Morphological Transformation and Surface Engineering of Glycovesicles Driven by Bioinspired Hydrogen Bonds of Nucleobases.

Glycopolymer-based supramolecular glycoassemblies with signal-driven cascade morphological deformation and accessible surface engineering toward bioinspired functional glycomaterials have attracted much attention due to their diverse applications in fundamental and practical scenarios. Herein, we achieved the cascade morphological transformation and surface engineering of a nucleobase-containing polymeric glycovesicle through exploiting the bioinspired complementary multiple hydrogen bonds of complementary nucleobases. First, the synthesized thymine-containing glycopolymers (PGal30-b-PTAm249) are capable of self-assembling into well-defined glycovesicles. Several kinds of amphiphilic adenine-containing block copolymers with neutral, positive, and negative charges were synthesized to engineer the glycovesicles through the multiple hydrogen bonds between adenine and thymine. A cascade of morphological transformations from vesicles to ruptured vesicles with tails, to worm-like micelles, and finally to spherical micelles were observed via continuously adding the adenine-containing polymer into the thymine-containing glycovesicles. Furthermore, the surface charge properties of these glyconano-objects can be facilely regulated through incorporating various adenine-containing polymers. This work demonstrates the potential application of a unique bioinspired approach to precisely engineer the morphology and surface properties of glycovesicles for boosting their biological applications.

N/O→B dative bonds supplemented by N-HN/HC hydrogen bonds make BN-cages an attractive candidate for DNA-nucleobase adsorption - an MP2 prediction.

The response of B12N12-nanocages towards DNA-nucleobases (adenine, guanine, cytosine, and thymine) is investigated using MP2 and DFT (M06-2X) levels of theory with the 6-311+G** basis set. Multiple BN-cage-nucleobase structures for each nucleobase emerged depending on the number of Lewis base centers of nucleobases. The main source of stability of these complexes is the N/O→B dative bond, where the N or O atom of nucleobases donates the lone-pair electron to one of the boron atoms of the nanocage. Nitrogen atoms of the BN-cage, adjacent to the B-site forming dative bond, act as a proton acceptor to form multiple (N-HN and N-HC) hydrogen bonds, where proton-donors NH and CH are part of nucleobases. MP2/6-311+G** adsorption energies are -43.1, -43.4 and -45.3 kcal mol-1 (B12N12-adenine), -37.1, -41.9 and -43.3 kcal mol-1 (B12N12-guanine), -41.3 and -43.4 (B12N12-cytosine), and -29.3 and -31.3 (B12N12-thymine). Similar adsorption energies were recorded for larger BN-fullerenes-nucleobases, namely B16N16 and B24N24. Changes in adsorption energies and structures of these nano-bio-hybrid materials in aqueous media are also discussed. Computationally cost-effective MP2 single point calculations at the M06-2X optimized geometries were found to be reliable in predicting adsorption energies. The effect of the BN-network and H-bonds on the adsorption process is assessed by comparing the results with simple BH3-nucleobase models. BSSE correction to the adsorption energy is not recommended.

Glyco-Nanoadjuvants: Sugar Structures on Carriers of a Small Molecule TLR7 Ligand Affect Their Immunostimulatory Activities.

Toll-like receptors (TLRs) are pattern recognition receptors that activate innate immunity, and their ligands are promising adjuvants for vaccines and immunotherapies. Small molecule TLR7 ligands are ideal vaccine adjuvants as they induce not only proinflammatory cytokines but also type I interferons. However, their application has only been approved for local administration due to severe systemic immune-related adverse events. In a previous study, we prepared the gold nanoparticles coimmobilized with synthetic small molecule TLR7 ligand, 1V209, and α-mannose (1V209-αMan-GNPs). 1V209-αMan-GNPs were selectively delivered via a cell surface sugar-binding protein, mannose receptor, which enabled selective delivery of TLR7 ligands to immune cells. Besides the mannose receptor, immune cells express various sugar-binding proteins such as macrophage galactose binding lectins and sialic acid-binding immunoglobulin-type lectins and recognize distinct sugar structures. Hence, in the present study, we investigated whether sugar structures on GNPs affect the efficiency and selectivity of intracellular delivery and subsequent immunostimulatory potencies. Five neutral sugars and two sialosides were selected and each sugar was coimmobilized with 1V209 onto GNPs (1V209-SGNPs) and their innate immunostimulatory potencies were compared to that of 1V209-αMan-GNPs. The in vitro study using mouse bone marrow derived dendritic cells (BMDCs) demonstrated that α-glucose, α-N-acetylglucosamine, or α-fucose immobilized 1V209-SGNPs increased interleukin-6 and type I interferon release similar to that of 1V209-αMan-GNPs, whereas galacto-type sugar immobilized 1V209-SGNPs predominantly enhanced type I interferon release. In contrast, sialoside immobilized 1V209-SGNPs did not enhance the potency of 1V209. In the in vivo immunization study using ovalbumin as a model antigen, neutral sugar immobilized 1V209-SGNPs induced comparable T helper-1 immune response to that of 1V209-αMan-GNPs and by 10-fold higher than that of sialoside immobilized 1V209-SGNPs. These results indicate that the sugar structures on 1V209-SGNPs affect their immunostimulatory activities, and functionalization of the carrier particles is important to shape immune responses.

