A collaborative, open-science team undertook discovery of novel small molecule inhibitors of the SARS-CoV-2 nsp16-nsp10 2'-O-methyltransferase using a high throughput screening approach with the potential to reveal new inhibition strategies. This screen yielded compound 5a, a ligand possessing an electron-deficient double bond, as an inhibitor of SARS-CoV-2 nsp16 activity. Surprisingly, X-ray crystal structures revealed that 5a covalently binds within a previously unrecognized cryptic pocket near the S-adenosylmethionine binding cleft in a manner that prevents occupation by S-adenosylmethionine. Using a multidisciplinary approach, we examined the mechanism of binding of compound 5a to the nsp16 cryptic pocket and developed 5a derivatives that inhibited nsp16 activity and murine hepatitis virus replication in rat lung epithelial cells but proved cytotoxic to cell lines canonically used to examine SARS-CoV-2 infection. Our study reveals the druggability of this newly discovered SARS-CoV-2 nsp16 cryptic pocket, provides novel tool compounds to explore the site, and suggests a new approach for discovery of nsp16 inhibition-based pan-coronavirus therapeutics through structure-guided drug design.

• MeSH
• COVID-19 * MeSH
• krysa rodu rattus MeSH
• methyltransferasy MeSH
• myši MeSH
• S-adenosylmethionin chemie   metabolismus   MeSH
• SARS-CoV-2 * metabolismus   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

The metabolism of sulfur-containing amino acids (SAAs) requires an orchestrated interplay among several dozen enzymes and transporters, and an adequate dietary intake of methionine (Met), cysteine (Cys), and B vitamins. Known human genetic disorders are due to defects in Met demethylation, homocysteine (Hcy) remethylation, or cobalamin and folate metabolism, in Hcy transsulfuration, and Cys and hydrogen sulfide (H2S) catabolism. These disorders may manifest between the newborn period and late adulthood by a combination of neuropsychiatric abnormalities, thromboembolism, megaloblastic anemia, hepatopathy, myopathy, and bone and connective tissue abnormalities. Biochemical features include metabolite deficiencies (e.g. Met, S-adenosylmethionine (AdoMet), intermediates in 1-carbon metabolism, Cys, or glutathione) and/or their accumulation (e.g. S-adenosylhomocysteine, Hcy, H2S, or sulfite). Treatment should be started as early as possible and may include a low-protein/low-Met diet with Cys-enriched amino acid supplements, pharmacological doses of B vitamins, betaine to stimulate Hcy remethylation, the provision of N-acetylcysteine or AdoMet, or experimental approaches such as liver transplantation or enzyme replacement therapy. In several disorders, patients are exposed to long-term markedly elevated Met concentrations. Although these conditions may inform on Met toxicity, interpretation is difficult due to the presence of additional metabolic changes. Two disorders seem to exhibit Met-associated toxicity in the brain. An increased risk of demyelination in patients with Met adenosyltransferase I/III (MATI/III) deficiency due to biallelic mutations in the MATIA gene has been attributed to very high blood Met concentrations (typically >800 μmol/L) and possibly also to decreased liver AdoMet synthesis. An excessively high Met concentration in some patients with cystathionine β-synthase deficiency has been associated with encephalopathy and brain edema, and direct toxicity of Met has been postulated. In summary, studies in patients with various disorders of SAA metabolism showed complex metabolic changes with distant cellular consequences, most of which are not attributable to direct Met toxicity.

• MeSH
• aminokyseliny sírové metabolismus   MeSH
• cystein metabolismus   MeSH
• glutathion metabolismus   MeSH
• homocystein metabolismus   MeSH
• homocystinurie etiologie   metabolismus   MeSH
• játra metabolismus   MeSH
• lidé MeSH
• metabolické nemoci genetika   metabolismus   patologie   terapie   MeSH
• methionin metabolismus   MeSH
• methioninadenosyltransferasa metabolismus   MeSH
• metylace MeSH
• nemoci mozku etiologie   metabolismus   MeSH
• S-adenosylmethionin metabolismus   MeSH
• síra metabolismus   MeSH
• siřičitany metabolismus   MeSH
• sloučeniny síry metabolismus   MeSH
• sulfan metabolismus   MeSH
• vrozené poruchy metabolismu patologie   terapie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

