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Nature Dec 2021The human microbiome encodes a large repertoire of biochemical enzymes and pathways, most of which remain uncharacterized. Here, using a metagenomics-based search...
The human microbiome encodes a large repertoire of biochemical enzymes and pathways, most of which remain uncharacterized. Here, using a metagenomics-based search strategy, we discovered that bacterial members of the human gut and oral microbiome encode enzymes that selectively phosphorylate a clinically used antidiabetic drug, acarbose, resulting in its inactivation. Acarbose is an inhibitor of both human and bacterial α-glucosidases, limiting the ability of the target organism to metabolize complex carbohydrates. Using biochemical assays, X-ray crystallography and metagenomic analyses, we show that microbiome-derived acarbose kinases are specific for acarbose, provide their harbouring organism with a protective advantage against the activity of acarbose, and are widespread in the microbiomes of western and non-western human populations. These results provide an example of widespread microbiome resistance to a non-antibiotic drug, and suggest that acarbose resistance has disseminated in the human microbiome as a defensive strategy against a potential endogenous producer of a closely related molecule.
Topics: Acarbose; Amylases; Animals; Drug Resistance, Bacterial; Gastrointestinal Microbiome; Humans; Hypoglycemic Agents; Inactivation, Metabolic; Metagenome; Models, Molecular; Mouth; Phosphotransferases (Alcohol Group Acceptor)
PubMed: 34819672
DOI: 10.1038/s41586-021-04091-0 -
Journal of the American Chemical Society Nov 2018Target residence time is emerging as an important optimization parameter in drug discovery, yet target and off-target engagement dynamics have not been clearly linked to...
Target residence time is emerging as an important optimization parameter in drug discovery, yet target and off-target engagement dynamics have not been clearly linked to the clinical performance of drugs. Here we developed high-throughput binding kinetics assays to characterize the interactions of 270 protein kinase inhibitors with 40 clinically relevant targets. Analysis of the results revealed that on-rates are better correlated with affinity than off-rates and that the fraction of slowly dissociating drug-target complexes increases from early/preclinical to late stage and FDA-approved compounds, suggesting distinct contributions by each parameter to clinical success. Combining binding parameters with PK/ADME properties, we illustrate in silico and in cells how kinetic selectivity could be exploited as an optimization strategy. Furthermore, using bio- and chemoinformatics we uncovered structural features influencing rate constants. Our results underscore the value of binding kinetics information in rational drug design and provide a resource for future studies on this subject.
Topics: Binding Sites; Drug Discovery; Humans; Kinetics; Molecular Structure; Phosphotransferases; Protein Kinase Inhibitors
PubMed: 30362749
DOI: 10.1021/jacs.8b08048 -
The Journal of Physical Chemistry. B Jul 2022The environmental condition is a critical regulation factor for protein behavior in solution. Several studies have shown that macromolecular crowders can modulate...
The environmental condition is a critical regulation factor for protein behavior in solution. Several studies have shown that macromolecular crowders can modulate protein structures, interactions, and functions. Recent publications described the regulation of specific interaction by macromolecular crowders. However, the other category of protein-protein interaction, namely, the transient interaction, is rarely investigated, especially from the perspective of protein structure to study transient interactions between proteins. Here, we used nuclear magnetic resonance and small-angle X-ray/neutron scattering methods to structurally investigate the ensemble of the protein complex in dilute buffer and crowded environments. Histidine phosphocarrier protein (HPr) and the N-terminal domain of enzyme I (EIN) are the important components of the bacterial phosphotransfer system. Our results show that the addition of Ficoll-70 promotes HPr molecules to form the encounter complex with EIN maintained by long-range electrostatic interaction. However, when macromolecular crowder BSA is used, the soft interaction between BSA and HPr perturbs the active site of HPr, driving HPr to form an encounter complex with EIN at the weakly charged interface. Our results indicate that different macromolecular crowders could influence transient EIN-HPr interaction through different mechanisms and provide new insights into protein-protein interaction regulation in native environments.