Determinants of adenine-mutagenesis in diversity-generating retroelements.

Diversity-generating retroelements (DGRs) vary protein sequences to the greatest extent known in the natural world. These elements are encoded by constituents of the human microbiome and the microbial 'dark matter'. Variation occurs through adenine-mutagenesis, in which genetic information in RNA is reverse transcribed faithfully to cDNA for all template bases but adenine. We investigated the determinants of adenine-mutagenesis in the prototypical Bordetella bacteriophage DGR through an in vitro system composed of the reverse transcriptase bRT, Avd protein, and a specific RNA. We found that the catalytic efficiency for correct incorporation during reverse transcription by the bRT-Avd complex was strikingly low for all template bases, with the lowest occurring for adenine. Misincorporation across a template adenine was only somewhat lower in efficiency than correct incorporation. We found that the C6, but not the N1 or C2, purine substituent was a key determinant of adenine-mutagenesis. bRT-Avd was insensitive to the C6 amine of adenine but recognized the C6 carbonyl of guanine. We also identified two bRT amino acids predicted to nonspecifically contact incoming dNTPs, R74 and I181, as promoters of adenine-mutagenesis. Our results suggest that the overall low catalytic efficiency of bRT-Avd is intimately tied to its ability to carry out adenine-mutagenesis.

Enzymatic synthesis of hypermodified DNA polymers for sequence-specific display of four different hydrophobic groups.

A set of modified 2'-deoxyribonucleoside triphosphates (dNTPs) bearing a linear or branched alkane, indole or phenyl group linked through ethynyl or alkyl spacer were synthesized and used as substrates for polymerase synthesis of hypermodified DNA by primer extension (PEX). Using the alkyl-linked dNTPs, the polymerase synthesized up to 22-mer fully modified oligonucleotide (ON), whereas using the ethynyl-linked dNTPs, the enzyme was able to synthesize even long sequences of >100 modified nucleotides in a row. In PCR, the combinations of all four modified dNTPs showed only linear amplification. Asymmetric PCR or PEX with separation or digestion of the template strand can be used for synthesis of hypermodified single-stranded ONs, which are monodispersed polymers displaying four different substituents on DNA backbone in sequence-specific manner. The fully modified ONs hybridized with complementary strands and modified DNA duplexes were found to exist in B-type conformation (B- or C-DNA) according to CD spectral analysis. The modified DNA can be replicated with high fidelity to natural DNA through PCR and sequenced. Therefore, this approach has a promising potential in generation and selection of hypermodified aptamers and other functional polymers.

Adenine-Functionalized Supramolecular Micelles for Selective Cancer Chemotherapy.

Functional supramolecular micelles containing self-complementary multiple hydrogen bonding adenine groups (A-PPG) can spontaneously self-assemble into stable nanosized micelles in aqueous solution. These micelles can be used to selectively deliver anticancer drugs to cancer cells and effectively promote tumor cell death via apoptosis, without harming normal cells. The drug-loaded micelles exhibit tunable drug-loading capacity and rapid pH-triggered drug release under acidic conditions, as well as a high drug-entrapment stability in serum-rich media due to the reversible hydrogen-bonded adenine-adenine interactions within the micellar interior; these properties are critical to achieving effective chemotherapeutic drug delivery and controlled drug release. In vitro assays show that the drug-loaded micelles exert significant cytotoxic effects on cancer cells, with minimal effects on normal cells under physiological conditions. Cytotoxicity assays using A-PPG micelles loaded with different anticancer drugs confirm these effects. Importantly, cellular internalization and flow cytometric analyses demonstrate that the adenine moieties within A-PPG micelles significantly increase selective endocytic uptake of the supramolecular micelles by cancer cells, which in turn induce apoptotic cell death and substantially enhance the response to chemotherapy. Thus, A-PPG micelles can improve the safety and efficacy of cancer chemotherapy.

