-
Nefrologia : Publicacion Oficial de La... 2013ImmuKnow is an in vitro diagnosis method that uses patient samples of whole blood polyclonally stimulated with phytohaemagglutinin. It also measures adenosine... (Review)
Review
ImmuKnow is an in vitro diagnosis method that uses patient samples of whole blood polyclonally stimulated with phytohaemagglutinin. It also measures adenosine triphosphate (ATP) production by CD4+ T cells. The test aims to offer an objective and overall measurement of each individual's cellular immune response. The assay was designed with the idea of individually monitoring the immunosuppression administered to transplant patients. At the same time, it aims to help achieve a balance as a way of avoiding immunosuppression excess and the associated adverse effects (infections, cancer, etc.) or an immunosuppression defect and the subsequent risk of allograft rejection. The majority of studies that have evaluated its clinical usefulness display great diversity in terms of patient recruitment, the immunosuppressant treatment received, the clinical variables analysed and, above all, the time between performing ImmuKnow and the evaluated clinical event. The most consistent data show that this assay on CD4+ T cell functioning is useful for predicting the risk of infection in renal transplant patients. However, its use as a rejection risk indicator is unclear. Lastly, given the great variability of immune response amongst individuals and that of existing publications, it can be deduced that the isolated ImmuKnow value does not have diagnostic capacity and only individual serial monitoring could provide definitive assistance in clinical decision making and immunosuppressant treatment changes. Other aspects of ImmuKnow application in the clinical routine, such as assay cycles, require randomised prospective studies for more comprehensive information.
Topics: Adenosine Triphosphate; CD4-Positive T-Lymphocytes; Humans; Intracellular Membranes; Kidney Transplantation
PubMed: 23640115
DOI: 10.3265/Nefrologia.pre2012.Oct.11540 -
Biology Direct Mar 2022Bacteria and archaea produce an enormous diversity of modified peptides that are involved in various forms of inter-microbial conflicts or communication. A vast class...
BACKGROUND
Bacteria and archaea produce an enormous diversity of modified peptides that are involved in various forms of inter-microbial conflicts or communication. A vast class of such peptides are Ribosomally synthesized, Postranslationally modified Peptides (RiPPs), and a major group of RiPPs are graspetides, so named after ATP-grasp ligases that catalyze the formation of lactam and lactone linkages in these peptides. The diversity of graspetides, the multiple proteins encoded in the respective Biosynthetic Gene Clusters (BGCs) and their evolution have not been studied in full detail. In this work, we attempt a comprehensive analysis of the graspetide-encoding BGCs and report a variety of novel graspetide groups as well as ancillary proteins implicated in graspetide biosynthesis and expression.
RESULTS
We compiled a comprehensive, manually curated set of graspetides that includes 174 families including 115 new families with distinct patterns of amino acids implicated in macrocyclization and further modification, roughly tripling the known graspetide diversity. We derived signature motifs for the leader regions of graspetide precursors that could be used to facilitate graspetide prediction. Graspetide biosynthetic gene clusters and specific precursors were identified in bacterial divisions not previously known to encode RiPPs, in particular, the parasitic and symbiotic bacteria of the Candidate phyla radiation. We identified Bacteroides-specific biosynthetic gene clusters (BGC) that include remarkable diversity of graspetides encoded in the same loci which predicted to be modified by the same ATP-grasp ligase. We studied in details evolution of recently characterized chryseoviridin BGCs and showed that duplication and horizonal gene exchange both contribute to the diversification of the graspetides during evolution.
CONCLUSIONS
We demonstrate previously unsuspected diversity of graspetide sequences, even those associated with closely related ATP-grasp enzymes. Several previously unnoticed families of proteins associated with graspetide biosynthetic gene clusters are identified. The results of this work substantially expand the known diversity of RiPPs and can be harnessed to further advance approaches for their identification.
Topics: Adenosine Triphosphate; Bacteria; Multigene Family; Peptides; Phylogeny; Protein Processing, Post-Translational
PubMed: 35313954
DOI: 10.1186/s13062-022-00320-2 -
Journal of the American Chemical Society Jun 2018Bisphosphonates are a major class of drugs used to treat osteoporosis, Paget's disease, and cancer. They have been proposed to act by inhibiting one or more targets...
