Articoli di riviste sul tema "Inhibition covalente"
Segui questo link per vedere altri tipi di pubblicazioni sul tema: Inhibition covalente.
Cita una fonte nei formati APA, MLA, Chicago, Harvard e in molti altri stili
Vedi i top-50 articoli di riviste per l'attività di ricerca sul tema "Inhibition covalente".
Accanto a ogni fonte nell'elenco di riferimenti c'è un pulsante "Aggiungi alla bibliografia". Premilo e genereremo automaticamente la citazione bibliografica dell'opera scelta nello stile citazionale di cui hai bisogno: APA, MLA, Harvard, Chicago, Vancouver ecc.
Puoi anche scaricare il testo completo della pubblicazione scientifica nel formato .pdf e leggere online l'abstract (il sommario) dell'opera se è presente nei metadati.
Vedi gli articoli di riviste di molte aree scientifiche e compila una bibliografia corretta.
1
Aljoundi, Aimen, Ahmed El Rashedy, Patrick Appiah-Kubi e Mahmoud E. S. Soliman. "Coupling of HSP72 α-Helix Subdomains by the Unexpected Irreversible Targeting of Lysine-56 over Cysteine-17; Coevolution of Covalent Bonding". Molecules 25, n. 18 (16 settembre 2020): 4239. http://dx.doi.org/10.3390/molecules25184239.
Abstract (sommario):
Covalent inhibition has recently gained a resurgence of interest in several drug discovery areas. The expansion of this approach is based on evidence elucidating the selectivity and potency of covalent inhibitors when bound to particular amino acids of a biological target. The unexpected covalent inhibition of heat shock protein 72 (HSP72) by covalently targeting Lys-56 instead of Cys-17 was an interesting observation. However, the structural basis and conformational changes associated with this preferential coupling to Lys-56 over Cys-17 remain unclear. To resolve this mystery, we employed structural and dynamic analyses to investigate the structural basis and conformational dynamics associated with the unexpected covalent inhibition. Our analyses reveal that the coupling of the irreversible inhibitor to Lys-56 is intrinsically less dynamic than Cys-17. Conformational dynamics analyses further reveal that the coupling of the inhibitor to Lys-56 induced a closed conformation of the nucleotide-binding subdomain (NBD) α-helices, in contrast, an open conformation was observed in the case of Cys-17. The closed conformation maintained the crucial salt-bridge between Glu-268 and Lys-56 residues, which strengthens the interaction affinity of the inhibitor nearly identical to adenosine triphosphate (ADP/Pi) bound to the HSP72-NBD. The outcome of this report provides a substantial shift in the conventional direction for the design of more potent covalent inhibitors.
2
Liu, S. Q., e P. A. Knauf. "Lys-430, site of irreversible inhibition of band 3 Cl- flux by eosin-5-maleimide, is not at the transport site". American Journal of Physiology-Cell Physiology 264, n. 5 (1 maggio 1993): C1155—C1164. http://dx.doi.org/10.1152/ajpcell.1993.264.5.c1155.
Abstract (sommario):
Although eosin-5-maleimide (EM) covalently labels band 3 and has been thought to react at the external-facing anion transport site, EM reversibly inhibits Cl- exchange at 0 degrees C in a noncompetitive fashion, indicating that under these conditions it does not bind to the transport site [Knauf, P.A., N.M. Strong, J. Penikas, R.B. Wheeler, Jr., and S.J. Liu. Am. J. Physiol. 264 (Cell Physiol. 33): C1144-C1154 1993]. To see whether or not the covalent labeling by EM takes place at the same noncompetitive site as the reversible binding, we examined the dependence of reaction rate on EM concentration. The reaction rate saturates with increasing EM concentration, indicating that reversible binding precedes covalent reaction and that EM therefore acts as an affinity label. A more complex model in which reversible binding prevents a bimolecular reaction at a different site cannot, however, be ruled out. Cl- gradients across the membrane affect EM reversible binding in a manner suggesting that EM binds preferentially to the Eo form of band 3, with the transport site unloaded and facing outward. Thus EM binds to and probably reacts covalently with a site that is different from the transport site, but whose conformation is affected by the orientation of the transport site. Lysine-430, the amino acid residue which is covalently labeled by EM (4), may be near the transport site but does not seem to be directly involved in the binding of transported substrates such as chloride. EM binding to one band 3 monomer decreases the reactivity of the adjacent monomer but does not decrease the affinity constant of the reversible binding step that precedes covalent reaction. Although a small fraction (approximately 1%) of band 3 monomers fail to react with EM, EM nearly completely inhibits transport in those monomers with which it reacts.
3
Yang, Jianhong, Yong Li, Wei Yan, Weimin Li, Qiang Qiu, Haoyu Ye e Lijuan Chen. "Covalent modification of Cys-239 in β-tubulin by small molecules as a strategy to promote tubulin heterodimer degradation". Journal of Biological Chemistry 294, n. 20 (2 aprile 2019): 8161–70. http://dx.doi.org/10.1074/jbc.ra118.006325.
Abstract (sommario):
Clinical microtubule-targeting drugs are functionally divided into microtubule-destabilizing and microtubule-stabilizing agents. Drugs from both classes achieve microtubule inhibition by binding different sites on tubulin and inhibiting or promoting polymerization with no concomitant effects on the protein levels of tubulin heterodimers. Here, we have identified a series of small molecules with diverse structures potentially representing a third class of novel tubulin inhibitors that promote degradation by covalent binding to Cys-239 of β-tubulin. The small molecules highlighted in this study include T0070907 (a peroxisome proliferator-activated receptor γ inhibitor), T007-1 (a T0070907 derivative), T138067, N,N′-ethylene-bis(iodoacetamide) (EBI), and allyl isothiocyanate (AITC). Label-free quantitative proteomic analysis revealed that T007-1 promotes tubulin degradation with high selectivity. Mass spectrometry findings showed covalent binding of both T0070907 and T007-01 to Cys-239 of β-tubulin. Furthermore, T007-1 exerted a degradative effect on tubulin isoforms possessing Cys-239 (β2, β4, and β5(β)) but not those containing Ser-239 (β3, β6) or mutant β-tubulin with a C239S substitution. Three small molecules (T138067, EBI, and AITC) also reported to bind covalently to Cys-239 of β-tubulin similarly induced tubulin degradation. Our results strongly suggest that covalent modification of Cys-239 of β-tubulin by small molecules could serve as a novel strategy to promote tubulin heterodimer degradation. We propose that these small molecules represent a third novel class of tubulin inhibitor agents that exert their effects through degradation activity.
4
Maksimenko, A. V., e R. Sh Beabealashvili. "Theoretical Grounding and Formation of Experimental Approaches to Hyaluronidase Structure Consolidation due to Its Computational Interactions with Shortchain Glycosaminoglycan Ligands". Биоорганическая химия 49, n. 4 (1 luglio 2023): 369–83. http://dx.doi.org/10.31857/s0132342323020161.
Abstract (sommario):
The computational study of 3D model hyaluronidase interaction with shortchain glycosaminoglycan ligands demonstrated the diversity and significance of their reaction on enzyme structure. It has been realized due to electrostatic noncovalent interactions (without specific coupling with active site) inducing the perceptible conformational alterations of biocatalyst molecule. As a result of this the inactivation and stabilization of enzyme globule are observed, change of inhibition of biocatalyst by heparin. The binding of chondroitin trimers (on centers cn6, cn3, cn1) to hyaluronidase molecular surface increased the enzyme stability, binding of chondroitin sulfate trimers (on centers cs2, cs4, cs7, cs8 or cs1, cs2, cs4, cs7, cs8) decreased the inhibition of enzyme by tetramer heparin. It should be noted the importance of ligand binding for regulation of enzyme functioning and existence of multiform and multicomponent microenvironment of enzyme. The sequence of preferable coupling of ligands with hyaluronidase is elicited in our study and with its help was evaluate reality of experimental selective modification of enzyme (possibly no covalently or covalently, for instance, with chondroitin sulfate trimers on centers cs7, cs1, cs5) for experimental obtaining of stabilized enzyme forms of medical destination. The perspective approaches for this aim may be the no covalent reaction on hyaluronidase by chondroitin or chondroitin sulfate trimers as well covalent modification of biocatalyst by chondroitin sulfate trimers.
5
Bhatia, Sumeena, Steven C. Almo, Stanley G. Nathenson e Richard J. Hodes. "Dynamic equilibrium of B7-1 dimers and monomers is important for regulation of TCR/CD28 – mediated T cell activation (33.28)". Journal of Immunology 182, n. 1_Supplement (1 aprile 2009): 33.28. http://dx.doi.org/10.4049/jimmunol.182.supp.33.28.