The anti-diabetic biguanide metformin may exert health-promoting effects via metabolic regulation of the epigenome. Here we show that metformin promotes global DNA methylation in non-cancerous, cancer-prone and metastatic cancer cells by decreasing S-adenosylhomocysteine (SAH), a strong feedback inhibitor of S-adenosylmethionine (SAM)-dependent DNA methyltransferases, while promoting the accumulation of SAM, the universal methyl donor for cellular methylation. Using metformin and a mitochondria/complex I (mCI)-targeted analog of metformin (norMitoMet) in experimental pairs of wild-type and AMP-activated protein kinase (AMPK)-, serine hydroxymethyltransferase 2 (SHMT2)- and mCI-null cells, we provide evidence that metformin increases the SAM:SAH ratio-related methylation capacity by targeting the coupling between serine mitochondrial one-carbon flux and CI activity. By increasing the contribution of one-carbon units to the SAM from folate stores while decreasing SAH in response to AMPK-sensed energetic crisis, metformin can operate as a metabolo-epigenetic regulator capable of reprogramming one of the key conduits linking cellular metabolism to the DNA methylation machinery.

• MeSH
• genom lidský * MeSH
• hypoglykemika farmakologie   MeSH
• lidé MeSH
• metformin farmakologie   MeSH
• metylace DNA účinky léků   MeSH
• mitochondrie účinky léků   metabolismus   patologie   MeSH
• myši MeSH
• nádorové biomarkery MeSH
• nádorové buňky kultivované MeSH
• nádory prsu farmakoterapie   enzymologie   patologie   MeSH
• nádory tračníku farmakoterapie   enzymologie   patologie   MeSH
• následné studie MeSH
• proteinkinasy aktivované AMP metabolismus   MeSH
• regulace genové exprese u nádorů účinky léků   MeSH
• respirační komplex I metabolismus   MeSH
• S-adenosylhomocystein metabolismus   MeSH
• S-adenosylmethionin metabolismus   MeSH
• uhlík metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

ALKB-8 is a 2-oxoglutarate-dependent dioxygenase homologous to bacterial AlkB, which oxidatively demethylates DNA substrates. The mammalian AlkB family contains AlkB homologues denominated ALKBH1 to 8 and FTO. The C. elegans genome includes five AlkB-related genes, homologues of ALKBH1, 4, 6, 7, and 8, but lacks homologues of ALKBH2, 3, and 5 and FTO. ALKBH8 orthologues differ from other AlkB family members by possessing an additional methyltransferase module and an RNA binding N-terminal module. The ALKBH8 methyltransferase domain generates the wobble nucleoside 5-methoxycarbonylmethyluridine from its precursor 5-carboxymethyluridine and its (R)- and (S)-5-methoxycarbonylhydroxymethyluridine hydroxylated forms in tRNA Arg/UCG and tRNA Gly/UCC. The ALKBH8/ALKB-8 methyltransferase domain is highly similar to yeast TRM9, which selectively modulates translation of mRNAs enriched with AGA and GAA codons under both normal and stress conditions. In this report, we studied the role of alkb-8 in C. elegans. We show that downregulation of alkb-8 increases detection of lysosome-related organelles visualized by Nile red in vivo. Reversely, forced expression of alkb-8 strongly decreases the detection of this compartment. In addition, overexpression of alkb-8 applied in a pulse during the L1 larval stage increases the C. elegans lifespan.

• MeSH
• Caenorhabditis elegans embryologie   enzymologie   genetika   MeSH
• dioxygenasy metabolismus   MeSH
• dlouhověkost MeSH
• down regulace genetika   MeSH
• embryo nesavčí metabolismus   MeSH
• geneticky modifikovaná zvířata MeSH
• kyseliny ketoglutarové metabolismus   MeSH
• larva metabolismus   MeSH
• lyzozomy metabolismus   MeSH
• methyltransferasy metabolismus   MeSH
• operon MeSH
• promotorové oblasti (genetika) MeSH
• proteiny Caenorhabditis elegans genetika   metabolismus   MeSH
• RNA interference MeSH
• S-adenosylmethionin metabolismus   MeSH
• stárnutí metabolismus   MeSH
• vývojová regulace genové exprese MeSH
• zelené fluorescenční proteiny metabolismus   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

