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Biochemical and Biophysical Research... Jan 2024Adenosine diphosphate (ADP) is a nucleotide that is structurally very similar to ATP but lacks one of the two high-energy bonds due to hydrolysis. In muscle studies, ADP...
Adenosine diphosphate (ADP) is a nucleotide that is structurally very similar to ATP but lacks one of the two high-energy bonds due to hydrolysis. In muscle studies, ADP is usually considered exclusively as a product formed during myosin cross-bridge cycling and is not otherwise involved in this process. In our study, we question the widely held view of ADP as a final product formed during muscle contraction. Using biophysical and biochemical methods, we managed to show that ADP can act as a substrate for myosins in at least three types of muscles: smooth and striated adductor muscles of bivalves (Mytilidae and Pectinidae), and also vertebrate skeletal muscles. According to our data, the differences in the effect of ATP and ADP on the optical, biochemical, and structural properties of actomyosins are exclusively quantitative. We explain the previous ideas about ADP as a compound capable of inhibiting the ATPase activity of actomyosin by the ability of ATP and ADP to depolymerize the polymeric myosin when the concentration in the medium reaches more than 0.3 mM.
Topics: Apyrase; Adenosine Triphosphate; Myosins; Actomyosin; Muscle, Skeletal; Adenosine Diphosphate; Actins; Kinetics
PubMed: 38096615
DOI: 10.1016/j.bbrc.2023.149371 -
Frontiers in Immunology 2021Chronic wounds are a public health problem worldwide, especially those related to diabetes. Besides being an enormous burden to patients, it challenges wound care...
Chronic wounds are a public health problem worldwide, especially those related to diabetes. Besides being an enormous burden to patients, it challenges wound care professionals and causes a great financial cost to health system. Considering the absence of effective treatments for chronic wounds, our aim was to better understand the pathophysiology of tissue repair in diabetes in order to find alternative strategies to accelerate wound healing. Nucleotides have been described as extracellular signaling molecules in different inflammatory processes, including tissue repair. Adenosine-5'-diphosphate (ADP) plays important roles in vascular and cellular response and is immediately released after tissue injury, mainly from platelets. However, despite the well described effect on platelet aggregation during inflammation and injury, little is known about the role of ADP on the multiple steps of tissue repair, particularly in skin wounds. Therefore, we used the full-thickness excisional wound model to evaluate the effect of local ADP application in wounds of diabetic mice. ADP accelerated cutaneous wound healing, improved new tissue formation, and increased both collagen deposition and transforming growth factor-β (TGF-β) production in the wound. These effects were mediated by P2Y receptor activation since they were inhibited by Clopidogrel (Clop) treatment, a P2Y receptor antagonist. Furthermore, P2Y receptor antagonist also blocked ADP-induced wound closure until day 7, suggesting its involvement early in repair process. Interestingly, ADP treatment increased the expression of P2Y and P2Y receptors in the wound. In parallel, ADP reduced reactive oxygen species (ROS) formation and tumor necrosis factor-α (TNF-α) levels, while increased IL-13 levels in the skin. Also, ADP increased the counts of neutrophils, eosinophils, mast cells, and gamma delta (γδ) T cells (Vγ4 and Vγ5 cells subtypes of γδ T cells), although reduced regulatory T (Tregs) cells in the lesion. In accordance, ADP increased fibroblast proliferation and migration, myofibroblast differentiation, and keratinocyte proliferation. In conclusion, we provide strong evidence that ADP acts as a pro-resolution mediator in diabetes-associated skin wounds and is a promising intervention target for this worldwide problem.
Topics: Adenosine Diphosphate; Administration, Cutaneous; Alloxan; Animals; Diabetes Mellitus, Experimental; Humans; Male; Mice; Purinergic P2Y Receptor Agonists; Receptors, Purinergic P2Y12; Skin; Wound Healing
PubMed: 33828561
DOI: 10.3389/fimmu.2021.651740 -
EMBO Reports Aug 2023The mitochondrial ADP/ATP carrier (SLC25A4), also called the adenine nucleotide translocase, imports ADP into the mitochondrial matrix and exports ATP, which are key...