Abstract (sommario):
Abstract Under steady state conditions, B7-1 is present as a mixed population of non-covalent dimers and monomers on the cell surface. Here we have examined the physiological significance of this unique dimer-monomer equilibrium state of B7-1. We demonstrate that altering B7-1 to uniformly covalent dimeric state results in increased frequency of CD28 mediated T cell-APC conjugates. This augmented T cell-APC conjugate formation correlates with persistent concentration of signaling molecules, PKC-θ and lck, at the immunological synapse (IS) and with a higher calcium flux in T cells. In contrast, B7-1 acquisition by T cells, an event that occurs as a consequence of CD28 engagement with B7-1/B7-2, is highly reduced when B7-1 is present in covalently dimeric state. The ability of covalently dimeric and wild type B7-1 to costimulate antigen-specific T cell proliferation was also assessed. In contrast to the enhanced ability of dimeric B7-1 to support proximal T cell activation events, sensitivity to competitive inhibition by soluble CTLA-4:Ig indicated that the covalent dimeric form of B7-1 is in fact less efficient in costimulating T cell proliferation. These findings suggest a novel model in which optimal T cell costimulatory function of B7-1 requires CD28 engagement by B7-1 in a non-covalent dimeric state, followed by dissociation of these B7-1 dimers, facilitating downregulation of CD28 and internalization of B7-1.
6
Kuznetsova, Anastasiya, Philipp Klein e Till Opatz. "Halogenated 2,1,3-benzoxadiazoles as Potential Fluorescent Warheads for Covalent Protease Inhibitors". Proceedings 9, n. 1 (14 novembre 2018): 54. http://dx.doi.org/10.3390/ecsoc-22-05670.
Abstract (sommario):
Recently there has been a growing interest in covalent protease inhibitors in both industry and academia, caused by their longer residence times, their higher potency and their high ligand efficiency. Covalently reactive moieties which interact with activated amino acid residues such as serine or cysteine in enzymes like proteases or esterases mostly act through nucleophilic addition, substitution or ring opening. In contrast, nucleophilic aromatic substitution (SNAr) is rarely employed. In our previous work, we prepared and investigated electrophilic “warheads”, which contain aromatic, heteroaromatic or quinoid fragments. Some of them show potent inhibition constants for cathepsin L, cathepsin B, rhodesain or dengue-protease, and depending on the exact nature of the electrophile they exhibit reversible covalent or irreversible inhibition modes. In the present work, we demonstrate the synthesis of fluorescent “warhead” candidates based on 2,1,3-benzoxadiazoles and the investigation of their physicochemical and photophysical properties. These molecules shall serve as probes for the detailed analysis of association/dissociation mechanism and of the kinetic parameters of the bond forming event.
7
Hognon, Cécilia, Marco Marazzi e Cristina García-Iriepa. "Atomistic-Level Description of the Covalent Inhibition of SARS-CoV-2 Papain-like Protease". International Journal of Molecular Sciences 23, n. 10 (23 maggio 2022): 5855. http://dx.doi.org/10.3390/ijms23105855.
Abstract (sommario):
Inhibition of the papain-like protease (PLpro) of SARS-CoV-2 has been demonstrated to be a successful target to prevent the spreading of the coronavirus in the infected body. In this regard, covalent inhibitors, such as the recently proposed VIR251 ligand, can irreversibly inactivate PLpro by forming a covalent bond with a specific residue of the catalytic site (Cys111), through a Michael addition reaction. An inhibition mechanism can therefore be proposed, including four steps: (i) ligand entry into the protease pocket; (ii) Cys111 deprotonation of the thiol group by a Brønsted–Lowry base; (iii) Cys111-S− addition to the ligand; and (iv) proton transfer from the protonated base to the covalently bound ligand. Evaluating the energetics and PLpro conformational changes at each of these steps could aid the design of more efficient and selective covalent inhibitors. For this aim, we have studied by means of MD simulations and QM/MM calculations the whole mechanism. Regarding the first step, we show that the inhibitor entry in the PLpro pocket is thermodynamically favorable only when considering the neutral Cys111, that is, prior to the Cys111 deprotonation. For the second step, MD simulations revealed that His272 would deprotonate Cys111 after overcoming an energy barrier of ca. 32 kcal/mol (at the QM/MM level), but implying a decrease of the inhibitor stability inside the protease pocket. This information points to a reversible Cys111 deprotonation, whose equilibrium is largely shifted toward the neutral Cys111 form. Although thermodynamically disfavored, if Cys111 is deprotonated in close proximity to the vinylic carbon of the ligand, then covalent binding takes place in an irreversible way (third step) to form the enolate intermediate. Finally, due to Cys111-S− negative charge redistribution over the bound ligand, proton transfer from the initially protonated His272 is favored, finally leading to an irreversibly modified Cys111 and a restored His272. These results elucidate the selectivity of Cys111 to enable formation of a covalent bond, even if a weak proton acceptor is available, as His272.
8
Bisconte, Angelina, Ronald Hill, Michael Bradshaw, Erik Verner, David Finkle, Ken Brameld, Jens Funk, David Goldstein e Phil Nunn. "Efficacy in collagen induced arthritis models with a selective, reversible covalent Bruton’s tyrosine kinase inhibitor PRN473 is driven by durable target occupancy rather than extended plasma exposure (THER5P.904)". Journal of Immunology 194, n. 1_Supplement (1 maggio 2015): 139.6. http://dx.doi.org/10.4049/jimmunol.194.supp.139.6.
Abstract (sommario):
Abstract Bruton’s Tyrosine Kinase (BTK) is an essential signaling element downstream of the B-cell receptor (BCR). Inhibition of BTK activity in B cells produces phenotypic changes consistent with blockade of the BCR, including inhibition of cell proliferation, differentiation, maturation, and survival. A selective BTK inhibitor has the potential to treat diseases involving inflammation and autoimmunity. Using Principia Biopharma’s proprietary Tailored Covalency™ technology, we discovered PRN473, a reversible covalent BTK inhibitor that selectively binds BTK with a slow off-rate as assessed in biochemical and cell based assays. A slow off-rate molecule with rapid systemic clearance may lead to a long duration of action and high efficacy, while reducing the potential for off target toxicities. The duration of binding of PRN473 was measured in vivo, with prolonged BTK occupancy confirmed after drug was cleared from plasma. In mouse and rat collagen-induced arthritis (CIA) models, PRN473 when dosed therapeutically demonstrated dose-dependent inhibition, reversal of arthritis, with almost complete abrogation of clinical scores and preservation of joint integrity and histology. BTK occupancy in splenocytes was found to be a robust measure of BTK inhibition and correlated with efficacy in the CIA models. These data in combination with GLP toxicology studies, where no organ specific toxicities were observed, validate using a reversible covalent BTK inhibitor to treat autoimmune diseases.
9
Beck, Philipp, Christian Dubiella e Michael Groll. "Covalent and non-covalent reversible proteasome inhibition". Biological Chemistry 393, n. 10 (1 ottobre 2012): 1101–20. http://dx.doi.org/10.1515/hsz-2012-0212.
Abstract (sommario):
Abstract The 20S proteasome core particle (CP) is the proteolytically active key element of the ubiquitin proteasome system that directs the majority of intracellular protein degradation in eukaryotic cells. Over the past decade, the CP has emerged as an anticancer therapy target after approval of the first-in-class drug bortezomib (Velcade®) by the US Food and Drug Administration. However, bortezomib and all second-generation CP inhibitors that are currently explored in clinical phase studies react covalently and most often irreversibly with the proteolytic sites of the CP, hereby causing permanent CP blockage. Furthermore, reactive head groups result in unspecific binding to proteasomal active centers and in substantial enzymatic off-target activities that translate to severe side effects. Thus, reversible proteasome inhibitors might be a promising alternative, overcoming these drawbacks, but are challenging with respect to their urge for thorough enthalpic and entropic optimization. This review describes developments in the hitherto neglected field of reversible proteasome inhibitors focusing on insights gained from crystal structures, which provide valuable knowledge and strategies for future directions in drug development.
10
Piestrzeniewicz, Mariola K., Dorota Wilmańska, Janusz Szemraj, Kazimierz Studzian e Marek Gniazdowski. "Interactions of Novel Morpholine and Hexamethylene Derivatives of Anthracycline Antibiotics with DNA". Zeitschrift für Naturforschung C 59, n. 9-10 (1 ottobre 2004): 739–48. http://dx.doi.org/10.1515/znc-2004-9-1020.
Abstract (sommario):
Abstract Doxorubicin (DOX), daunorubicin (DRB), epidoxorubicin (EDOX) and their analogues with a 3′-NH2 group in daunosamine form a covalent bond with a 2-NH2 group of guanine via a methylene group from formaldehyde (CH2O). It is assumed that a Schiff base type intermediate is formed between CH2O and the 3′-NH2 group in the reaction. This reaction is supposed to occur in the cell. New analogues of anthracyclines with formamidine functionality bound to C-3′ of daunosamine and containing the bulky morpholine (DRBM, DOXM and EDOXM) or hexamethyleneimine rings attached are studied in our laboratory. These substituents decrease the association of the drugs to DNA and potentially hinder the formation of Schiff base-intermediates. Our experiments indicate that the formation of the covalent complexes by DRB, DOX and EDOX under these conditions is confirmed by a high enhancement (17-40x) of the inhibition of overall RNA synthesis by E. coli RNA polymerase on T7 DNA. DRBM and DOXM exhibit a lower enhancement of the inhibition by CH2O (7-13x). The other analogues show a 1.6-3x increase of inhibition. Hence, their covalent binding is lower than that of the parent compounds. These conclusions are confirmed by spectrophotometric estimations following removal of non-covalently associated drugs. Electrophoretic analysis of drug-DNA complexes formed in the presence of CH2O indicates that DRBM and DOXM as their parent compounds induce labile cross-links in DNA. Comparison of the results obtained at the subcellular level with cytotoxicity estimations indicates that there is a correlation between cytotoxicity of the anthracyclines on L1210 cells and transcriptional template activity of drug-DNA complexes formed in the presence of CH2O (r = 0.64; n = 9). These data confirm a notion that covalent attachment of anthracyclines to DNA is an essential event leading to cytotoxicity.