Adenine-coated magnetic multiwalled carbon nanotubes for the selective extraction of aristolochic acids based on multiple interactions.

A method is described for the functionalization of magnetic carbon nanotubes to recognize aristolochic acid Ⅰ and Ⅱ. 3-Glycidyloxypropyltrimethoxysilane was used as a coupling agent to immobilize adenine on a solid support. The morphology and structure of adenine-coated magnetic carbon nanotubes was investigated using transmission electron microscopy (TEM), scanning electron microscopy (SEM), X-ray diffraction (XRD) and a vibrating sample magnetometer (VSM). The adsorption performance of the adenine-coated magnetic carbon nanotubes was evaluated via adsorption isotherms, the kinetics and selectivity tests. The adsorption capacity of the adenine-functionalized sorbent for aristolochic acid Ⅰ was determined to be 24.5 µg mg-1. By combining magnetic solid phase extraction with HPLC detection, a method was developed to enrich and detect aristolochic acids used in traditional Chinese medicine. A satisfactory recovery (92.7 - 97.5% for aristolochic acid Ⅰ and 92.6 - 99.4% for aristolochic acid Ⅱ) and an acceptable relative standard deviation (<4.0%) were obtained.

Structure of a DNA polymerase abortive complex with the 8OG:dA base pair at the primer terminus.

Adenine frequently pairs with the Hoogsteen edge of an oxidized guanine base (8OG) causing G to T transversions. The (syn)8OG:dA base pair is indistinguishable from the cognant base pair and can be extended by DNA polymerases with reduced efficiency. To examine the structural basis of this reduced efficiency, we sought to obtain the structure of the "product" complex of DNA polymerase (pol) ß with the (syn)8OG:dA base pair at the primer terminus by soaking the binary complex crystals with a hydrolysable dCTP analogue complementary to the template base G. Crystallographic refinement of the structure revealed that the adenine of the (syn)8OG:dA base pair had been expelled from the primer terminus and a dCMP was inserted opposite 8OG in a reverse orientation; another uninserted molecule of the analogue was bound to the templating base G. This leads to an abortive complex that could form the basis of oxidatively-induced pol ß stalling.

Self-assembled nucleo-tripeptide hydrogels provide local and sustained doxorubicin release.

Self-assembled nucleo-peptide hydrogels have a nanofibril structure composed of noncovalent molecular interactions between peptide groups as well as π-π stacking and Watson-Crick interactions via complementary nucleobases. These hydrogels have specific benefits for biomedical applications due to their DNA-like interactions in addition to the well-known advantages of peptide biomaterials: biocompatibility, extracellular matrix (ECM)-like structure, and bottom-up design. Inspired by the nucleobase stacking structure, we hypothesized that nucleo-peptides would be able to deliver the DNA-intercalating chemotherapeutic, doxorubicin (Dox) in a sustained manner when delivered locally to a solid tumor. Ade-FFF nucleo-peptide hydrogels were able to load a high concentration of Dox (1 mM) and demonstrated continuous release under in vitro degradation conditions. We adopted an in vivo tumor-bearing mouse model to evaluate the delivery of Dox by Ade-FFF hydrogels. We found that Dox-containing hydrogels reduced tumor growth and resulted in greater apoptosis-mediated cell death in the tumor as evidenced by caspase-3 expression. Pharmacokinetics and biodistribution studies also supported the observation that Dox delivery by an Ade-FFF hydrogel improves sustained delivery in the local tumor site. This study demonstrates the potential of self-assembled nucleo-peptides in biomedical applications by using their distinctive DNA-like structure.

Functional Systems Derived from Nucleobase Self-assembly.

Dynamic and reversible non-covalent interactions endow synthetic systems and materials with smart adaptive functions that allow them to response to diverse stimuli, interact with external agents, or repair structural defects. Inspired by the outstanding performance and selectivity of DNA in living systems, scientists are increasingly employing Watson-Crick nucleobase pairing to control the structure and properties of self-assembled materials. Two sets of complementary purine-pyrimidine pairs (guanine:cytosine and adenine:thymine(uracil)) are available that provide selective and directional H-bonding interactions, present multiple metal-coordination sites, and exhibit rich redox chemistry. In this review, we highlight several recent examples that profit from these features and employ nucleobase interactions in functional systems and materials, covering the fields of energy/electron transfer, charge transport, adaptive nanoparticles, porous materials, macromolecule self-assembly, or polymeric materials with adhesive or self-healing ability.

