Protein kinase CK2, a heterotetrameric holoenzyme composed of two catalytic chains (CK2α) attache... more Protein kinase CK2, a heterotetrameric holoenzyme composed of two catalytic chains (CK2α) attached to a homodimer of regulatory subunits (CK2β), is a target for drug development for cancer therapy. Here, we describe the tetraiodobenzimidazole derivative ARC-3140, a bisubstrate inhibitor addressing the ATP site and the substrate-binding site of CK2 with extraordinary affinity (Ki = 84 pM). In a crystal structure of ARC-3140 in complex with CK2α, three copies of the inhibitor are visible, one of them at the CK2β interface of CK2α. Subsequent interaction studies based on microscale thermophoresis and fluorescence anisotropy changes revealed a significant impact of ARC-3140 and of its tetrabromo equivalent ARC-1502 on the CK2α/CK2β interaction. A structural inspection revealed that ARC-3140, unlike CK2β antagonists described so far, interferes with both sub-interfaces of the bipartite CK2α/CK2β interaction. Thus, ARC-3140 is a lead for the further development of highly effective compounds perturbating the quaternary structure of the CK2α2β2 holoenzyme.
ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was e... more ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.
Alkylation of adenine in solution and on solid phase was accelerated by phosphazene base P1-tBu c... more Alkylation of adenine in solution and on solid phase was accelerated by phosphazene base P1-tBu compared to mineral bases. The reactions in solution afforded regioselectively the appropriate N9-alkylated adenines with high preparative yields while the reaction with polystyrene resin-bound N-bromoacetylated peptides gave three regioisomers (alkylated at the N9, N7, and N3 position of adenine) in a 4:2:1 molar ratio. Ten novel nonphosphate nucleotide analogues were tested in an ADP-induced platelet aggregation assay.
The conjugates of an adenosine mimetic and oligo‐l‐arginine or oligo‐d‐arginine (ARCs) were initi... more The conjugates of an adenosine mimetic and oligo‐l‐arginine or oligo‐d‐arginine (ARCs) were initially designed in our research group as inhibitors and photoluminescent probes targeting basophilic protein kinases. Here, we explored a panel of ARCs and their fluorescent derivatives in biochemical assays with members of the protein arginine methyltransferase (PRMT) family, focusing specifically on PRMT1. In the binding/displacement assay with detection of fluorescence anisotropy, we found that ARCs and arginine‐rich peptides could serve as high‐affinity ligands for PRMT1, whereas the equilibrium dissociation constant values depended dramatically on the number of arginine residues within the compounds. The fluorescently labeled probe ARC‐1081 was displaced from its complex with PRMT1 by both S‐adenosyl‐l‐methionine (SAM) and S‐adenosyl‐l‐homocysteine (SAH), indicating binding of the adenosine mimetic of ARCs to the SAM/SAH‐binding site within PRMT1. The ARCs that had previously shown microsecond‐lifetime photoluminescence in complex with protein kinases did not feature such property in complex with PRMT1, demonstrating the selectivity of the time‐resolved readout format. When tested against a panel of PRMT family members in single‐dose inhibition experiments, a micromolar concentration of ARC‐902 was required for the inhibition of PRMT1 and PRMT7. Overall, our results suggest that the compounds containing multiple arginine residues (including the well‐known cell‐penetrating peptides) are likely to inhibit PRMT and thus interfere with the epigenetic modification status in complex biological systems, which should be taken into consideration during interpretation of the experimental data.
ABSTRACT No Gi-linked P2Y receptors have been cloned to date but the presence of such receptors i... more ABSTRACT No Gi-linked P2Y receptors have been cloned to date but the presence of such receptors is thought to be restricted to platelets and certain clonal cell lines. Using the functional approach of [35S]guanosine 5′-[γ-thio]-triphosphate autoradiography, we uncovered the widespread presence of such receptors in the CNS. Under conditions in which the prominent signal due to tonic adenosine receptor activity is masked, ADP and ATP stimulated G-protein activity in multiple grey and white matter regions. Localization in the grey matter suggests inhibitory auto-/heteroreceptor function. In the white matter, activated G proteins appeared as ‘hot spots’ (presumed oligodendrocyte progenitors) with scattered distribution along the main fibre tracts. Responses to ATP were diminished under conditions that inhibited degradation, suggesting that prior conversion to ADP explained agonist action. Uracil nucleotides were ineffective but 2-methylthio-ADP activated G proteins ≈ 500-fold more potently than ADP, although both were similarly degraded. Throughout the brain, ADP-dependent G-protein activity was reversed by 2-hexylthio-AdoOC(O)Asp2, a non-phosphate ATP analogue, whereas selective P2Y1 receptor antagonists proved ineffective. A similar receptor was also disclosed from the adrenal medulla. These data witness a hitherto unrecognized abundance of Gi/o-linked ADP receptors in the nervous system. Biochemical and pharmacological behaviour suggests striking similarities to the elusive platelet P2YADP receptor.
