls numerous biosynthetic reactions that... (Review)
Review
Inorganic pyrophosphatase (PPase) is a ubiquitous enzyme that converts pyrophosphate (PP) to phosphate and, in this way, controls numerous biosynthetic reactions that produce PP as a byproduct. PPase activity is generally assayed by measuring the product of the hydrolysis reaction, phosphate. This reaction is reversible, allowing PP synthesis measurements and making PPase an excellent model enzyme for the study of phosphoanhydride bond formation. Here we summarize our long-time experience in measuring PPase activity and overview three types of the assay that are found most useful for (a) low-substrate continuous monitoring of PP hydrolysis, (b) continuous and fixed-time measurements of PP synthesis, and (c) high-throughput procedure for screening purposes. The assays are based on the color reactions between phosphomolybdic acid and triphenylmethane dyes or use a coupled ATP sulfurylase/luciferase enzyme assay. We also provide procedures to estimate initial velocity from the product formation curve and calculate the assay medium's composition, whose components are involved in multiple equilibria.
Topics: Diphosphates; Enzyme Assays; Humans; Hydrolysis; Inorganic Pyrophosphatase; Luciferases; Phosphates
PubMed: 33919593
DOI: 10.3390/molecules26082356 -
Microbiology (Reading, England) Dec 2020
Topics: Bacillus; Biological Control Agents; COVID-19; Genetic Techniques; Humans; Microbiology; Molecular Imaging; Periodicals as Topic; Pyrophosphatases; SARS-CoV-2; Vibrio; Nudix Hydrolases
PubMed: 33353584
DOI: 10.1099/mic.0.001019