Honey as Source of Nitrogen Compounds: Aromatic Amino Acids, Free Nucleosides and Their Derivatives.

The content of selected major nitrogen compounds including nucleosides and their derivatives was evaluated in 75 samples of seven varieties of honey (heather, buckwheat, black locust, goldenrod, canola, fir, linden) by targeted ultra-high performance liquid chromatography-diode array detector - high-resolution quadrupole time-of-flight mass spectrometry (UHPLC-DAD-QqTOF-MS) and determined by UHPLC-DAD. The honey samples contained nucleosides, nucleobases and their derivatives (adenine: 8.9 to 18.4 mg/kg, xanthine: 1.2 to 3.3 mg/kg, uridine: 17.5 to 51.2 mg/kg, guanosine: 2.0 to 4.1 mg/kg; mean amounts), aromatic amino acids (tyrosine: 7.8 to 263.9 mg/kg, phenylalanine: 9.5 to 64.1 mg/kg; mean amounts). The amounts of compounds significantly differed between some honey types. For example, canola honey contained a much lower amount of uridine (17.5 ± 3.9 mg/kg) than black locust where it was most abundant (51.2 ± 7.8 mg/kg). The presence of free nucleosides and nucleobases in different honey varieties is reported first time and supports previous findings on medicinal activities of honey reported in the literature as well as traditional therapy and may contribute for their explanation. This applies, e.g., to the topical application of honey in herpes infections, as well as its beneficial activity on cognitive functions as nootropic and neuroprotective, in neuralgia and is also important for the understanding of nutritional values of honey.

Optimization of 8-oxoadenines with toll-like-receptor 7 and 8 activity.

Toll-like receptors 7 and 8 (TLR7/8) agonists are potent immunostimulants that are attracting considerable interest as vaccine adjuvants. We recently reported the synthesis of a new series of 2-O-butyl-8-oxoadenines substituted at the 9-position with various linkers and N-heterocycles, and showed that TLR7/8 selectivity, potency and cytokine induction could be modulated by varying the alkyl linker length and the N-heterocyclic ring. In the present study, we further optimized the oxoadenine scaffold by investigating the effect of different substituents at the 2-position of the oxoadenine on TLR7/8 potency/selectivity, cytokine induction and DC maturation in human PBMCs. The results show that introducing a 1-(S)-methylbutoxy group at the 2-position of the oxoadenine significantly increased potency for TLR7/8 activity, cytokine induction and DC maturation.

Theoretical analyses and experimental validation of the effects caused by the fluorinated substituent modification of DNA.

Halogen-modified nucleic acid molecules, such as trifluorothymidine (FTD) and 5-fluorouracil, are widely used in medical science and clinical site. These compounds have a very similar nucleobase structure. It is reported that both of these compounds could be incorporated into DNA. The incorporation of FTD produces highly anti-tumor effect. However, it is not known whether to occur a significant effect by the incorporation of 5-fluorouracil. Nobody knows why such a difference will occur. To understand the reason why there is large differences between trifluorothymidine and 5-fluorouracil, we have performed the molecular dynamics simulations and molecular orbital calculations. Although the active interaction energy between Halogen-modified nucleic acids or and complementary adenine was increased, in only FTD incorporated DNA, more strongly dispersion force interactions with an adjacent base were detected in many thermodynamic DNA conformations. As the results, the conformational changes occur even if it is in internal body temperature. Then the break of hydrogen bonding between FTD and complementary adenine base occur more frequently. The double helix structural destabilization of DNA with FTD is resulted from autoagglutination caused by the bonding via halogen orbitals such as halogen bonding and the general van der Waals interactions such as CH-[Formula: see text], lone pair (LP)-[Formula: see text], and [Formula: see text]-[Formula: see text] interactions. Therefore, it is strongly speculated that such structural changes caused by trifluoromethyl group is important for the anti-tumor effect of FTD alone.

Modulating Morphologies and Surface Properties of Nanoparticles from Cellulose-Grafted Bottlebrush Copolymers Using Complementary Hydrogen-Bonding between Nucleobases.