The mitochondrial ADP/ATP carrier (SLC25A4), also called the adenine nucleotide translocase, imports ADP into the mitochondrial matrix and exports ATP, which are key steps in oxidative phosphorylation. Historically, the carrier was thought to form a homodimer and to operate by a sequential kinetic mechanism, which involves the formation of a ternary complex with the two exchanged substrates bound simultaneously. However, recent structural and functional data have demonstrated that the mitochondrial ADP/ATP carrier works as a monomer and has a single substrate binding site, which cannot be reconciled with a sequential kinetic mechanism. Here, we study the kinetic properties of the human mitochondrial ADP/ATP carrier by using proteoliposomes and transport robotics. We show that the Km/Vmax ratio is constant for all of the measured internal concentrations. Thus, in contrast to earlier claims, we conclude that the carrier operates with a ping-pong kinetic mechanism in which substrate exchange across the membrane occurs consecutively rather than simultaneously. These data unite the kinetic and structural models, showing that the carrier operates with an alternating access mechanism.
Topics: Humans; Mitochondrial ADP, ATP Translocases; Mitochondria; Adenosine Triphosphate; Adenosine Diphosphate; Kinetics; Adenine Nucleotide Translocator 1
PubMed: 37278158
DOI: 10.15252/embr.202357127 -
Proceedings of the National Academy of... May 2023Poly(ADP-ribose) (PAR) is a homopolymer of adenosine diphosphate ribose that is added to proteins as a posttranslational modification to regulate numerous cellular...
Poly(ADP-ribose) (PAR) is a homopolymer of adenosine diphosphate ribose that is added to proteins as a posttranslational modification to regulate numerous cellular processes. PAR also serves as a scaffold for protein binding in macromolecular complexes, including biomolecular condensates. It remains unclear how PAR achieves specific molecular recognition. Here, we use single-molecule fluorescence resonance energy transfer (smFRET) to evaluate PAR flexibility under different cation conditions. We demonstrate that, compared to RNA and DNA, PAR has a longer persistence length and undergoes a sharper transition from extended to compact states in physiologically relevant concentrations of various cations (Na, Mg, Ca, and spermine). We show that the degree of PAR compaction depends on the concentration and valency of cations. Furthermore, the intrinsically disordered protein FUS also served as a macromolecular cation to compact PAR. Taken together, our study reveals the inherent stiffness of PAR molecules, which undergo switch-like compaction in response to cation binding. This study indicates that a cationic environment may drive recognition specificity of PAR.
Topics: Poly Adenosine Diphosphate Ribose; Adenosine Diphosphate Ribose; Protein Processing, Post-Translational; Protein Binding; Cell Physiological Phenomena
PubMed: 37126687
DOI: 10.1073/pnas.2215068120 -
Urologie (Heidelberg, Germany) Dec 2023Metastatic castration-resistant prostate cancer (mCRPC) is a heterogeneous disease with varying clinical and molecular subtypes. Almost one-third of patients have... (Review)
Review
[Combination therapy with poly(adenosine diphosphate-ribose) polymerase (PARPi) and androgen receptor signaling pathway (ARPi) inhibitors for metastatic castration-resistant prostate cancer].
Metastatic castration-resistant prostate cancer (mCRPC) is a heterogeneous disease with varying clinical and molecular subtypes. Almost one-third of patients have abnormalities in homologous recombinant repair genes. Again, about one third of these mutations affect the BReast CAncer 1 or 2 (BRCA 1 or BRCA 2) genes, which generally render tumours receptive to treatment with poly(adenosine diphosphate-ribose) polymerase inhibitors (PARPi). In 2020 the PARPi olaparib was approved for the treatment of mCRPC after progression with a new hormonal drug (androgen receptor signaling pathway inhibitors, ARPi). In 2022 and 2023 approval of two combination therapies followed, each combining a PARPi and an ARPi (olaparib plus abiraterone and niraparib plus abiraterone). The combination of talazoparib plus enzalutamide will be approved soon. This article introduces the pivotal clinical trials that led to the approval of the respective substances, reports the side effects that may occur during therapy with PARPi plus ARPi, and offers recommendations for management of these side effects.