Topics: Bacterial Proteins; Catalytic Domain; Histidine; Macromolecular Substances; Phosphoenolpyruvate Sugar Phosphotransferase System
PubMed: 35731981
DOI: 10.1021/acs.jpcb.2c02713 -
Frontiers in Bioscience (Landmark... Mar 2020Chemokines are small regulatory proteins that play a crucial role in the coordinated migration of cell populations to the site of infection/inflammation by binding to... (Review)
Review
Chemokines are small regulatory proteins that play a crucial role in the coordinated migration of cell populations to the site of infection/inflammation by binding to their cognate receptors. In principle, chemokine receptors, which are serpentine G protein-coupled receptors (GPCRs), mediate the series of downstream intracellular signaling events that occur inside the cells to resolve the pathogenicity. Intracellular signaling pathways regulated by the kinase protein sub-families are the center of attention for chemokine derived functional responses. Kinases potentially influence cell migration, cell growth, transcriptional activation, and other essential molecular events. The regulation and flow of the signals by the kinases are different for each physiological and pathological event and are tightly regulated by the nature and pairing of chemokine(s) with its receptor(s). For example, phosphoinositide 3-kinase (PI3K) is activated during the initial steps of the chemokine induced signaling cascade to regulate chemotaxis, transcription, and cell survival. G protein-coupled receptor kinase (GRKs) plays a crucial role in the desensitization and internalization of the chemokine receptors. The regulation of chemokine receptor is also governed by kinases like protein kinase A (PKA), protein kinase C (PKC), mitogen-activated protein kinases / extracellular signal-regulated kinases (MAPK/ERK), . It was also established that tyrosine-protein kinases (TECs) such as ITK and RLK play a significant role in chemokine signaling in T lymphocytes. On a similar note, many others like janus kinases (JAKs), Protein kinase B (PKB), PKC, etc. are also studied in chemokine mediated disease models. The present review elucidates the role of different kinases involved in the chemokine/chemokine receptor mediated signaling cascade during various pathophysiological processes.
Topics: Chemokines; Humans; Janus Kinases; Mitogen-Activated Protein Kinases; Phosphatidylinositol 3-Kinase; Phosphotransferases; Protein-Tyrosine Kinases; Receptors, Chemokine; Receptors, G-Protein-Coupled; Signal Transduction
PubMed: 32114437
DOI: 10.2741/4860 -
Advances in Biological Regulation Jan 2019The protein kinase family is characterized by substantial conservation of architectural elements that are required for both ATP binding and phosphotransferase activity.... (Review)
Review
The protein kinase family is characterized by substantial conservation of architectural elements that are required for both ATP binding and phosphotransferase activity. Many of these structural features have also been identified in homologous enzymes that phosphorylate a variety of alternative, non-protein substrates. A comparative structural analysis of these different kinase sub-classes is a portal to a greater understanding of reaction mechanisms, enzyme regulation, inhibitor-development strategies, and superfamily-level evolutionary relationships. To serve such advances, we review structural elements of the protein kinase fold that are conserved in the subfamily of inositol phosphate kinases (InsPKs) that share a PxxxDxKxG catalytic signature: inositol 1,4,5-trisphosphate kinase (IP3K), inositol hexakisphosphate kinase (IP6K), and inositol polyphosphate multikinase (IPMK). We describe conservation of the fundamental two-lobe kinase architecture: an N-lobe constructed upon an anti-parallel β-strand scaffold, which is coupled to a largely helical C-lobe by a single, adenine-binding hinge. This equivalency also includes a G-loop that embraces the β/γ-phosphates of ATP, a transition-state stabilizing residue (Lys/His), and a Mg-positioning aspartate residue within a catalytic triad. Furthermore, we expand this list of conserved structural features to include some not previously identified in InsPKs: a 'gatekeeper' residue in the N-lobe, and an 'αF'-like helix in the C-lobe that anchors two structurally-stabilizing, hydrophobic spines, formed from non-consecutive residues that span the two lobes. We describe how this wide-ranging structural homology can be exploited to develop lead inhibitors of IP6K and IPMK, by using strategies similar to those that have generated ATP-competing inhibitors of protein-kinases. We provide several examples to illustrate how such an approach could benefit human health.