• Klíčová slova
• Silybin, ALAVIS SAMMY,
• MeSH
• antioxidancia terapeutické užití   MeSH
• demence * farmakoterapie   MeSH
• kočky MeSH
• kognitivní stárnutí MeSH
• nemoci jater farmakoterapie   veterinární   MeSH
• ostropestřec mariánský MeSH
• psi MeSH
• S-adenosylmethionin * metabolismus   terapeutické užití   MeSH
• zvířata MeSH
• Check Tag
• kočky MeSH
• psi MeSH
• zvířata MeSH

p-Cresol and indole are volatile biologically active products of the bacterial degradation of tyrosine and tryptophan respectively. They are typically produced by bacteria in animal intestines, soil and various sediments. Here, we demonstrate that the free-living eukaryote Mastigamoeba balamuthi and its pathogenic relative Entamoeba histolytica produce significant amounts of indole via tryptophanase activity. Unexpectedly, M. balamuthi also produces p-cresol in concentrations that are bacteriostatic to non-p-cresol-producing bacteria. The ability of M. balamuthi to produce p-cresol, which has not previously been observed in any eukaryotic microbe, was gained due to the lateral acquisition of a bacterial gene for 4-hydroxyphenylacetate decarboxylase (HPAD). In bacteria, the genes for HPAD and the S-adenosylmethionine-dependent activating enzyme (AE) are present in a common operon. In M. balamuthi, HPAD displays a unique fusion with the AE that suggests the operon-mediated transfer of genes from a bacterial donor. We also clarified that the tyrosine-to-4-hydroxyphenylacetate conversion proceeds via the Ehrlich pathway. The acquisition of the bacterial HPAD gene may provide M. balamuthi a competitive advantage over other microflora in its native habitat.

• MeSH
• Archamoebae genetika   MeSH
• Bacteria genetika   MeSH
• bakteriální geny * MeSH
• indoly metabolismus   MeSH
• karboxylyasy MeSH
• kresoly metabolismus   MeSH
• operon MeSH
• přenos genů horizontální * MeSH
• S-adenosylmethionin metabolismus   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Both cardiovascular disease and liver injury are major public health issues. Hyperhomocysteinemia has been linked to cardiovascular diseases, and defects in methyl group metabolism, often resulting in hyperhomocysteinemia, are among the key molecular events postulated to play a role in liver injury. We employed proteomics and metabolomics analyses of human hepatocytes in primary cell culture to explore the spectrum of proteins and associated metabolites affected by the disruption of methyl group metabolism. We treated the hepatocytes with homocysteine (Hcy, 0.1mM and 2mM) to follow the impact of hyperhomocysteinemia, and in parallel, we used a specific inhibitor of betaine-homocysteine S-methyltransferase (BHMT) to extend our understanding of the physiological functions of the enzyme. The major effect of BHMT inhibition was a 50% decrease in S-adenosylmethionine levels. The treatments with Hcy resulted in multiple changes in the metabolite levels depending on the treatment modality. The BHMT inhibition and 0.1mM Hcy treatment induced only moderate changes in the hepatocyte proteome and secretome, while the changes induced by the 2mM Hcy treatment were extensive. Phosphatidylethanolamine carboxykinase and ornithine aminotransferase were up-regulated about two fold indicating an intervention into metabolism. Cellular proliferation was suspended, secretome composition was changed and signs of apoptosis were discernible. We have detected fibrinogen gamma dimers, which might have a role as a potentially new biomarker of early liver injury. Finally, we have demonstrated the failed maturation of apolipoprotein A1, which might be a new mechanism of disruption of cholesterol efflux from tissues.

• MeSH
• 2D gelová elektroforéza MeSH
• apolipoprotein A-I metabolismus   MeSH
• apoptóza MeSH
• betain-homocystein-S-methyltransferasa antagonisté a inhibitory   metabolismus   MeSH
• fibrinogen metabolismus   MeSH
• hepatocyty účinky léků   metabolismus   MeSH
• homocystein farmakologie   MeSH
• hyperhomocysteinemie metabolismus   MeSH
• kolorektální nádory farmakoterapie   metabolismus   patologie   MeSH
• kultivované buňky MeSH
• lidé středního věku MeSH
• lidé MeSH
• metabolomika * MeSH
• multimerizace proteinu MeSH
• nádory jater farmakoterapie   metabolismus   patologie   MeSH
• proliferace buněk MeSH
• proteom analýza   metabolismus   MeSH
• S-adenosylmethionin metabolismus   MeSH
• spektrometrie hmotnostní - ionizace laserem za účasti matrice MeSH
• western blotting MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH

DNA methylation inhibitors are being extensively studied as potential anticancer agents. In the present study, we compared the capability of alpha anomer of 5-aza-2'-deoxycytidine (alpha-5-azadCyd) to induce down-regulation of hTERT expression in HL-60 cells with other nucleoside analogs that act as DNA methylation inhibitors: beta-5-azadCyd (decitabine), (S)-9-(2,3-dihydroxypropyl)adenine [(S)-DHPA], isobutyl ester of (R,S)-3-(adenin-9-yl)-2-hydroxypropanoic acid [(R,S)-AHPA-ibu] and prospective DNA methylation inhibitors (S)-1-[3-hydroxy-2-(phosphonomethoxy)propyl]-5-azacytosine [(S)-HPMPazaC] and 5-fluoro-zebularine (F-PymRf). Exposure to alpha-5-azadCyd induced the down-regulation of hTERT expression in low micromolar concentrations (0.05-50 microM). A more cytotoxic beta anomer caused a transient up-regulation of hTERT and a subsequent reduction in hTERT mRNA levels at concentrations more than 10 times below its GIC50 value. In this respect, (S)-DHPA and (R,S)-AHPA-ibu were less efficient, since a similar effect was achieved at concentrations above their GIC(50). In contrast, F-PymRf treatment resulted in up to a three-fold induction of hTERT expression within a broad range of concentrations. In all cases, the down-regulation of hTERT expression was concentration-dependent. The correlation was found between c-myc overexpression and transiently elevated hTERT expression after treatment with all tested compounds except for alpha-5-azadCyd and (S)-HPMPazaC. Although the primary task of hypomethylating agents in anticancer therapy lies in reactivation of silenced tumour-suppressor genes, the inhibition of hTERT expression might also be a fruitful clinical effect of this approach.

• MeSH
• antimetabolity antitumorózní farmakologie   MeSH
• azacytidin analogy a deriváty   farmakologie   chemie   MeSH
• DNA metabolismus   MeSH
• down regulace MeSH
• financování organizované MeSH
• HL-60 buňky MeSH
• lidé MeSH
• messenger RNA biosyntéza   MeSH
• metylace DNA účinky léků   MeSH
• polymerázová řetězová reakce s reverzní transkripcí MeSH
• S-adenosylhomocystein metabolismus   MeSH
• S-adenosylmethionin metabolismus   MeSH
• stereoizomerie MeSH
• telomerasa biosyntéza   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• Check Tag
• lidé MeSH

S-adenosylmethionin (SAMe) je látka produkovaná v cytosolu prakticky všech lidských buněk, hlavním místem syntézy a degradace SAMe jsou však játra. SAMe se podílí na řadě reakcí, které vedou mimo jiné k udržování fluidity membrán a tvorbě významného buněčného antioxidantu glutathionu. Jaterní onemocnění jsou velmi často provázena poklesem SAMe v buňkách, který má řadu příčin. Podávání SAMe pak může různými mechanizmy působit preventivně proti poškození jaterních buněk a rozvoji jaterního onemocnění.

Although S-adenosylmethionine (SAMe) is a substance produced in cytosol in almost all human cells, SAMe is mainly synthesized in the liver. SAMe plays a role in numerous metabolic reactions including synthesis of glutathione, the main cellular antioxidant, and methylations whereby membrane stability is restored. Liver diseases are frequently accompanied with decrease in SAMe content in the cells resulting from a variety of causes. Then treatment with SAMe can prevent hepatocyte injury and progression of liver disease by different mechanisms.

• MeSH
• glutathion biosyntéza   metabolismus   účinky léků   MeSH
• lidé MeSH
• methioninadenosyltransferasa metabolismus   účinky léků   MeSH
• mitochondrie metabolismus   účinky léků   MeSH
• nemoci jater farmakoterapie   prevence a kontrola   terapie   MeSH
• primární prevence metody   MeSH
• S-adenosylmethionin metabolismus   sekrece   terapeutické užití   MeSH
• Check Tag
• lidé MeSH

• MeSH
• Alzheimerova nemoc komplikace   metabolismus   MeSH
• cerebrovaskulární poruchy komplikace   metabolismus   MeSH
• cévní mozková příhoda komplikace   metabolismus   MeSH
• hyperhomocysteinemie * komplikace   krev   metabolismus   MeSH
• kyselina listová izolace a purifikace   MeSH
• lidé MeSH
• metaanalýza jako téma MeSH
• N-methylaspartát izolace a purifikace   krev   metabolismus   MeSH
• nemoci nervového systému * klasifikace   metabolismus   MeSH
• neurokognitivní poruchy etiologie   metabolismus   MeSH
• S-adenosylmethionin izolace a purifikace   metabolismus   MeSH
• schizofrenie komplikace   metabolismus   MeSH
• stárnutí * fyziologie   metabolismus   patologie   MeSH
• statistika jako téma MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• přehledy MeSH