Topics: Male; Humans; Prostatic Neoplasms, Castration-Resistant; Receptors, Androgen; Ribose; Poly(ADP-ribose) Polymerases; Signal Transduction; Adenosine Diphosphate
PubMed: 37978072
DOI: 10.1007/s00120-023-02230-1 -
Methods in Molecular Biology (Clifton,... 2022Mitochondrial energy production is crucial for normal daily activities and maintenance of life. Herein, the logic and execution of two main classes of measurements are...
Mitochondrial energy production is crucial for normal daily activities and maintenance of life. Herein, the logic and execution of two main classes of measurements are outlined to delineate mitochondrial function: ATP production and oxygen consumption. Aerobic ATP production is quantified by phosphorus magnetic resonance spectroscopy (PMRS) in vivo in both human subjects and animal models using the same protocols and maintaining the same primary assumptions. Mitochondrial oxygen consumption is quantified by oxygen polarography and applied in isolated mitochondria, cultured cells, and permeabilized fibers derived from human or animal tissue biopsies. Traditionally, mitochondrial functional measures focus on maximal oxidative capacity-a flux rate that is rarely, if ever, observed outside of experimental conditions. Perhaps more physiologically relevant, both measurement classes herein focus on one principal design paradigm; submaximal mitochondrial fluxes generated by graded levels of ADP to map the function for ADP sensitivity. We propose this function defines the bioenergetic role that mitochondria fill within the myoplasm to sense and match ATP demands. Any deficit in this vital role for ATP homeostasis leads to symptoms often seen in cardiovascular and cardiopulmonary diseases, diabetes, and metabolic syndrome.
Topics: Adenosine Diphosphate; Adenosine Triphosphate; Animals; Energy Metabolism; Humans; Mitochondria; Oxidative Phosphorylation; Oxygen Consumption; Polarography
PubMed: 35771441
DOI: 10.1007/978-1-0716-2309-1_10 -
Cancer Jun 2023
Topics: Humans; Poly(ADP-ribose) Polymerases; Adenosine Diphosphate Ribose
PubMed: 37060235
DOI: 10.1002/cncr.34705 -
Proceedings of the National Academy of... Oct 2022The major cytoskeleton protein actin undergoes cyclic transitions between the monomeric G-form and the filamentous F-form, which drive organelle transport and cell...
The major cytoskeleton protein actin undergoes cyclic transitions between the monomeric G-form and the filamentous F-form, which drive organelle transport and cell motility. This mechanical work is driven by the ATPase activity at the catalytic site in the F-form. For deeper understanding of the actin cellular functions, the reaction mechanism must be elucidated. Here, we show that a single actin molecule is trapped in the F-form by fragmin domain-1 binding and present their crystal structures in the ATP analog-, ADP-Pi-, and ADP-bound forms, at 1.15-Å resolutions. The G-to-F conformational transition shifts the side chains of Gln137 and His161, which relocate four water molecules including W1 (attacking water) and W2 (helping water) to facilitate the hydrolysis. By applying quantum mechanics/molecular mechanics calculations to the structures, we have revealed a consistent and comprehensive reaction path of ATP hydrolysis by the F-form actin. The reaction path consists of four steps: 1) W1 and W2 rotations; 2) P-O bond cleavage; 3) four concomitant events: W1-PO formation, OH and proton cleavage, nucleophilic attack by the OH against P, and the abstracted proton transfer; and 4) proton relocation that stabilizes the ADP-Pi-bound F-form actin. The mechanism explains the slow rate of ATP hydrolysis by actin and the irreversibility of the hydrolysis reaction. While the catalytic strategy of actin ATP hydrolysis is essentially the same as those of motor proteins like myosin, the process after the hydrolysis is distinct and discussed in terms of Pi release, F-form destabilization, and global conformational changes.
Topics: Actins; Adenosine Diphosphate; Adenosine Triphosphate; Dalteparin; Hydrolysis; Myosins; Protons; Water
PubMed: 36252034
DOI: 10.1073/pnas.2122641119 -
Spectrochimica Acta. Part A, Molecular... Jan 2024Nucleoside polyphosphate (NPP) anions are important for enzymatic activity and should be monitored by scientists in industry and medicine. By elucidating enzyme kinetics...