11
Müller, Patrick, Mergim Meta, Jan Laurenz Meidner, Marvin Schwickert, Jessica Meyr, Kevin Schwickert, Christian Kersten et al. "Investigation of the Compatibility between Warheads and Peptidomimetic Sequences of Protease Inhibitors—A Comprehensive Reactivity and Selectivity Study". International Journal of Molecular Sciences 24, n. 8 (13 aprile 2023): 7226. http://dx.doi.org/10.3390/ijms24087226.
Abstract (sommario):
Covalent peptidomimetic protease inhibitors have gained a lot of attention in drug development in recent years. They are designed to covalently bind the catalytically active amino acids through electrophilic groups called warheads. Covalent inhibition has an advantage in terms of pharmacodynamic properties but can also bear toxicity risks due to non-selective off-target protein binding. Therefore, the right combination of a reactive warhead with a well-suited peptidomimetic sequence is of great importance. Herein, the selectivities of well-known warheads combined with peptidomimetic sequences suited for five different proteases were investigated, highlighting the impact of both structure parts (warhead and peptidomimetic sequence) for affinity and selectivity. Molecular docking gave insights into the predicted binding modes of the inhibitors inside the binding pockets of the different enzymes. Moreover, the warheads were investigated by NMR and LC-MS reactivity assays against serine/threonine and cysteine nucleophile models, as well as by quantum mechanics simulations.
12
Gerling, Katharina, Sabrina Ölschläger, Meltem Avci-Adali, Bernd Neumann, Ernst Schweizer, Christian Schlensak, Hans-Peter Wendel e Sandra Stoppelkamp. "A Novel C1-Esterase Inhibitor Oxygenator Coating Prevents FXII Activation in Human Blood". Biomolecules 10, n. 7 (13 luglio 2020): 1042. http://dx.doi.org/10.3390/biom10071042.
Abstract (sommario):
The limited hemocompatibility of currently used oxygenator membranes prevents long-term use of artificial lungs in patients with lung failure. To improve hemocompatibility, we developed a novel covalent C1-esterase inhibitor (C1-INH) coating. Besides complement inhibition, C1-INH also prevents FXII activation, a very early event of contact phase activation at the crossroads of coagulation and inflammation. Covalently coated heparin, as the current anticoagulation gold standard, served as control. Additionally, a combination of both coatings (C1-INH/heparin) was established. The coatings were tested for their hemocompatibility by dynamic incubation with freshly drawn human whole blood. The analysis of various blood and plasma parameters revealed that C1-INH-containing coatings were able to markedly reduce FXIIa activity compared to heparin coating. Combined C1-INH/heparin coatings yielded similarly low levels of thrombin-antithrombin III complex formation as heparin coating. In particular, adhesion of monocytes and platelets as well as the diminished formation of fibrin networks were observed for combined coatings. We could show for the first time that a covalent coating with complement inhibitor C1-INH was able to ameliorate hemocompatibility. Thus, the early inhibition of the coagulation cascade is likely to have far-reaching consequences for the other cross-reacting plasma protein pathways.
13
Stellmacher, Lena, Tatyana Sandalova, Sarah Schneider, Gunter Schneider, Georg A. Sprenger e Anne K. Samland. "Novel mode of inhibition byD-tagatose 6-phosphate through a Heyns rearrangement in the active site of transaldolase B variants". Acta Crystallographica Section D Structural Biology 72, n. 4 (24 marzo 2016): 467–76. http://dx.doi.org/10.1107/s2059798316001170.
Abstract (sommario):
Transaldolase B (TalB) and D-fructose-6-phosphate aldolase A (FSAA) fromEscherichia coliare C—C bond-forming enzymes. Using kinetic inhibition studies and mass spectrometry, it is shown that enzyme variants of FSAA and TalB that exhibit D-fructose-6-phosphate aldolase activity are inhibited covalently and irreversibly by D-tagatose 6-phosphate (D-T6P), whereas no inhibition was observed for wild-type transaldolase B fromE. coli. The crystal structure of the variant TalBF178Ywith bound sugar phosphate was solved to a resolution of 1.46 Å and revealed a novel mode of covalent inhibition. The sugar is bound covalentlyviaits C2 atom to the ∊-NH2group of the active-site residue Lys132. It is neither bound in the open-chain form nor as the closed-ring form of D-T6P, but has been converted to β-D-galactofuranose 6-phosphate (D-G6P), a five-membered ring structure. The furanose ring of the covalent adduct is formedviaa Heyns rearrangement and subsequent hemiacetal formation. This reaction is facilitated by Tyr178, which is proposed to act as acid–base catalyst. The crystal structure of the inhibitor complex is compared with the structure of the Schiff-base intermediate of TalBE96Qformed with the substrate D-fructose 6-phosphate determined to a resolution of 2.20 Å. This comparison highlights the differences in stereochemistry at the C4 atom of the ligand as an essential determinant for the formation of the inhibitor adduct in the active site of the enzyme.
14
Betori, Rick C., Yue Liu, Rama K. Mishra, Scott B. Cohen, Stephen J. Kron e Karl A. Scheidt. "Targeted Covalent Inhibition of Telomerase". ACS Chemical Biology 15, n. 3 (4 febbraio 2020): 706–17. http://dx.doi.org/10.1021/acschembio.9b00945.
15
Ghosh, Avick Kumar, Indranil Samanta, Anushree Mondal e Wenshe Ray Liu. "Covalent Inhibition in Drug Discovery". ChemMedChem 14, n. 9 (26 marzo 2019): 889–906. http://dx.doi.org/10.1002/cmdc.201900107.
16
Rao, AK, e MA Kowalska. "ADP-induced platelet shape change and mobilization of cytoplasmic ionized calcium are mediated by distinct binding sites on platelets: 5'- p-fluorosulfonylbenzoyladenosine is a weak platelet agonist". Blood 70, n. 3 (1 settembre 1987): 751–56. http://dx.doi.org/10.1182/blood.v70.3.751.751.
Abstract (sommario):
Abstract Platelet stimulation with ADP results in several responses, including shape change, increase in cytoplasmic ionized calcium concentration [Ca2+]i, an inhibition of adenylate cyclase. 5′-p-Fluorosulphonyl benzoyladenosine (FSBA), which covalently labels an ADP binding site on platelets, blocks platelet shape change but not the inhibition of cyclic AMP levels by ADP, whereas p-chloromercuribenzenesulfonate (pCMBS), a nonpenetrating thiol reagent, has the opposite effects. We examined the effect of FSBA and pCMBS on ADP-induced increase in [Ca2+]i using platelets loaded with fluorescent Ca2+ indicators quin2 and fura-2. FSBA (50 to 200 mumol/L) induced a dose-dependent rise in [Ca2+]i, indicating that it is a weak platelet agonist. Under conditions of covalent labeling of the ADP binding sites, FSBA (50 to 100 mumol/L) did not inhibit the ADP-induced increase in [Ca2+]i or its inhibition of adenylate cyclase, whereas pCMBS (up to 1 mmol/L) abolished both these responses but not shape change. These findings suggest that ADP-induced Ca2+ mobilization and inhibition of adenylate cyclase are mediated by platelet binding sites distinct from those mediating shape change.
17
Rao, AK, e MA Kowalska. "ADP-induced platelet shape change and mobilization of cytoplasmic ionized calcium are mediated by distinct binding sites on platelets: 5'- p-fluorosulfonylbenzoyladenosine is a weak platelet agonist". Blood 70, n. 3 (1 settembre 1987): 751–56. http://dx.doi.org/10.1182/blood.v70.3.751.bloodjournal703751.
Abstract (sommario):
Platelet stimulation with ADP results in several responses, including shape change, increase in cytoplasmic ionized calcium concentration [Ca2+]i, an inhibition of adenylate cyclase. 5′-p-Fluorosulphonyl benzoyladenosine (FSBA), which covalently labels an ADP binding site on platelets, blocks platelet shape change but not the inhibition of cyclic AMP levels by ADP, whereas p-chloromercuribenzenesulfonate (pCMBS), a nonpenetrating thiol reagent, has the opposite effects. We examined the effect of FSBA and pCMBS on ADP-induced increase in [Ca2+]i using platelets loaded with fluorescent Ca2+ indicators quin2 and fura-2. FSBA (50 to 200 mumol/L) induced a dose-dependent rise in [Ca2+]i, indicating that it is a weak platelet agonist. Under conditions of covalent labeling of the ADP binding sites, FSBA (50 to 100 mumol/L) did not inhibit the ADP-induced increase in [Ca2+]i or its inhibition of adenylate cyclase, whereas pCMBS (up to 1 mmol/L) abolished both these responses but not shape change. These findings suggest that ADP-induced Ca2+ mobilization and inhibition of adenylate cyclase are mediated by platelet binding sites distinct from those mediating shape change.