Topics: Animals; Binding Sites; Humans; Inositol Phosphates; Phosphorylation; Phosphotransferases (Alcohol Group Acceptor); Phosphotransferases (Phosphate Group Acceptor); Protein Kinases; Protein Structure, Secondary
PubMed: 30392847
DOI: 10.1016/j.jbior.2018.10.006 -
Current Medicinal Chemistry 2015One-third of epileptic patients are drug refractory due to the limited efficacy of antiepileptic therapy. Thus, there is an immense need to find more effective, safer... (Review)
Review
One-third of epileptic patients are drug refractory due to the limited efficacy of antiepileptic therapy. Thus, there is an immense need to find more effective, safer and well-tolerated antiepileptic drugs. A great deal of results suggests that adenosine (Ado), guanosine (Guo), inosine (Ino) or uridine (Urd) are endogenous antiepileptogenic modulators. Furthermore, nucleosides and their derivatives may be safe and effective potential drugs in the treatment of epilepsy. Conversely, nucleosidergic modulatory system implying nucleoside levels, metabolism, receptors and transporters may also be involved in seizure pathomechanisms. Application of Ado receptor agonists as well as antagonists, elevation of nucleoside levels (e.g., by nucleoside metabolism inhibitors, and Adoreleasing implants) or utilization of non-Ado nucleosides may also turn to be useful approaches to decrease epileptic activity. However, all drugs exerting their effects on the nucleosidergic modulatory system may affect the fine regulation of glia-neuron interactions that are intimately governed by various nucleosidergic processes. Perturbation of the complex, bidirectional communication between neurons and astrocytes through these nucleosidergic modulatory mechanisms may lead to pathological changes in the central nervous system (CNS) and therefore may cause significant side effects. Thus, a deeper understanding of the nucleosidergic modulatory control over glia-neuron interactions is essential in order to develop more effective and safe nucleoside-based antiepileptic drugs. In this review article we focus on the role of Ado and Urd in glia-neuron interactions, placing emphasis on their implications for the treatment of epilepsy.
Topics: Animals; Anticonvulsants; Enzyme Inhibitors; Humans; Neuroglia; Neurons; Nucleosides; Phosphotransferases
PubMed: 25666791
DOI: 10.2174/0929867322666150212153210 -
Biomedicine & Pharmacotherapy =... Jan 2024Polycystic ovary syndrome (PCOS) is a highly prevalent endocrine and metabolic disorder that is closely associated with the proliferation and apoptosis of ovarian...
Polycystic ovary syndrome (PCOS) is a highly prevalent endocrine and metabolic disorder that is closely associated with the proliferation and apoptosis of ovarian granulosa cells (GCs). Ampelopsis japonica (AJ) is the dried tuberous root of Ampelopsis japonica (Thunb.) Makino (A. japonica), with anti-inflammatory, antioxidant, antibacterial, antiviral, wound-healing, and antitumor properties; however, it is unclear whether this herb has a therapeutic effect on PCOS. Therefore, this study aimed to investigate the pharmacological effect of AJ on PCOS and reveal its potential mechanism of action. A PCOS rat model was established using letrozole. After establishing the PCOS model, the rats received oral treatment of AJ and Diane-35 (Positive drug: ethinylestradiol + cyproterone tablets) for 2 weeks. Lipidomics was conducted using liquid-phase mass spectrometry and chromatography. AJ significantly regulated serum hormone levels and attenuated pathological variants in the ovaries of rats with PCOS. Furthermore, AJ significantly reduced the apoptotic rate of ovarian GCs. Lipidomic analysis revealed that AJ modulated glycerolipid and glycerophospholipid metabolic pathways mediated by lipoprotein lipase (Lpl), diacylglycerol choline phosphotransferase (Chpt1), and choline/ethanolamine phosphotransferase (Cept1). Therefore, we established that AJ may reduce ovarian GC apoptosis by modulating lipid metabolism, ultimately improving ovulatory dysfunction in PCOS. Therefore, AJ is a novel candidate for PCOS treatment.
Topics: Female; Humans; Rats; Animals; Polycystic Ovary Syndrome; Ampelopsis; Lipid Metabolism; Phosphotransferases; Choline
PubMed: 38159378
DOI: 10.1016/j.biopha.2023.116093 -
Current Topics in Medicinal Chemistry 2017
Topics: Drug Discovery; Humans; Phosphorylation; Phosphotransferases; Protein Kinase Inhibitors
PubMed: 28799507
DOI: 10.2174/156802661722170726105918 -
Journal of Cellular Biochemistry Jun 2017Purinergic signaling maintains local tissue homeostasis in blood vessels via the regulation of vascular tone, blood platelet aggregation, cell proliferation, and...