Bisphosphonates are a major class of drugs used to treat osteoporosis, Paget's disease, and cancer. They have been proposed to act by inhibiting one or more targets including protein prenylation, the epidermal growth factor receptor, or the adenine nucleotide translocase. Inhibition of the latter is due to formation in cells of analogs of ATP: the isopentenyl ester of ATP (ApppI) or an AppXp-type analog of ATP, such as AMP-clodronate (AppCClp). We screened both ApppI as well as AppCClp against a panel of 369 kinases finding potent inhibition of some tyrosine kinases by AppCClp, attributable to formation of a strong hydrogen bond between tyrosine and the terminal phosphonate. We then synthesized bisphosphonate preprodrugs that are converted in cells to other ATP-analogs, finding low nM kinase inhibitors that inhibited cell signaling pathways. These results help clarify our understanding of the mechanisms of action of bisphosphonates, potentially opening up new routes to the development of bone resorption, anticancer, and anti-inflammatory drug leads.
Topics: Adenosine Triphosphate; Cell Line, Tumor; Diphosphonates; Humans; Hydrogen Bonding; Models, Chemical; Prodrugs; Protein Kinase Inhibitors; Protein-Tyrosine Kinases; Signal Transduction
PubMed: 29787268
DOI: 10.1021/jacs.8b02363 -
Chemistry (Weinheim An Der Bergstrasse,... May 2020Simple and robust assays to monitor enzymatic ATP cleavage with high efficiency in real-time are scarce. To address this shortcoming, we developed fluorescently labelled...
Simple and robust assays to monitor enzymatic ATP cleavage with high efficiency in real-time are scarce. To address this shortcoming, we developed fluorescently labelled adenosine tri-, tetra- and pentaphosphate analogues of ATP. The novel ATP analogues bear - in contrast to earlier reports - only a single acridone-based dye at the terminal phosphate group. The dye's fluorescence is quenched by the adenine component of the ATP analogue and is restored upon cleavage of the phosphate chain and dissociation of the dye from the adenosine moiety. Thereby the activity of ATP-cleaving enzymes can be followed in real-time. We demonstrate this proficiency for ubiquitin activation by the ubiquitin-activating enzymes UBA1 and UBA6 which represents the first step in an enzymatic cascade leading to the covalent attachment of ubiquitin to substrate proteins, a process that is highly conserved from yeast to humans. We found that the efficiency to serve as cofactor for UBA1/UBA6 very much depends on the length of the phosphate chain of the ATP analogue: triphosphates are used poorly while pentaphosphates are most efficiently processed. Notably, the novel pentaphosphate-harbouring ATP analogue supersedes the efficiency of recently reported dual-dye labelled analogues and thus, is a promising candidate for broad applications.
Topics: Adenosine Triphosphate; Fluorescent Dyes; Humans; Ubiquitin-Activating Enzymes
PubMed: 32154932
DOI: 10.1002/chem.202001091 -
Current Neuropharmacology 2022P2X7 receptors (Rs) are prominent members of the P2XR family, which after binding ATP, open non-selective cationic channels, thereby allowing the transmembrane passage...
P2X7 receptors (Rs) are prominent members of the P2XR family, which after binding ATP, open non-selective cationic channels, thereby allowing the transmembrane passage of Na+, Ca, and K+. Long-lasting and repetitive stimulation of the receptor by its agonist leads to the formation of large membrane pores permeable for organic cations of up to 900 Da molecular size. These pores are believed to play a role in apoptosis and inflammation. P2X7Rs are located primarily at peripheral macrophages and microglial cells, the resident macrophages of the CNS. The coactivation of toll-like receptors 4 (TLR4) by lipopolysaccharide, a constituent of the cell membrane of gram-negative bacteria, and the P2X7R by ATP leads to the generation and release of the proinflammatory cytokines interleukin-1β (IL-1β), IL-6, and tumor necrosis factor-α. Together with the microglial release of chemokines, reactive oxygen and nitrogen species, proteases, and excitotoxic glutamate, these cytokines result in neurodegeneration. P2X7Rs were found not only to amplify various neurodegenerative illnesses, such as Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, and multiple sclerosis, but also to participate in a range of psychiatric diseases, such as major depression, bipolar disorder, schizophrenia, and autism spectrum disorder. Based on the prevention/reversal of neuroinflammation, pharmacological antagonists of P2X7Rs and their genetic deletion in animal experiments counteract these deleterious psychiatric conditions. Hence, brain penetrant P2X7R antagonists are potential therapeutics for psychiatric diseases, although the available evidence still needs to be extended and validated by further clinical data.