18
Bjij, Imane, Fisayo A. Olotu, Clement Agoni, Emmanuel Adeniji, Shama Khan, Ahmed El Rashedy, Driss Cherqaoui e Mahmoud E. S. Soliman. "Covalent Inhibition in Drug Discovery: Filling the Void in Literature". Current Topics in Medicinal Chemistry 18, n. 13 (4 ottobre 2018): 1135–45. http://dx.doi.org/10.2174/1568026618666180731161438.
Abstract (sommario):
The serendipitous discovery of covalent inhibitors and their characteristic potency of inducing irreversible and complete inhibition in therapeutic targets have caused a paradigm shift from the use of non-covalent drugs in disease treatment. This has caused a significant evolution in the field of covalent targeting to understand their inhibitory mechanisms and facilitate the systemic design of novel covalent modifiers for ‘undruggable’ targets. Computational techniques have evolved over the years and have significantly contributed to the process of drug discovery by mirroring the pattern of biological occurrences thereby providing insights into the dynamics and conformational transitions associated with biomolecular interactions. Moreover, our previous contributions towards the systematic design of selective covalent modifiers have revealed the various setbacks associated with the use of these conventional techniques in the study of covalent systems, hence there is a need for distinct approaches. In this review, we highlight the modifications and development of computational techniques suitable for covalent systems, their lapses, shortcomings and recent advancements.
19
Strelow, John M. "A Perspective on the Kinetics of Covalent and Irreversible Inhibition". SLAS DISCOVERY: Advancing the Science of Drug Discovery 22, n. 1 (5 ottobre 2016): 3–20. http://dx.doi.org/10.1177/1087057116671509.
Abstract (sommario):
The clinical and commercial success of covalent drugs has prompted a renewed and more deliberate pursuit of covalent and irreversible mechanisms within drug discovery. A covalent mechanism can produce potent inhibition in a biochemical, cellular, or in vivo setting. In many cases, teams choose to focus on the consequences of the covalent event, defined by an IC50 value. In a biochemical assay, the IC50 may simply reflect the target protein concentration in the assay. What has received less attention is the importance of the rate of covalent modification, defined by kinact/KI. The kinact/KI is a rate constant describing the efficiency of covalent bond formation resulting from the potency (KI) of the first reversible binding event and the maximum potential rate (kinact) of inactivation. In this perspective, it is proposed that the kinact/KI should be employed as a critical parameter to identify covalent inhibitors, interpret structure-activity relationships (SARs), translate activity from biochemical assays to the cell, and more accurately define selectivity. It is also proposed that a physiologically relevant kinact/KI and an (unbound) AUC generated from a pharmacokinetic profile reflecting direct exposure of the inhibitor to the target protein are two critical determinants of in vivo covalent occupancy. A simple equation is presented to define this relationship and improve the interpretation of covalent and irreversible kinetics.
20
Kilbourn, R., e G. Lopez-Berestein. "Protease inhibitors block the macrophage-mediated inhibition of tumor cell mitochondrial respiration." Journal of Immunology 144, n. 3 (1 febbraio 1990): 1042–45. http://dx.doi.org/10.4049/jimmunol.144.3.1042.
Abstract (sommario):
Abstract The antitumor activity of activated macrophages toward tumor cells, in vitro, appears to involve the production of toxic nitrogen intermediates. These intermediates, particularly nitric oxide, have been shown to cause the inhibition of cell division and to decrease cellular respiration by inhibiting electron transport. We studied the effects of proteolytic inhibitors on macrophage-mediated inhibition of L1210 tumor cell respiration and DNA synthesis, and found that chloromethyl ketone derivatives, which covalently modify serine proteases, can block macrophage cytotoxicity. Furthermore, these inhibitors decrease nitrite production by activated macrophages suggesting that the mechanism of action involves the inhibition of nitric oxide production.
21
Colman, RW, WR Figures, LM Scearce, AM Strimpler, FX Zhou e AK Rao. "Inhibition of collagen-induced platelet activation by 5'-p- fluorosulfonylbenzoyl adenosine: evidence for an adenosine diphosphate requirement and synergistic influence of prostaglandin endoperoxides". Blood 68, n. 2 (1 agosto 1986): 565–70. http://dx.doi.org/10.1182/blood.v68.2.565.565.
Abstract (sommario):
Abstract The relative roles of platelet autacoids such as adenosine diphosphate (ADP), prostaglandin endoperoxides, and thromboxane A2 (TXA2) in collagen-induced platelet activation are not fully understood. We reexamined this relationship using the ADP affinity analogue, 5'-p- fluorosulfonylbenzoyl adenosine (FSBA), which covalently modifies a receptor for ADP on the platelet surface, thereby inhibiting ADP- induced platelet activation. Collagen-induced shape change, aggregation, and fibrinogen binding were each fully inhibited under conditions in which FSBA is covalently incorporated and could not be overcome by raising the collagen used to supramaximal concentrations. In contrast, TXA2 synthesis stimulated by collagen under conditions that produced maximum aggregation was only minimally inhibited by FSBA. Since covalent incorporation of FSBA has been previously shown to specifically inhibit ADP-induced activation of platelets, the present study supports the contention that ADP is required for collagen-induced platelet activation. Under similar conditions, indomethacin, an inhibitor of cyclooxygenase, inhibited collagen-induced shape change, indicating that endoperoxides and/or TXA2 also play a role in this response. Shape change induced by low concentrations (10 nmol/L) of the stable prostaglandin endoperoxide, azo-PGH2, was also inhibited by FSBA. These observations indicate a role for ADP in responses elicited by low concentrations of endoperoxides. However, at higher concentrations of azo-PGH2 (100 nmol/L), inhibition by FSBA could be overcome. Thus, the effect of collagen apparently has an absolute requirement for ADP for aggregation and fibrinogen binding and for both ADP and prostaglandins for shape change. Aggregation and fibrinogen binding induced by prostaglandin endoperoxides also required ADP as a mediator, but ADP is not absolutely required at high endoperoxide concentration to induce shape change.
22
Colman, RW, WR Figures, LM Scearce, AM Strimpler, FX Zhou e AK Rao. "Inhibition of collagen-induced platelet activation by 5'-p- fluorosulfonylbenzoyl adenosine: evidence for an adenosine diphosphate requirement and synergistic influence of prostaglandin endoperoxides". Blood 68, n. 2 (1 agosto 1986): 565–70. http://dx.doi.org/10.1182/blood.v68.2.565.bloodjournal682565.
Abstract (sommario):
The relative roles of platelet autacoids such as adenosine diphosphate (ADP), prostaglandin endoperoxides, and thromboxane A2 (TXA2) in collagen-induced platelet activation are not fully understood. We reexamined this relationship using the ADP affinity analogue, 5'-p- fluorosulfonylbenzoyl adenosine (FSBA), which covalently modifies a receptor for ADP on the platelet surface, thereby inhibiting ADP- induced platelet activation. Collagen-induced shape change, aggregation, and fibrinogen binding were each fully inhibited under conditions in which FSBA is covalently incorporated and could not be overcome by raising the collagen used to supramaximal concentrations. In contrast, TXA2 synthesis stimulated by collagen under conditions that produced maximum aggregation was only minimally inhibited by FSBA. Since covalent incorporation of FSBA has been previously shown to specifically inhibit ADP-induced activation of platelets, the present study supports the contention that ADP is required for collagen-induced platelet activation. Under similar conditions, indomethacin, an inhibitor of cyclooxygenase, inhibited collagen-induced shape change, indicating that endoperoxides and/or TXA2 also play a role in this response. Shape change induced by low concentrations (10 nmol/L) of the stable prostaglandin endoperoxide, azo-PGH2, was also inhibited by FSBA. These observations indicate a role for ADP in responses elicited by low concentrations of endoperoxides. However, at higher concentrations of azo-PGH2 (100 nmol/L), inhibition by FSBA could be overcome. Thus, the effect of collagen apparently has an absolute requirement for ADP for aggregation and fibrinogen binding and for both ADP and prostaglandins for shape change. Aggregation and fibrinogen binding induced by prostaglandin endoperoxides also required ADP as a mediator, but ADP is not absolutely required at high endoperoxide concentration to induce shape change.
23
Tian, Yafeng, Mi Zhang, Panpan Heng, Hua Hou e Baoshan Wang. "Computational Investigations on Reaction Mechanisms of the Covalent Inhibitors Ponatinib and Analogs Targeting the Extracellular Signal-Regulated Kinases". International Journal of Molecular Sciences 24, n. 20 (16 ottobre 2023): 15223. http://dx.doi.org/10.3390/ijms242015223.