Protein kinase CK2, a heterotetrameric holoenzyme composed of two catalytic chains (CK2α) attache... more Protein kinase CK2, a heterotetrameric holoenzyme composed of two catalytic chains (CK2α) attached to a homodimer of regulatory subunits (CK2β), is a target for drug development for cancer therapy. Here, we describe the tetraiodobenzimidazole derivative ARC-3140, a bisubstrate inhibitor addressing the ATP site and the substrate-binding site of CK2 with extraordinary affinity (Ki = 84 pM). In a crystal structure of ARC-3140 in complex with CK2α, three copies of the inhibitor are visible, one of them at the CK2β interface of CK2α. Subsequent interaction studies based on microscale thermophoresis and fluorescence anisotropy changes revealed a significant impact of ARC-3140 and of its tetrabromo equivalent ARC-1502 on the CK2α/CK2β interaction. A structural inspection revealed that ARC-3140, unlike CK2β antagonists described so far, interferes with both sub-interfaces of the bipartite CK2α/CK2β interaction. Thus, ARC-3140 is a lead for the further development of highly effective compounds perturbating the quaternary structure of the CK2α2β2 holoenzyme.
ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was e... more ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.
Alkylation of adenine in solution and on solid phase was accelerated by phosphazene base P1-tBu c... more Alkylation of adenine in solution and on solid phase was accelerated by phosphazene base P1-tBu compared to mineral bases. The reactions in solution afforded regioselectively the appropriate N9-alkylated adenines with high preparative yields while the reaction with polystyrene resin-bound N-bromoacetylated peptides gave three regioisomers (alkylated at the N9, N7, and N3 position of adenine) in a 4:2:1 molar ratio. Ten novel nonphosphate nucleotide analogues were tested in an ADP-induced platelet aggregation assay.
The conjugates of an adenosine mimetic and oligo‐l‐arginine or oligo‐d‐arginine (ARCs) were initi... more The conjugates of an adenosine mimetic and oligo‐l‐arginine or oligo‐d‐arginine (ARCs) were initially designed in our research group as inhibitors and photoluminescent probes targeting basophilic protein kinases. Here, we explored a panel of ARCs and their fluorescent derivatives in biochemical assays with members of the protein arginine methyltransferase (PRMT) family, focusing specifically on PRMT1. In the binding/displacement assay with detection of fluorescence anisotropy, we found that ARCs and arginine‐rich peptides could serve as high‐affinity ligands for PRMT1, whereas the equilibrium dissociation constant values depended dramatically on the number of arginine residues within the compounds. The fluorescently labeled probe ARC‐1081 was displaced from its complex with PRMT1 by both S‐adenosyl‐l‐methionine (SAM) and S‐adenosyl‐l‐homocysteine (SAH), indicating binding of the adenosine mimetic of ARCs to the SAM/SAH‐binding site within PRMT1. The ARCs that had previously shown microsecond‐lifetime photoluminescence in complex with protein kinases did not feature such property in complex with PRMT1, demonstrating the selectivity of the time‐resolved readout format. When tested against a panel of PRMT family members in single‐dose inhibition experiments, a micromolar concentration of ARC‐902 was required for the inhibition of PRMT1 and PRMT7. Overall, our results suggest that the compounds containing multiple arginine residues (including the well‐known cell‐penetrating peptides) are likely to inhibit PRMT and thus interfere with the epigenetic modification status in complex biological systems, which should be taken into consideration during interpretation of the experimental data.
ABSTRACT No Gi-linked P2Y receptors have been cloned to date but the presence of such receptors i... more ABSTRACT No Gi-linked P2Y receptors have been cloned to date but the presence of such receptors is thought to be restricted to platelets and certain clonal cell lines. Using the functional approach of [35S]guanosine 5′-[γ-thio]-triphosphate autoradiography, we uncovered the widespread presence of such receptors in the CNS. Under conditions in which the prominent signal due to tonic adenosine receptor activity is masked, ADP and ATP stimulated G-protein activity in multiple grey and white matter regions. Localization in the grey matter suggests inhibitory auto-/heteroreceptor function. In the white matter, activated G proteins appeared as ‘hot spots’ (presumed oligodendrocyte progenitors) with scattered distribution along the main fibre tracts. Responses to ATP were diminished under conditions that inhibited degradation, suggesting that prior conversion to ADP explained agonist action. Uracil nucleotides were ineffective but 2-methylthio-ADP activated G proteins ≈ 500-fold more potently than ADP, although both were similarly degraded. Throughout the brain, ADP-dependent G-protein activity was reversed by 2-hexylthio-AdoOC(O)Asp2, a non-phosphate ATP analogue, whereas selective P2Y1 receptor antagonists proved ineffective. A similar receptor was also disclosed from the adrenal medulla. These data witness a hitherto unrecognized abundance of Gi/o-linked ADP receptors in the nervous system. Biochemical and pharmacological behaviour suggests striking similarities to the elusive platelet P2YADP receptor.
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