BACKGROUND
Nucleoside polyphosphate (NPP) anions are important for enzymatic activity and should be monitored by scientists in industry and medicine. By elucidating enzyme kinetics and processes, it aids in the discovery of effective inhibitors and activators. Nucleoside polyphosphate (NPP) anions are used by kinases, GTPases, and glycosyltransferases (GTs). Phosphorylation of certain amino acid residues (Ser, Thr, and Tyr) on proteins requires the breakdown of ATP by protein kinases, which produces ADP. Protein kinases, breakdown of ATP, and NPP are the focus of oncology drug development because the aberrant control of kinase activity is a common cause of cancer.
RESULTS
However, a discriminative turn-on fluorescent property is exhibited by non-fluorescent p-tertbutylcalix[4]arene modified 1,2,3-triazole containing bis-ruthenium polypyridyl complex (RL) upon the addition of phosphate anions such as (dihydrogen pyrophosphate (HPO) and dihydrogen phosphate (HPO)) in CHCN solvent and Adenosine Diphosphate (ADP) in CHCN/HEPES (pH = 7.4) buffer (9/1, v/v). The probe RL shows a better-recognizing ability with pyrophosphate anion (HPO) than dihydrogen phosphate anion (HPO). With HPO and HPO anions, the RL detection limit was calculated to be as low as 83 nM and 198 nM, respectively.
SIGNIFICANCE
The calix[4]arene macrocycle's excellent size and binding cone conformation make it a good host-guest interface for the pyrophosphate anion and ADP. The bis-ruthenium polypyridyl complex's connection to the p-tertbutyl calix[4]arene moiety creates the ADP selectivity turn-on sensor. When moving from mono-nuclear to bi-nuclear ruthenium complex anchored on p-tertbutyl calix[4]arene, the probe can differentiate ADP, ATP, and AMP. Furthermore, this platform is a great resource for creating devices to simultaneously assess phosphate anions in environmental samples.
Topics: Phosphates; Diphosphates; Ruthenium; Nucleosides; Anions; Adenosine Diphosphate; Adenosine Triphosphate; Protein Kinases
PubMed: 37804708
DOI: 10.1016/j.saa.2023.123483 -
GeroScience Dec 2023Aging muscle experiences functional decline in part mediated by impaired mitochondrial ADP sensitivity. Elamipretide (ELAM) rapidly improves physiological and...
The mitochondrially targeted peptide elamipretide (SS-31) improves ADP sensitivity in aged mitochondria by increasing uptake through the adenine nucleotide translocator (ANT).
Aging muscle experiences functional decline in part mediated by impaired mitochondrial ADP sensitivity. Elamipretide (ELAM) rapidly improves physiological and mitochondrial function in aging and binds directly to the mitochondrial ADP transporter ANT. We hypothesized that ELAM improves ADP sensitivity in aging leading to rescued physiological function. We measured the response to ADP stimulation in young and old muscle mitochondria with ELAM treatment, in vivo heart and muscle function, and compared protein abundance, phosphorylation, and S-glutathionylation of ADP/ATP pathway proteins. ELAM treatment increased ADP sensitivity in old muscle mitochondria by increasing uptake of ADP through the ANT and rescued muscle force and heart systolic function. Protein abundance in the ADP/ATP transport and synthesis pathway was unchanged, but ELAM treatment decreased protein s-glutathionylation incuding of ANT. Mitochondrial ADP sensitivity is rapidly modifiable. This research supports the hypothesis that ELAM improves ANT function in aging and links mitochondrial ADP sensitivity to physiological function. ELAM binds directly to ANT and ATP synthase and ELAM treatment improves ADP sensitivity, increases ATP production, and improves physiological function in old muscles. ADP (adenosine diphosphate), ATP (adenosine triphosphate), VDAC (voltage-dependent anion channel), ANT (adenine nucleotide translocator), H (proton), ROS (reactive oxygen species), NADH (nicotinamide adenine dinucleotide), FADH (flavin adenine dinucleotide), O (oxygen), ELAM (elamipretide), -SH (free thiol), -SSG (glutathionylated protein).
Topics: Mitochondria; Adenosine Triphosphate; Adenosine Diphosphate; Peptides
PubMed: 37462785
DOI: 10.1007/s11357-023-00861-y