Abstract (sommario):
As an important cancer therapeutic target, extracellular signal-regulated kinases (ERK) are involved in triggering various cellular responses in tumors. Regulation of the ERK signaling pathway by the small molecular inhibitors is highly desired for the sake of cancer therapy. In contrast to the routine inhibitors targeting ERKs through long-range non-bonding interactions, Ponatinib, a covalent inhibitor to ERK2 with a macrocyclic structure characterized by the α,β-C=C unsaturated ketone, can form the stable -C(S)-C(H)-type complex via the four-center barrier due to the nucleophilic addition reaction of the thiol group of the Cys166 residue of ERK2 with the C=C double bond of Ponatinib with reaction free-energy barrier of 47.2 kcal/mol. Reaction mechanisms for the covalent binding were calculated using QM/MM methods and molecular dynamics simulations. The interaction modes and the corresponding binding free energies were obtained for the non-covalent and covalent complexation. The binding free energies of the non-covalent and covalent inhibitions are 14.8 kcal/mol and 33.4 kcal/mol, respectively. The mechanistic study stimulated a rational design on the modified Ponatinib structure by substituting the C=C bond with the C=N bond. It was demonstrated that the new compound exhibits better inhibition activity toward ERK2 in term of both thermodynamic and kinetic aspects through the covalent binding with a lower reaction free-energy barrier of 23.1 kcal/mol. The present theoretical work sheds new light on the development of the covalent inhibitors for the regulation of ERKs.
24
Zhou, X. Edward, Kelly Suino-Powell, Chad R. Schultz, Bilal Aleiwi, Joseph S. Brunzelle, Jared Lamp, Irving E. Vega, Edmund Ellsworth, André S. Bachmann e Karsten Melcher. "Structural basis of binding and inhibition of ornithine decarboxylase by 1-amino-oxy-3-aminopropane". Biochemical Journal 478, n. 23 (6 dicembre 2021): 4137–49. http://dx.doi.org/10.1042/bcj20210647.
Abstract (sommario):
Ornithine decarboxylase (ODC) is the rate-limiting enzyme for the synthesis of polyamines (PAs). PAs are oncometabolites that are required for proliferation, and pharmaceutical ODC inhibition is pursued for the treatment of hyperproliferative diseases, including cancer and infectious diseases. The most potent ODC inhibitor is 1-amino-oxy-3-aminopropane (APA). A previous crystal structure of an ODC–APA complex indicated that APA non-covalently binds ODC and its cofactor pyridoxal 5-phosphate (PLP) and functions by competing with the ODC substrate ornithine for binding to the catalytic site. We have revisited the mechanism of APA binding and ODC inhibition through a new crystal structure of APA-bound ODC, which we solved at 2.49 Å resolution. The structure unambiguously shows the presence of a covalent oxime between APA and PLP in the catalytic site, which we confirmed in solution by mass spectrometry. The stable oxime makes extensive interactions with ODC but cannot be catabolized, explaining APA's high potency in ODC inhibition. In addition, we solved an ODC/PLP complex structure with citrate bound at the substrate-binding pocket. These two structures provide new structural scaffolds for developing more efficient pharmaceutical ODC inhibitors.
25
Kinoshita, T., A. W. Dodds, S. K. A. Law e K. Inoue. "The low C5 convertase activity of the C4A6 allotype of human complement component C4". Biochemical Journal 261, n. 3 (1 agosto 1989): 743–48. http://dx.doi.org/10.1042/bj2610743.
Abstract (sommario):
We have compared the C5-convertase-forming ability of different C4 allotypes, including the C4A6 allotype, which has low haemolytic activity and which has previously been shown to be defective in C5-convertase formation. Recent studies suggest that C4 plays two roles in the formation of the C5 convertase from the C3 convertase. Firstly, C4b acts as the binding site for C3 which, upon cleavage by C2, forms a covalent linkage with the C4b. Secondly, C4b with covalently attached C3b serves to form a high-affinity binding site for C5. Purified allotypes C4A3, C4B1 and C4A6 were used to compare these two activities of C4. Covalently linked C4b-C3b complexes were formed on sheep erythrocytes with similar efficiency by using C4A3 and C4B1, indicating that the two isotypes behave similarly as acceptors for covalent attachment of C3b. C4A6 showed normal efficiency in this function. However, cells bearing C4b-C3b complexes made from C4A6 contained only a small number of high-affinity binding sites for C5. Therefore a lack of binding of C5 to the C4b C3b complexes is the reason for the inefficient formation of C5 convertase by C4A6. The small number of high-affinity binding sites created, when C4A6 was used, were tested for inhibition by anti-C3 and anti-C4. Anti-C4 did not inhibit C5 binding, whereas anti-C3 did. This suggests that the sites created when C4A6 is used to make C3 convertase may be C3b-C3b dimers, and hence the low haemolytic activity of C4A6 results from the creation of low numbers of alternative-pathway C5-convertase sites.
26
Stevic, Ivan, Howard H. W. Chan, Ankush Chander, Leslie R. Berry e Anthony K. C. Chan. "Covalently linking heparin to antithrombin enhances prothrombinase inhibition on activated platelets". Thrombosis and Haemostasis 109, n. 06 (2013): 1016–24. http://dx.doi.org/10.1160/th12-10-0766.
Abstract (sommario):
SummaryFactor (F)Xa within the prothrombinase complex is protected from inhibition by unfractionated heparin (UFH), enoxaparin and fondaparinux. We have developed a covalent antithrombin-heparin complex (ATH) with enhanced anticoagulant activity. We have also demonstrated that ATH is superior at inhibiting coagulation factors when assembled on artificial surfaces. The objective of the present study is to determine the ability of ATH vs AT+UFH to inhibit FXa within the prothrombinase complex when the enzyme complex is assembled on the more native platelet system. Discontinuous inhibition assays were performed to determine final k 2-values for inhibition of FXa, FXa within the platelet-prothrombinase, or FXa within prothrombinase devoid of various components. Thrombin generation and plasma clotting was also assayed in the presence of resting/activated platelets ± inhibitors. Protection of FXa was not observed for ATH, whereas a moderate 60% protection was observed for AT+UFH. ATH inhibited platelet-prothrombinase ∼4-fold faster than AT+UFH. Relative to intact prothrombinase, rates for FXa inhibition by AT+UFH in prothrombinase complexes devoid of either prothrombin (II)/activated platelets/FVa were higher. However, inhibition by AT+UFH of prothrombinase devoid of FII yielded slightly lower rates compared to free FXa inhibition. Thrombin generation and plasma clotting was enhanced with activated platelets, while inhibition was better by ATH compared to AT+UFH, thus suggesting an overall enhanced anticoagulant activity of ATH against platelet-bound prothrombinase complexes.
27
Sim, E., A. W. Dodds e A. Goldin. "Inhibition of the covalent binding reaction of complement component C4 by penicillamine, an anti-rheumatic agent". Biochemical Journal 259, n. 2 (15 aprile 1989): 415–19. http://dx.doi.org/10.1042/bj2590415.
Abstract (sommario):
D(-)-Penicillamine [D(-)-beta beta-dimethylcysteine] is an anti-arthritic drug, but its use is limited by adverse side effects, which include problems in immune-complex clearance. Complement is important as a source of inflammatory mediators in rheumatoid arthritis and is also involved in immune-complex clearance. Thus inhibition of the complement cascade would be likely to contribute to both the therapeutic and the toxic effects of penicillamine. It is shown that penicillamine and cysteine are potent inhibitors of the covalent binding of activated complement component C4 to immune complexes. [35S]Cysteine itself becomes covalently bound to C4b through the thioester site. Penicillamine and cysteine are more reactive with the C4A isotype than with the C4B isotype of the HLA class III protein C4. The limited amino acid sequence differences between C4A and C4B include a cysteine/serine interchange, and it is suggested that the cysteine residue in C4A contributes to the increased rate of reaction of C4A with the alpha-amino-beta-thiol compounds.
28
Lu, Jia-Hui, Zhen Li, Jia-Hui Chen, Shu-Liang Li, Jie-Hao He, Song Gu, Bo-Wen Liu, Li Chen e Yu-Zhong Wang. "Adaptable Phosphate Networks towards Robust, Reprocessable, Weldable, and Alertable-Yet-Extinguishable Epoxy Vitrimer". Research 2022 (6 ottobre 2022): 1–12. http://dx.doi.org/10.34133/2022/9846940.
Abstract (sommario):
Covalent adaptable networks (CANs) combine the uniqueness of thermoplastics and thermosets to allow for reprocessability while being covalently crosslinked. However, it is highly desirable but rarely achieved for CANs to simultaneously demonstrate reversibility and mechanical robustness. Herein, we report a feasible strategy to develop a novel epoxy vitrimer (EV) composed of adaptable phosphate networks (APNs), by which the EVs exhibit promising mechanical properties (tensile strength of 62.5 ~ 87.8 MPa and tensile modulus of 1360.1 ~ 2975.3 MPa) under ambient conditions. At elevated temperatures, the topology rearrangement occurs relied on phosphate transesterification, which contributes to the shape memory performance, self-healing, reprocessing, and welding behaviors. Moreover, the incorporation of APNs allows for improvements in anti-ignition and also the inhibition of both heat release and smoke generation to avoid empyrosis, asphyxiation, and toxication during burning, showing expected intrinsic fire safety. Thermal, mechanical properties, and flame retardancy of the reprocessed EVs after hot pressing are very close to those of the original EVs, which is attributed to the sufficient reversibility of APNs. Accordingly, combining the aforementioned features, EVs are manufactured as flame-triggered switches for fire alarms, which symbolizes the innovative development of high-performance covalent adaptable polymeric materials.