Topics: Animals; Receptors, Purinergic P2X7; Autism Spectrum Disorder; Microglia; Mental Disorders; Adenosine Triphosphate
PubMed: 35236262
DOI: 10.2174/1570159X20666220302152400 -
Plant Physiology Oct 2003
Review
Topics: Adenosine Triphosphate; Arabidopsis; Ion Channels; Plant Physiological Phenomena; Purines; Pyrimidines; Signal Transduction
PubMed: 14555773
DOI: 10.1104/pp.103.024091 -
Neurogastroenterology and Motility Sep 2016Adenosine 5'-triphosphate (ATP) is released extracellularly as a neurotransmitter and an autocrine or paracrine mediator in numerous systems, including the...
BACKGROUND
Adenosine 5'-triphosphate (ATP) is released extracellularly as a neurotransmitter and an autocrine or paracrine mediator in numerous systems, including the gastrointestinal tract. It is rapidly degraded to active and inactive metabolites by membrane-bound enzymes. Investigators frequently use inhibitors of ATP hydrolysis such as ARL-67156 and POM-1 to suppress the catabolism of ATP and prolong its effects in pharmacological studies. Our aim was to investigate directly the effects of ARL-67156 and POM-1 on the degradation of ATP and adenosine 5'-diphosphate (ADP) in mouse colonic muscles.
METHODS
The degradation of ATP and ADP was evaluated by superfusing tissues with 1,N(6) -etheno-ATP (eATP) and 1,N(6) -etheno-ADP (eADP) as substrates and monitoring the decrease in substrate and increase in products (i.e., eADP, eAMP, and e-adenosine) by high-performance liquid chromatography techniques with fluorescence detection. Relaxation responses to etheno-derivatized and non-derivatized ATP and ADP were examined in isometric tension experiments.
KEY RESULTS
ARL-67156 inhibits the degradation of ADP but not of ATP, whereas POM-1 inhibits the degradation of ATP but not of ADP in murine colonic muscles. Consequently, ARL-67156 enhances relaxation responses to both ATP and ADP, whereas POM-1 reduces relaxation to ATP and does not affect relaxation to ADP.
CONCLUSIONS & INFERENCES
Studies that use ARL-67156 to inhibit ATP degradation in smooth muscle likely evaluate responses to accumulated ADP rather than ATP. POM-1 appears to be a more selective inhibitor of ATP degradation in the mouse colon. The choice of pharmacological tools in studies on extracellular ATP signaling may affect the interpretation of experimental data in functional studies.
Topics: Adenosine Diphosphate; Adenosine Triphosphate; Animals; Colon; Mice
PubMed: 27060478
DOI: 10.1111/nmo.12836 -
The Journal of Physiology Dec 19681. The synthesis of ATP has been studied in human erythrocytes. Fresh cells showed no net synthesis of ATP when incubated with adenine or adenosine, although labelled...
1. The synthesis of ATP has been studied in human erythrocytes. Fresh cells showed no net synthesis of ATP when incubated with adenine or adenosine, although labelled adenine was incorporated into ATP in small amounts.2. Cold-stored cells (3-6 weeks old) became progressively depleted of adenine nucleotides but incubation with adenosine or adenine plus inosine restored the ATP concentration to normal within 4 hr. Incorporation of labelled adenine or adenosine into the ATP of incubated stored cells corresponded to net ATP synthesis by these cells.3. Synthesis of ATP from adenosine plus adenine together was 75% derived from adenine and only 25% from adenosine, indicating that nucleotide synthesis from adenine inhibits the simultaneous synthesis of nucleotide from adenosine.
