The 3'-5', 3'-5' cyclic dinucleotides (3'3'CDNs) are bacterial second messengers that can also bind to the stimulator of interferon genes (STING) adaptor protein in vertebrates and activate the host innate immunity. Here, we profiled the substrate specificity of four bacterial dinucleotide synthases from Vibrio cholerae (DncV), Bacillus thuringiensis (btDisA), Escherichia coli (dgcZ), and Thermotoga maritima (tDGC) using a library of 33 nucleoside-5'-triphosphate analogues and then employed these enzymes to synthesize 24 3'3'CDNs. The STING affinity of CDNs was evaluated in cell-based and biochemical assays, and their ability to induce cytokines was determined by employing human peripheral blood mononuclear cells. Interestingly, the prepared heterodimeric 3'3'CDNs bound to the STING much better than their homodimeric counterparts and showed similar or better potency than bacterial 3'3'CDNs. We also rationalized the experimental findings by in-depth STING-CDN structure-activity correlations by dissecting computed interaction free energies into a set of well-defined and intuitive terms. To this aim, we employed state-of-the-art methods of computational chemistry, such as quantum mechanics/molecular mechanics (QM/MM) calculations, and complemented the computed results with the {STING:3'3'c-di-ara-AMP} X-ray crystallographic structure. QM/MM identified three outliers (mostly homodimers) for which we have no clear explanation of their impaired binding with respect to their heterodimeric counterparts, whereas the R2 = 0.7 correlation between the computed ΔG'int_rel and experimental ΔTm's for the remaining ligands has been very encouraging.
- MeSH
- Bacillus thuringiensis enzymologie ultrastruktura MeSH
- cytokiny chemie genetika MeSH
- Escherichia coli enzymologie ultrastruktura MeSH
- krystalografie rentgenová MeSH
- kvantová teorie MeSH
- leukocyty mononukleární chemie enzymologie MeSH
- lidé MeSH
- membránové proteiny chemie genetika ultrastruktura MeSH
- nukleotidy biosyntéza chemie genetika MeSH
- přirozená imunita genetika MeSH
- substrátová specifita MeSH
- Thermotoga maritima enzymologie ultrastruktura MeSH
- Vibrio cholerae enzymologie ultrastruktura MeSH
- vztahy mezi strukturou a aktivitou * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Plastids, organelles that evolved from cyanobacteria via endosymbiosis in eukaryotes, provide carbohydrates for the formation of biomass and for mitochondrial energy production to the cell. They generate their own energy in the form of the nucleotide adenosine triphosphate (ATP). However, plastids of non-photosynthetic tissues, or during the dark, depend on external supply of ATP. A dedicated antiporter that exchanges ATP against adenosine diphosphate (ADP) plus inorganic phosphate (Pi) takes over this function in most photosynthetic eukaryotes. Additional forms of such nucleotide transporters (NTTs), with deviating activities, are found in intracellular bacteria, and, surprisingly, also in diatoms, a group of algae that acquired their plastids from other eukaryotes via one (or even several) additional endosymbioses compared to algae with primary plastids and higher plants. In this review, we summarize what is known about the nucleotide synthesis and transport pathways in diatom cells, and discuss the evolutionary implications of the presence of the additional NTTs in diatoms, as well as their applications in biotechnology.
Synthesis of base-modified dNTPs through the Suzuki or Sonogashira cross-coupling reactions of halogenated dNTPs with boronic acids or alkynes is reported, as well as the use of these modified dNTPs in polymerase incorporations to oligonucleotides or DNA by primer extension or PCR.
Our investigation in the chemistry of nucleoside phosphonates has provided an impressive number of novel structurally diverse compounds. Among them, several potent inhibitors of enzymes of nucleosides and nucleotide salvage pathways were found. The findings obtained in synthesis of nucleoside phosphonates are unique. The nucleoside phosphonates provided several types of monomers for the synthesis of modified oligonucleotides. The first non-silyl protecting group was developed and used in the solid-phase synthesis of modified oligoribonucleotides in the reverse direction. The superior nuclease stability of phosphonate internucleotide linkages, their ability to enhance hybridization together with the ability to elicit RNase H activity may classify these compounds as useful in biochemistry and biology
- MeSH
- biochemické jevy MeSH
- fosforylasy chemie metabolismus MeSH
- inhibitory enzymů chemie metabolismus MeSH
- lidé MeSH
- molekulární konformace MeSH
- nukleotidy biosyntéza chemie metabolismus MeSH
- oligonukleotidy * biosyntéza chemie metabolismus MeSH
- organofosfonáty * chemická syntéza chemie metabolismus MeSH
- ribonukleasy chemie metabolismus MeSH
- toll-like receptory chemie metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
- Klíčová slova
- Inosin 5-mono-fosfát dehydrogenáza, inkluze, ultrastruktura,
- MeSH
- elektronová mikroskopie využití MeSH
- inhibitory enzymů * izolace a purifikace terapeutické užití MeSH
- konformace proteinů * účinky léků MeSH
- lidé MeSH
- nukleotidy biosyntéza MeSH
- prospektivní studie MeSH
- zmrazování MeSH
- Check Tag
- lidé MeSH
- Klíčová slova
- PREVENTAN, JUWIM, JUWÍK,
- MeSH
- lidé MeSH
- nukleotidy biosyntéza imunologie metabolismus MeSH
- polynukleotidy aplikace a dávkování imunologie terapeutické užití MeSH
- potraviny využití MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- klinické zkoušky MeSH
- MeSH
- adenin MeSH
- azacytidin MeSH
- morčata MeSH
- mozková kůra MeSH
- nukleotidy biosyntéza MeSH
- puriny MeSH
- techniky in vitro MeSH
- Check Tag
- morčata MeSH