29
Ruddraraju, Kasi Viswanatharaju, e Zhong-Yin Zhang. "Covalent inhibition of protein tyrosine phosphatases". Molecular BioSystems 13, n. 7 (2017): 1257–79. http://dx.doi.org/10.1039/c7mb00151g.
30
Zhao, Yuguang, Fredrik Svensson, David Steadman, Sarah Frew, Amy Monaghan, Magda Bictash, Tiago Moreira et al. "Structural Insights into Notum Covalent Inhibition". Journal of Medicinal Chemistry 64, n. 15 (22 luglio 2021): 11354–63. http://dx.doi.org/10.1021/acs.jmedchem.1c00701.
31
Westover, Kenneth D., Pasi A. Jänne e Nathanael S. Gray. "Progress on Covalent Inhibition of KRASG12C". Cancer Discovery 6, n. 3 (marzo 2016): 233–34. http://dx.doi.org/10.1158/2159-8290.cd-16-0092.
32
Jacobs, Amy. "Covalent Inhibition of HIV Membrane Fusion". Biophysical Journal 96, n. 3 (febbraio 2009): 359a. http://dx.doi.org/10.1016/j.bpj.2008.12.1814.
33
Huang, Huang, Christina A. Howard, Sergei Zari, Hyo Je Cho, Shirish Shukla, Hao Li, Juliano Ndoj et al. "Covalent inhibition of NSD1 histone methyltransferase". Nature Chemical Biology 16, n. 12 (31 agosto 2020): 1403–10. http://dx.doi.org/10.1038/s41589-020-0626-6.
34
Amara, Neri, Roi Mashiach, Dotan Amar, Pnina Krief, Stéphane A. H. Spieser, Matthew J. Bottomley, Amir Aharoni e Michael M. Meijler. "Covalent Inhibition of Bacterial Quorum Sensing". Journal of the American Chemical Society 131, n. 30 (5 agosto 2009): 10610–19. http://dx.doi.org/10.1021/ja903292v.
35
Kentner, Taryn A., Leslie R. Berry e Anthony K. C. Chan. "Inhibition of Factor Xa in Prothrombinase Is Enhanced by Covalent Linkage of Antithrombin to Heparin." Blood 104, n. 11 (16 novembre 2004): 1051. http://dx.doi.org/10.1182/blood.v104.11.1051.1051.
Abstract (sommario):
Abstract Prothrombinase is the surface-bound complex in which factor Xa (Xa) converts prothrombin to thrombin in vivo. Studies have shown that Xa within prothrombinase is resistant to inhibition by antithrombin + heparin (AT+H). Previously we found that, unlike AT+H, a covalent conjugate of AT and H (ATH) was able to neutralize fibrin-bound thrombin. In this study, AT+H and ATH were compared in their reaction with Xa in prothrombinase. Mixtures of CaCl2, PCPS vesicles, factor Va (Va) and prothrombin in TSP buffer were combined with either AT+H or ATH. Following addition of Xa, time samples were neutralized with Na2EDTA + polybrene + substrate (S-2222) and residual Xa activity measured. Second order rate constants (k2) were calculated from plots of activity versus time. Results were compared to those in corresponding experiments with Xa alone. Inhibition of Xa in prothrombinase by AT+H had a k2 (x 108 M−1min−1) of 0.688 +/− 0.030. Conversely, free Xa neutralization by AT+H was significantly more rapid (k2 = 1.53 +/− 0.35, p = 0.041). ATH reaction with prothrombinase Xa proceeded at a rate that was insignificantly different from that with Xa alone (2.36 +/− 0.31 and 2.83 +/− 0.83, respectively) and similar to AT+H reaction with free Xa. Varying concentrations of prothrombinase components showed similar effects. We conclude that covalently linked complexes of AT + H undergo unhindered reaction with Xa in prothrombinase, while non-covalently linked AT+H encounters resistance. It is possible that ATH may effectively prevent plasma thrombin generation by neutralization of Xa in prothrombinase and thus may prohibit feedback activation of coagulation.
36
NAGUMO, Yoko, Hideaki KAKEYA, Mitsuru SHOJI, Yujiro HAYASHI, Naoshi DOHMAE e Hiroyuki OSADA. "Epolactaene binds human Hsp60 Cys442 resulting in the inhibition of chaperone activity". Biochemical Journal 387, n. 3 (26 aprile 2005): 835–40. http://dx.doi.org/10.1042/bj20041355.
Abstract (sommario):
Epolactaene is a microbial metabolite isolated from Penicillium sp., from which we synthesized its derivative ETB (epolactaene tertiary butyl ester). In the present paper, we report on the identification of the binding proteins of epolactaene/ETB, and the results of our investigation into its inhibitory mechanism. Using biotin-labelled derivatives of epolactaene/ETB, human Hsp (heat-shock protein) 60 was identified as a binding protein of epolactaene/ETB in vitro as well as in situ. In addition, we found that Hsp60 pre-incubated with epolactaene/ETB lost its chaperone activity. The in vitro binding study showed that biotin-conjugated epolactaene/ETB covalently binds to Hsp60. In order to investigate the binding site, binding experiments with alanine mutants of Hsp60 cysteine residues were conducted. As a result, it was suggested that Cys442 is responsible for the covalent binding with biotin-conjugated epolactaene/ETB. Furthermore, the replacement of Hsp60 Cys442 with an alanine residue renders the chaperone activity resistant to ETB inhibition, while the alanine replacement of other cysteine residues do not. These results indicate that this cysteine residue is alkylated by ETB, leading to Hsp60 inactivation.
37
Jebaraj, Billy Michael Chelliah, Annika Müller, Rashmi Priyadharshini Dheenadayalan, Sascha Endres, Philipp M. Roessner, Felix Seyfried, Claudia Walliser et al. "Evaluation of vecabrutinib as a model for noncovalent BTK/ITK inhibition for treatment of chronic lymphocytic leukemia". Blood 139, n. 6 (10 febbraio 2022): 859–75. http://dx.doi.org/10.1182/blood.2021011516.
Abstract (sommario):
Abstract Covalent Bruton tyrosine kinase (BTK) inhibitors, such as ibrutinib, have proven to be highly beneficial in the treatment of chronic lymphocytic leukemia (CLL). Interestingly, the off-target inhibition of IL-2-inducible T-cell kinase (ITK) by ibrutinib may also play a role in modulating the tumor microenvironment, potentially enhancing the treatment benefit. However, resistance to covalently binding BTK inhibitors can develop as the result of a mutation in cysteine 481 of BTK (C481S), which prevents irreversible binding of the drugs. In the present study we performed preclinical characterization of vecabrutinib, a next-generation noncovalent BTK inhibitor that has ITK-inhibitory properties similar to those of ibrutinib. Unlike ibrutinib and other covalent BTK inhibitors, vecabrutinib showed retention of the inhibitory effect on C481S BTK mutants in vitro, similar to that of wild-type BTK. In the murine Eμ-TCL1 adoptive transfer model, vecabrutinib reduced tumor burden and significantly improved survival. Vecabrutinib treatment led to a decrease in CD8+ effector and memory T-cell populations, whereas the naive populations were increased. Of importance, vecabrutinib treatment significantly reduced the frequency of regulatory CD4+ T cells in vivo. Unlike ibrutinib, vecabrutinib treatment showed minimal adverse impact on the activation and proliferation of isolated T cells. Lastly, combination treatment with vecabrutinib and venetoclax augmented treatment efficacy, significantly improved survival, and led to favorable reprogramming of the microenvironment in the murine Eμ-TCL1 model. Thus, noncovalent BTK/ITK inhibitors, such as vecabrutinib, may be efficacious in C481S BTK mutant CLL while preserving the T-cell immunomodulatory function of ibrutinib.
38
Wágner, Gábor, Tamara A. M. Mocking, Albert J. Kooistra, Inna Slynko, Péter Ábrányi-Balogh, György M. Keserű, Maikel Wijtmans, Henry F. Vischer, Iwan J. P. de Esch e Rob Leurs. "Covalent Inhibition of the Histamine H3 Receptor". Molecules 24, n. 24 (11 dicembre 2019): 4541. http://dx.doi.org/10.3390/molecules24244541.
Abstract (sommario):
Covalent binding of G protein-coupled receptors by small molecules is a useful approach for better understanding of the structure and function of these proteins. We designed, synthesized and characterized a series of 6 potential covalent ligands for the histamine H3 receptor (H3R). Starting from a 2-amino-pyrimidine scaffold, optimization of anchor moiety and warhead followed by fine-tuning of the required reactivity via scaffold hopping resulted in the isothiocyanate H3R ligand 44. It shows high reactivity toward glutathione combined with appropriate stability in water and reacts selectively with the cysteine sidechain in a model nonapeptide equipped with nucleophilic residues. The covalent interaction of 44 with H3R was validated with washout experiments and leads to inverse agonism on H3R. Irreversible binder 44 (VUF15662) may serve as a useful tool compound to stabilize the inactive H3R conformation and to study the consequences of prolonged inhibition of the H3R.