Herein we report the synthesis of a cellulose-grafted bottlebrush copolymer with nucleobases as hydrophobic moieties. Well-defined spherical micelles from this bottlebrush copolymer were fabricated via a solvent switch method. A morphological transition from spheres to worms was only observed to occur when a diblock copolymer with a complementary nucleobase functionality was introduced. Hydrophobic interaction is not capable of triggering the morphological transformation, and the diblock copolymer with the heterogeneous acrylamide nucleobase monomer can induce the morphological transition at higher A:T molar ratios, which might be caused by the weak H-bonding interaction. This supramolecular "grafting to" method enables the preparation of a series of nanoparticles with similar shapes and dimensions but distinct surface properties such as zeta potentials. Moreover, reversible morphological transitions between worm-like micelles and spheres can be achieved using a reversible collapsing and swelling of a thermoresponsive polymer. This work highlights that a supramolecular "grafting to" approach between complementary nucleobases can be utilized to tune morphologies and surface properties of nanoparticles.

Adenine nucleobase directed supramolecular architectures based on ferrimagnetic heptanuclear copper(II) entities and benzenecarboxylate anions.

Two planar organic anions, benzoate and benzene-1,4-dicarboxylate (terephthalate), have been selected as potential π-stacking intercalators among ferrimagnetic [Cu7(µ-adeninato)6(µ3-OH)6(µ-H2O)6]2+ heptameric discrete entities. The resulting supramolecular architecture is highly dependent on the negative charge density distribution, mainly located in the carboxylate groups of the organic anions. In this sense, the benzoate anion, with just one carboxylate group, does not allow its intercalation between the adeninato ligands as it would imply a high steric hindrance among the heptameric entities. As a consequence, these benzoate anions are located inside the voids of the crystal structure reducing the accessible volume of compound [Cu7(µ-adeninato)6(µ3-OH)6(µ-H2O)6](benzoate)2·~17H2O (1). On the contrary, the terephthalate anion, containing two carboxylate groups at opposite sites, adopts a π-stacking sandwich arrangement between two adeninato ligands that affords the porous open structure of formula [Cu7(µ-adeninato)6(µ3-OH)6(µ-H2O)6](terephthalate)·nH2O (2a, 2b; n: 12 and 24, respectively). In addition to that, the less directional nature of the π-stacking interactions in comparison to the complementary hydrogen bonding based supramolecular metal-organic frameworks (SMOFs), suits them with a flexible architecture able to reversibly adsorb/desorb water (up to a 25-30% at 20 °C) altogether with the expansion/shrinkage of the crystal structure. The bridging adeninato and hydroxido ligands are effective magnetic exchange mediators to provide a ST = 5/2 ferrimagnetic state for the heptanuclear entity.

Gold Nanoparticles Coimmobilized with Small Molecule Toll-Like Receptor 7 Ligand and α-Mannose as Adjuvants.

Adjuvants enhance the immune response during vaccination. Among FDA-approved adjuvants, aluminum salts are most commonly used in vaccines. Although aluminum salts enhance humoral immunity, they show a limited effect for cell-mediated immune responses. Thus, further development of adjuvants that induce T-cell-mediated immune response is needed. Toll-like receptors (TLRs) recognizing specific pathogen-associated molecular patterns activate innate immunity, which is crucial to shape adaptive immunity. Using TLR ligands as novel adjuvants in vaccines has therefore attracted substantial attention. Among them a small molecule TLR7 ligand, imiquimod, has been approved for clinical use, but its use is restricted to local administration due to unwanted adverse side effects when used systematically. Since TLR7 is mainly located in the endosomal compartment of immune cells, efficient transport of the ligand into the cells is important for improving the potency of the TLR7 ligand. In this study we examined gold nanoparticles (GNPs) immobilized with α-mannose as carriers for a TLR7 ligand to target immune cells. The small molecule synthetic TLR7 ligand, 2-methoxyethoxy-8-oxo-9-(4-carboxy benzyl)adenine (1V209), and α-mannose were coimmobilized via linker molecules consisting of thioctic acid on the GNP surface (1V209-αMan-GNPs). The in vitro cytokine production activity of 1V209-αMan-GNPs was higher than that of the unconjugated 1V209 derivative in mouse bone marrow-derived dendritic cells and in human peripheral blood mononuclear cells. In the in vivo immunization study, 1V209-αMan-GNPs induced significantly higher titers of IgG2c antibody specific to ovalbumin as an antigen than did unconjugated 1V209, and splenomegaly and weight loss were not observed. These results indicate that 1V209-αMan-GNPs could be useful as safe and effective adjuvants for development of vaccines against infectious diseases and cancer.

DFT and MD Studies of Formaldehyde-Derived DNA Adducts: Molecular-Level Insights into the Differential Mispairing Potentials of the Adenine, Cytosine, and Guanine Lesions.