Gene Expression and Activity Profiling Reveal a Significant Contribution of Exo-Phosphotransferases to the Extracellular Nucleotides Metabolism in HUVEC Endothelial Cells.
Purinergic signaling maintains local tissue homeostasis in blood vessels via the regulation of vascular tone, blood platelet aggregation, cell proliferation, and differentiation as well as inflammatory responses. Extracellular purines are important signaling molecules in the vasculature, and both purine-hydrolysing as well as -phosphorylating enzymes are considered to selectively govern extracellular nucleotide/nucleoside metabolism. Recent studies have provided some evidence for the existence of these enzymes in a soluble form in human blood and their secretion into the extracellular space under physiological and pathological conditions. However, the comprehensive analysis of endothelium-derived enzymes involved in purine metabolic pathways has received no attention so far. In the presented study, in vitro cultured human umbilical vein endothelial cells (HUVEC) are shown to be an abundant source of exo-nucleotidases comprising 5'-nucleotidase (exo-5'-NT), and nucleoside triphosphate diphosphohydrolases (exo-NTPDase) as well as phosphotransferases, represented by nucleoside diphosphate kinase (exo-NDPK) and adenylate kinase (exo-AK). An attempt is also made to demonstrate that, in contrast to the metabolic pattern determined on the endothelial cell surface, exo-phosphorylating activities markedly predominate over exo-hydrolytic ones. We present for the first time the expression profiles of genes encoding isoenzymes belonging to distinct nucleotide kinase and nucleotidase families. The genes encoding NDPK1, NDPK2, AK1, and AK2 phosphotransferases have been shown to be expressed at the highest level in HUVEC cells. The data indicate the coexistence of secreted and cell-associated factors of endothelial origin mediating ATP-consuming and ATP-generating pathways with the predominance of exo-phosphotransferases activity. The described enzymes contribute to the regulation of purinergic signal duration and extent in the venous vasculature. J. Cell. Biochem. 118: 1341-1348, 2017. © 2016 Wiley Periodicals, Inc.
Topics: Adenosine Triphosphate; Adenylate Kinase; Gene Expression Profiling; Human Umbilical Vein Endothelial Cells; Humans; Nucleoside-Diphosphate Kinase; Phosphotransferases (Phosphate Group Acceptor); Purines; Signal Transduction
PubMed: 27859553
DOI: 10.1002/jcb.25791 -
Acta Crystallographica. Section D,... Dec 2019Aminoglycoside phosphotransferases (APHs) are one of three families of aminoglycoside-modifying enzymes that confer high-level resistance to the aminoglycoside...
Aminoglycoside phosphotransferases (APHs) are one of three families of aminoglycoside-modifying enzymes that confer high-level resistance to the aminoglycoside antibiotics via enzymatic modification. This has now rendered many clinically important drugs almost obsolete. The APHs specifically phosphorylate hydroxyl groups on the aminoglycosides using a nucleotide triphosphate as the phosphate donor. The APH(2'') family comprises four distinct members, isolated primarily from Enterococcus sp., which vary in their substrate specificities and also in their preference for the phosphate donor (ATP or GTP). The structure of the ternary complex of APH(2'')-IIIa with GDP and kanamycin was solved at 1.34 Å resolution and was compared with substrate-bound structures of APH(2'')-Ia, APH(2'')-IIa and APH(2'')-IVa. In contrast to the case for APH(2'')-Ia, where it was proposed that the enzyme-mediated hydrolysis of GTP is regulated by conformational changes in its N-terminal domain upon GTP binding, APH(2'')-IIa, APH(2'')-IIIa and APH(2'')-IVa show no such regulatory mechanism, primarily owing to structural differences in the N-terminal domains of these enzymes.
Topics: Bacterial Proteins; Binding Sites; Crystallography, X-Ray; Enterococcus; Guanosine Triphosphate; Kanamycin; Models, Molecular; Phosphotransferases (Alcohol Group Acceptor); Protein Conformation; Substrate Specificity
PubMed: 31793906
DOI: 10.1107/S2059798319015079