39
Thøgersen, I. B., G. Salvesen, F. H. Brucato, S. V. Pizzo e J. J. Enghild. "Purification and characterization of an α-macroglobulin proteinase inhibitor from the mollusc Octopus vulgaris". Biochemical Journal 285, n. 2 (15 luglio 1992): 521–27. http://dx.doi.org/10.1042/bj2850521.
Abstract (sommario):
The cell-free haemolymph of the mollusc Octopus vulgaris inhibited the proteolytic activity of the thermolysin against the high-molecular-mass substrate hide powder azure. The purified inhibitor was a glycoprotein composed of two identical 180 kDa disulphide-linked subunits. In addition to the inhibition of the metalloproteinase thermolysin, the protein inhibited the serine proteinases human neutrophil elastase, pig pancreatic elastase, bovine chymotrypsin, bovine trypsin and the cysteine proteinase papain. A fraction of the proteinase-inhibitor complex resisted dissociation after denaturation indicating that some of the proteinase molecules became covalently bound. The nucleophile beta-aminopropionitrile decreased the covalent binding of proteinases to the Octopus vulgaris protein, suggesting that this interaction is mediated by an internal thiol ester; the reactivity and the amino acid sequence flanking the reactive residues of the putative thiol ester were consistent with this hypothesis. Bound trypsin remained active against the low-molecular-mass chromatogenic substrate H-D-Pro-Phe-Arg p-nitroanilide and was protected from inhibition by active-site-directed protein inhibitors of trypsin; however, the bound trypsin was readily inhibited by small synthetic inhibitors. This indicates that the inhibition of proteinases is accomplished by steric hindrance. The proteinase-inhibitory activity of this protein is characteristic of inhibition by mammalian alpha-macroglobulins and the presence of a putative thiol ester suggests that the Octopus vulgaris proteinase inhibitor is a homologue of human alpha 2-macroglobulin.
40
Shatan, Anastasiia B., Vitalii Patsula, Hana Macková, Andrii Mahun, Renáta Lehotská, Elena Piecková e Daniel Horák. "Silver-Sulfamethazine-Conjugated β-Cyclodextrin/Dextran-Coated Magnetic Nanoparticles for Pathogen Inhibition". Nanomaterials 14, n. 4 (17 febbraio 2024): 371. http://dx.doi.org/10.3390/nano14040371.
Abstract (sommario):
In the fight against antibiotic resistance, which is rising to dangerously high levels worldwide, new strategies based on antibiotic-conjugated biocompatible polymers bound to magnetic nanoparticles that allow the drug to be manipulated and delivered to a specific target are being proposed. Here, we report the direct surface engineering of nontoxic iron oxide nanoparticles (IONs) using biocompatible dextran (Dex) covalently linked to β-cyclodextrin (β-CD) with the ability to form non-covalent complexes with silver-sulfamethazine (SMT-Ag). To achieve a good interaction of β-CD-modified dextran with the surface of the nanoparticles, it was functionalized with diphosphonic acid (DPA) that provides strong binding to Fe atoms. The synthesized polymers and nanoparticles were characterized by various methods, such as nuclear magnetic resonance (NMR), Fourier transform infrared (FTIR) and ultraviolet–visible (UV–Vis) spectroscopies, transmission electron microscopy (TEM), thermogravimetric analysis (TGA), atomic absorption spectroscopy (AAS), dynamic light scattering (DLS), etc. The resulting magnetic ION@DPA-Dex-β-CD-SMT-Ag nanoparticles were colloidally stable in water and contained 24 μg of antibiotic per mg of the particles. When tested for in vitro antimicrobial activity on Gram-positive (Staphylococcus aureus) and Gram-negative (Escherichia coli) bacteria and fungi (yeast Candida albicans and mold Aspergillus niger), the particles showed promising potential.
41
Lee, Jesang, e Seung Bum Park. "Extended Applications of Small-Molecule Covalent Inhibitors toward Novel Therapeutic Targets". Pharmaceuticals 15, n. 12 (27 novembre 2022): 1478. http://dx.doi.org/10.3390/ph15121478.
Abstract (sommario):
Recently, small-molecule covalent inhibitors have been accepted as a practical tool for targeting previously “undruggable” proteins. The high target selectivity of modern covalent inhibitors is now alleviating toxicity concerns regarding the covalent modifications of proteins. However, despite the tremendous clinical success of current covalent inhibitors, there are still unmet medical needs that covalent inhibitors have not yet addressed. This review categorized representative covalent inhibitors based on their mechanism of covalent inhibition: conventional covalent inhibitors, targeted covalent inhibitors (TCIs), and expanded TCIs. By reviewing both Food and Drug Administration (FDA)-approved drugs and drug candidates from recent literature, we provide insight into the future direction of covalent inhibitor development.
42
Walderveen, Maria Christina Van, Leslie Roy Berry, Helen Mary Atkinson e Anthony Kam Chuen Chan. "Covalent antithrombin-heparin effect on thrombin-thrombomodulin and activated protein C reaction with factor V/Va". Thrombosis and Haemostasis 103, n. 05 (2010): 910–19. http://dx.doi.org/10.1160/th09-07-0473.
Abstract (sommario):
SummaryThrombomodulin (TM), which variably contains a chondroitin sulfate (±CS), forms an anticoagulant complex with thrombin (IIa). IIaTM(±CS) converts protein C (PC) into activated PC (APC), which then inactivates activated factors V (FVa) and VIII (FVIIIa). This reduces prothrombinase and tenase complexes that generate IIa. Heparin (H) increases the rate of IIa-TM inhibition by antithrombin (AT) and enhances FV cleavage by APC. Our novel covalent AT-H (ATH) product, has superior anticoagulant activity compared to AT + unfractionated H (UFH). We studied mechanisms by which ATH versus AT + UFH inhibits IIaTM(±CS), and ATH influences on APC cleavage of FV/FVa compared to UFH. Findings would determine how these reactions moderate ATH’s overall effects as an anticoagulant. Discontinuous second order rate inhibition assays of IIa-TM(±CS) inhibition by AT + UFH or ATH were performed in presence or absence of human umbilical vein endothelial cells (HUVECs). FV/FVa cleavage by APC in the presence of UFH or ATH was analysed by Western blots. ATH increased IIa-TM(±CS) inhibition to a greater degree than AT + UFH, both on plastic and HUVEC surfaces. Unlike UFH, ATH did not accelerate FV cleavage by APC, but ATH did enhance FVa cleavage relative to UFH. Increased IIa-TM inhibition by ATH downregulates PC activation. However, ATH does accelerate downstream inactivation of FVa, which increases its potency for IIa generation inhibition compared to UFH. This trend holds true in the presence of APC’s cofactor, protein S. Overall, ATH may have a balanced function towards inhibiting or accelerating PC pathway activities.
43
Allgardsson, Anders, Lotta Berg, Christine Akfur, Andreas Hörnberg, Franz Worek, Anna Linusson e Fredrik J. Ekström. "Structure of a prereaction complex between the nerve agent sarin, its biological target acetylcholinesterase, and the antidote HI-6". Proceedings of the National Academy of Sciences 113, n. 20 (2 maggio 2016): 5514–19. http://dx.doi.org/10.1073/pnas.1523362113.
Abstract (sommario):
Organophosphorus nerve agents interfere with cholinergic signaling by covalently binding to the active site of the enzyme acetylcholinesterase (AChE). This inhibition causes an accumulation of the neurotransmitter acetylcholine, potentially leading to overstimulation of the nervous system and death. Current treatments include the use of antidotes that promote the release of functional AChE by an unknown reactivation mechanism. We have used diffusion trap cryocrystallography and density functional theory (DFT) calculations to determine and analyze prereaction conformers of the nerve agent antidote HI-6 in complex with Mus musculus AChE covalently inhibited by the nerve agent sarin. These analyses reveal previously unknown conformations of the system and suggest that the cleavage of the covalent enzyme–sarin bond is preceded by a conformational change in the sarin adduct itself. Together with data from the reactivation kinetics, this alternate conformation suggests a key interaction between Glu202 and the O-isopropyl moiety of sarin. Moreover, solvent kinetic isotope effect experiments using deuterium oxide reveal that the reactivation mechanism features an isotope-sensitive step. These findings provide insights into the reactivation mechanism and provide a starting point for the development of improved antidotes. The work also illustrates how DFT calculations can guide the interpretation, analysis, and validation of crystallographic data for challenging reactive systems with complex conformational dynamics.
44
Lee, Chang-Uk, e Tom N. Grossmann. "Reversible Covalent Inhibition of a Protein Target". Angewandte Chemie International Edition 51, n. 35 (13 luglio 2012): 8699–700. http://dx.doi.org/10.1002/anie.201203341.
45
Ahmed, Vanessa F., Nunzio Bottini e Amy M. Barrios. "Covalent Inhibition of the Lymphoid Tyrosine Phosphatase". ChemMedChem 9, n. 2 (8 gennaio 2014): 296–99. http://dx.doi.org/10.1002/cmdc.201300404.