4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone is a potent nicotine-based carcinogen that generates many DNA lesions, including the HOCH2-C, HOCH2-G, and HOCH2-A hydroxymethyl adducts. Despite all lesions containing an altered exocyclic amino group, which allows the hydroxymethyl group to be directed away from the Watson-Crick binding face, only the most persistent adenine adduct is mutagenic. As a first step toward understanding this differential mutagenicity, density functional theory (DFT) and molecular dynamics (MD) simulations were used to gain atomic-level structural details of these DNA damage products. DFT calculations reveal that all three lesions exhibit conformational diversity. However, regardless of the hydroxymethyl-nucleobase orientation, both DFT and MD simulations highlight that HOCH2-C and HOCH2-G form pairs with the canonical complementary base (G and C, respectively) that are structural and energetically preferred over mispairs. In contrast, depending on the hydroxymethyl-nucleobase orientation, the Watson-Crick HOCH2-A:T pair can become significantly destabilized relative to undamaged A:T. As a result, HOCH2-A mispairs with G, C, and A are energetically accessible and maintain key geometrical features of canonical DNA. Overall, our data directly correlate with the reported differential mutagenicity of the hydroxylmethyl lesions and will encourage future studies to further uncover the cellular impact of the most persistent adenine lesion.

Entrapment of an adenine derivative by a photo-irradiated uracil-functionalized micelle confers controlled self-assembly behavior.

HYPOTHESIS: Invoking cooperative assembly of the uracil-functionalized supramolecular polymer BU-PPG [uracil end-capped poly(propylene glycol)] upon association with the nucleobase adenine derivative A-MA [methyl 3-(6-amino-9H-purin-9-yl)propanoate] as a model drug provides a new concept to control and tune the properties of supramolecular complexes and holds significant potential for the development of safer, more effective drug delivery systems. EXPERIMENTS: BU-PPG and A-MA were successfully developed and exhibited specific recognition and high affinity, which enabled reversible complementary adenine-uracil (A-U) hydrogen bonding-induced formation of spherical micelles in aqueous solution. The self-assembly and controllable A-MA release behavior of BU-PPG/A-MA micelles were studied using morphological analysis and optical and light scattering techniques to investigate the effect of photoirradiation and temperature on the complementary hydrogen bond interactions between BU-PPG and A-MA. FINDINGS: The resulting micelles possess unusual physical properties, including controlled photoreactivity kinetics, controllable self-assembled morphology and low cytotoxicity in vitro, as well as reversible temperature-responsive behavior. Importantly, irradiated micelles exhibited excellent long-term structural stability under normal physiological conditions and serum disturbance. Increasing the temperature triggered rapid release of A-MA by disrupting A-U complexes. These findings represent an entirely new, promising strategy for the development of multi-controlled release drug delivery nanocarriers based on complementary hydrogen bonding interactions.

Dual control of NAD+ synthesis by purine metabolites in yeast.

Metabolism is a highly integrated process resulting in energy and biomass production. While individual metabolic routes are well characterized, the mechanisms ensuring crosstalk between pathways are poorly described, although they are crucial for homeostasis. Here, we establish a co-regulation of purine and pyridine metabolism in response to external adenine through two separable mechanisms. First, adenine depletion promotes transcriptional upregulation of the de novo NAD+ biosynthesis genes by a mechanism requiring the key-purine intermediates ZMP/SZMP and the Bas1/Pho2 transcription factors. Second, adenine supplementation favors the pyridine salvage route resulting in an ATP-dependent increase of intracellular NAD+. This control operates at the level of the nicotinic acid mononucleotide adenylyl-transferase Nma1 and can be bypassed by overexpressing this enzyme. Therefore, in yeast, pyridine metabolism is under the dual control of ZMP/SZMP and ATP, revealing a much wider regulatory role for these intermediate metabolites in an integrated biosynthesis network.

Synthesis and Antiviral Evaluation of 3'-Fluoro-4'-modified-5'-norcarbocyclic Nucleoside Phosphonates.

The synthesis and anti-HIV evaluation of hitherto unknown 3'-fluoro-5'-norcarbocyclic nucleoside phosphonates bearing adenine with modifications at the 4' position (ethynyl, vinyl, ethyl, hydroxymethyl) is described. One of the synthesized compounds was found to be an inhibitor of HIV-1 replication, but with moderate efficiency relative to (R)-9-(2-phosphonylmethoxypropyl)adenine ((R)-PMPA, tenofovir), with no concomitant cytotoxicity.