Topics: Adenine; Adenosine Triphosphate; Cold Temperature; Erythrocytes; Humans; Male; Nucleosides; Time Factors
PubMed: 5723519
DOI: 10.1113/jphysiol.1968.sp008664 -
Biophysical Journal Jun 1985We have studied the inhibition of the contraction of glycerinated rabbit psoas muscle caused by ligands that bind to the ATPase site of myosin. Two ligands, adenosine 5'...
We have studied the inhibition of the contraction of glycerinated rabbit psoas muscle caused by ligands that bind to the ATPase site of myosin. Two ligands, adenosine 5' (beta, gamma-imido) triphosphate (AMPPNP) and pyrophosphate (PPi), decreased the force and stiffness developed in isometric contractions and the velocity of shortening of isotonic contractions. The force exerted by isometric fibers was measured as a function of MgATP in the presence and absence of a constant concentration of the ligands. As the MgATP concentration decreased, the inhibition of tension caused by the ligand increased, reaching approximately 50% at 25 microM MgATP and either 2 mM MgPPi or 2 mM MgAMPPNP. The maximum velocity of shortening was also measured as a function of MgATP concentration in the presence of 1 and 2 mM MgPPi and 2.5 and 5 mM MgAMPPNP. Both ligands acted as pure competitive inhibitors with Ki = 3.0 mM for PPi and 5.1 mM for MgAMPPNP. These data show that both ligands are weak inhibitors of the contraction of fibers. The results provided information on the energetics of actin-myosin-ligand states that occur in the portion of the cross-bridge cycle where MgATP binds to myosin. A simple analysis of the inhibition of velocity suggests that MgAMPPNP binds to the actomyosin complex at this step of the cycle with an effective affinity constant of approximately 2 X 10(2) M-1.
Topics: Adenosine Triphosphate; Adenylyl Imidodiphosphate; Animals; Diphosphates; Isometric Contraction; Kinetics; Muscle Contraction; Muscles; Rabbits
PubMed: 2990586
DOI: 10.1016/S0006-3495(85)83980-1 -
Applied Microbiology Sep 1973EXISTING DATA ON ADENOSINE TRIPHOSPHATE (ATP) POOLS IN MICROBES ARE DEFICIENT FOR TWO REASONS: (i) incomplete extractions of ATP, and (ii) the failure to take into... (Comparative Study)
Comparative Study
EXISTING DATA ON ADENOSINE TRIPHOSPHATE (ATP) POOLS IN MICROBES ARE DEFICIENT FOR TWO REASONS: (i) incomplete extractions of ATP, and (ii) the failure to take into account that the adverse effects of extracting procedures on standard ATP exert analogous effects on the ATP released from bacterial cells. Methods for correcting observed yields and calculating ATP pools have been demonstrated. Three bacterial species were used in the studies on extraction of ATP: Escherichia coli, Mycobacterium phlei, and Mycobacterium lepraemurium. Perchloric acid and n-butanol were disqualified because of their failure to extract total bacterial ATP even from E. coli and because of inconvenient procedures. The new extraction procedure had minimal effects on standard ATP, liberated 100% of the ATP pools from the three representative species of microbes, and caused no ionic imbalance or quenching of bioluminescence. This method involves vortexing of cell suspensions for 10 s with 23% chloroform (vol/vol), heating at 98 C for the required time (E. coli, 3 min; M. phlei, 5 min; M. lepraemurium, 10 min) and then 1 min at 98 C with vacuum to dry the samples. Heat or chloroform alone may suffice for some microbes and release total ATP from plant and animal cells.
Topics: Adenosine Triphosphate; Butanols; Chloroform; Escherichia coli; Hot Temperature; Indicators and Reagents; Luminescent Measurements; Methods; Mycobacterium; Mycobacterium lepraemurium; Perchlorates; Solvents; Species Specificity
PubMed: 4356463
DOI: 10.1128/am.26.3.399-403.1973