46
Stevic, Ivan, Howard H. W. Chan, Ankush Chander, Leslie R. Berry e Anthony K. C. Chan. "Inhibition of Platelet-Prothrombianse by a Covalent Antithrombin-Heparin Complex." Blood 120, n. 21 (16 novembre 2012): 2212. http://dx.doi.org/10.1182/blood.v120.21.2212.2212.
Abstract (sommario):
Abstract Abstract 2212 Introduction: Factor Xa is protected within the prothrombinase complex from inhibition by heparin. We have developed a covalent antithrombin-heparin complex (ATH) with enhanced anticoagulant activity. Previously, we have shown that ATH is able to inhibit coagulation enzymes much more efficiently than regular antithrombin+heparin (AT+UFH). For example, ATH inhibited TF/VIIa ∼30-fold faster compared to AT+UFH. Furthermore, we have also demonstrated that ATH is capable of inhibiting Xa within a prothrombinase complex assembled on synthetic phospholipid vesicles better than AT+UFH. However, ATH's effect on prothrombinase when the complex is formed on a more native system such as platelets has never been explored. Thus, the objective of the present study is to determine the ability of ATH vs AT+UFH to inhibit Xa within the prothrombinase complex when the enzyme complex is assembled on the platelet system. Methods: Discontinuous second order rate constant assays were performed to obtain k2-values for inhibition of free or prothrombinase-bound Xa by AT+UFH or ATH. Freshly prepared resting platelets were subjected to inhibition analysis by first incubating them with Xa, Va, Ca2+ and pefabloc®-TH (thrombin inhibitor) in different wells of a 96-well plate for 3 min at 37 °C. Prothrombin was then simultaneously added to all wells to initiate thrombin generation, followed by addition of AT+UFH or ATH inhibitors to each well at specific time intervals. Reactions were neutralized by simultaneous addition of polybrene, Na2EDTA and Xa-specific substrate S-2222™ in buffer. The remaining Xa enzyme activity was obtained and final k2-values calculated. For experiments requiring activated platelets, freshly isolated platelets were activated with 5 μM calcium ionophore A23187 + 4 mM CaCl2 for 15 min at room temperature. The activated platelets were then tested in inhibition assays as described above. To investigate the roles of individual components of the prothrombinase complex on the anticoagulant effects of AT+UFH and ATH, additional experiments were performed where components of the complex (prothrombin, activated platelets or Va) were omitted prior to reaction with inhibitors. Thrombin generation was used to assess functionality of the activated platelet-prothrombinase system in the presence of inhibitors using a thrombin-specific substrate S-2238™. Results: The k2-values (×108M−1min−1) for inhibition of free Xa or resting platelet-prothrombinase were similar for both inhibitors, although the overall inhibition rates achieved by the ATH were 2-fold faster than AT+UFH (p<0.001). Since activated platelets are required for enhanced prothrombinase function, we then compared inhibition of free vs activated platelet-prothrombinase by the two inhibitors (platelet activation was confirmed with flow cytometry using an anti CD-41 antibody). No differences were observed in the k2-values between free Xa (3.96±0.23) and activated platelet prothrombinase (3.83±0.39) for ATH reactions. However, the k2-values for inhibition of free Xa by AT+UFH was 2.37±0.32, and assembly of Xa within the activated platelet-prothrombinase resulted in a reduction in the k2-values to 0.99±0.22 (p<0.001), thus confirming a moderate 60% protection of Xa by the prothrombinase components. However, omitting the components (prothrombin, activated platelets or Va) from the complex resulted in higher k2-values (1.76±0.37, 2.29±0.26 and 2.52±0.32, respectively p<0.01) for AT+UFH, and as expected, no net effect was observed for ATH. Thrombin generation was inhibited significantly by both AT+UFH and ATH compared to the control (p<0.001), but further analysis of thrombin potential yielded greater inhibition by ATH compared to AT+UFH (p<0.05). Conclusion: In this study, we report inhibition of the prothrombinase complex on the surface of resting and activated platelets. Consistent with previous investigations, a moderate protection of Xa was observed when the activated platelet-prothrombinase was inhibited by AT+UFH. ATH on the other hand, targets and inhibits prothrombinase complexed-Xa as fast as free Xa, and at inhibition rates that were significantly faster than AT+UFH. Thus, overall the covalent conjugate enhances anticoagulation of surface-bound enzymes and offers advantages over conventional heparin for the treatment of cell-based coagulation in vivo. Disclosures: No relevant conflicts of interest to declare.
47
Zhou, Yuxin, Ji Tao, Dingshuainan Jin, Shiping Zhang, Yan He e Longlong Niu. "The Inhibition Effect and Mechnism of a Thiadiazole Derivative on Q235 Carbon Steel in 1 M HCl Solution". Applied Sciences 13, n. 4 (6 febbraio 2023): 2103. http://dx.doi.org/10.3390/app13042103.
Abstract (sommario):
N,N-dihydroxyethyl-(5-methyl-[1,3,4] thiadiazol-2-sulfur)-carbonyl acetamide was synthesized and used as an inhibitor to protect Q235 carbon steel in a 1 M HCl solution. The results showed an increased inhibition efficiency with the increase in the concentration of this inhibitor, and an inhibition efficiency higher than 96% at 40 mg/L can be obtained from weight loss, electrochemical impedance spectroscopy, and potentiodynamic polarization results. The inhibition effect was determined by the adsorption film according to the surface morphology and elemental distribution of the carbon steel surface. The adsorption consists of physical adsorption and chemical adsorption in view of the free adsorption energy of −40.64 kJ/mol derived from the Langmuir adsorption isotherm line. The lone pair electrons of N, O and S and π electronics of double bonds in molecules form stable covalent coordination bonds with the empty d orbitals of iron atoms, which is beneficial to chemical adsorption of the inhibitor. The high inhibiton efficiency of this inhibitor is important for the potential application in pickling field.
48
Buneeva, O. A., L. N. Aksenova e A. E. Medvedev. "A Simple Approach for Pilot Analysis of Time-dependent Enzyme Inhibition: Discrimination Between Mechanism-based Inactivation and Tight Binding Inhibitor Behavior". Biomedical Chemistry: Research and Methods 3, n. 1 (2020): e00115. http://dx.doi.org/10.18097/bmcrm00115.
Abstract (sommario):
The increase in enzyme inhibition developed during prolonged incubation of an enzyme preparation with a chemical substance may be associated with both the non-covalent and also with covalent enzyme-inhibitor complex formation. The latter case involves catalytic conversion of a mechanism-based irreversible inhibitor (a poor substrate) into a reactive species forming covalent adduct(s) with the enzyme and thus irreversibly inactivating the enzyme molecule. Using a simple approach, based on comparison of enzyme inhibition after preincubation with a potential inhibitor at 4ºC or 37ºC we have analyzed inhibition of monoamine oxidase A (MAO A) by known MAO inhibitors pargyline and pirlindole (pyrazidol). MAO A inhibitory activity of pirlindole (reversible tight binding inhibitor of MAO A) assayed after mitochondrial wash was basically the same for the incubation at both 4ºC and 37ºC. In contrast to pirlindole, the effect of pargyline (mechanism based irreversible MAO inhibitor) strongly depended on the temperature of the incubation medium. At 37ºC the residual activity MAO A in the mitochondrial fraction after washing was significantly lower than in the mitochondrial samples incubated with pargyline at 4ºC. Results of this study suggest that using analysis of both time- and temperature-dependence of inhibition it is possible to discriminate mechanism-based irreversible inhibition and reversible tight binding inhibition of target enzym
49
Wang, Qian, Guofeng Chen, Jian He, Jiameng Li, Muya Xiong, Haixia Su, Minjun Li, Hangchen Hu e Yechun Xu. "Structure-Based Design of Potent Peptidomimetic Inhibitors Covalently Targeting SARS-CoV-2 Papain-like Protease". International Journal of Molecular Sciences 24, n. 10 (11 maggio 2023): 8633. http://dx.doi.org/10.3390/ijms24108633.
Abstract (sommario):
The papain-like protease (PLpro) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) plays a critical role in the proteolytic processing of viral polyproteins and the dysregulation of the host immune response, providing a promising therapeutic target. Here, we report the structure-guide design of novel peptidomimetic inhibitors covalently targeting SARS-CoV-2 PLpro. The resulting inhibitors demonstrate submicromolar potency in the enzymatic assay (IC50 = 0.23 μM) and significant inhibition of SARS-CoV-2 PLpro in the HEK293T cells using a cell-based protease assay (EC50 = 3.61 μM). Moreover, an X-ray crystal structure of SARS-CoV-2 PLpro in complex with compound 2 confirms the covalent binding of the inhibitor to the catalytic residue cysteine 111 (C111) and emphasizes the importance of interactions with tyrosine 268 (Y268). Together, our findings reveal a new scaffold of SARS-CoV-2 PLpro inhibitors and provide an attractive starting point for further optimization.
50
Abdeldayem, Ayah, Yasir S. Raouf, Stefan N. Constantinescu, Richard Moriggl e Patrick T. Gunning. "Advances in covalent kinase inhibitors". Chemical Society Reviews 49, n. 9 (2020): 2617–87. http://dx.doi.org/10.1039/c9cs00720b.