Synthesis and Pairing Properties of 3′‐Deoxyribopyranose (4′→2′)‐Oligonucleotides (‘p‐DNA')

The preparation and the pairing properties of the new 3′‐deoxyribopyranose (4′→2′)‐oligonucleotide (=p‐DNA) pairing system, based on 3′‐deoxy‐β‐D ‐ribopyranose nucleosides is presented. D ‐Xylose was efficiently converted to the prefunctionalized 3‐deoxyribopyranose derivative 4‐O‐[(tert‐butyl)dimethylsilyl]‐3‐deoxy‐D ‐ribopyranose 1,2‐diacetate 8 (obtained as a 4 : 1 mixture of α‐ and β‐D ‐anomers; Scheme 1). From this sugar building block, the corresponding, appropriately protected thymine, guanine, 5‐methylcytosine, and purine‐2,6‐diamine nucleoside phosphoramidites 29 – 32 were prepared in a minimal number of steps (Schemes 2–4). These building blocks were assembled on a DNA synthesizer, and the corresponding p‐DNA oligonucleotides were obtained in good yields after a one‐step deprotection under standard conditions, followed by HPLC purification (Scheme 5 and Table 1). Qualitatively, p‐DNA shows the same pairing behavior as p‐RNA, forming antiparallel, exclusively Watson‐Crick‐paired duplexes that are much stronger than corresponding DNA duplexes. Duplex stabilities within the three related (i.e., based on ribopyranose nucleosides) oligonucleotide systems p‐RNA, p‐DNA, and 3′‐O‐Me‐p‐RNA were compared with each other (Table 2). Intrinsically, p‐RNA forms the strongest duplexes, followed by p‐DNA, and 3′‐O‐Me‐p‐RNA. However, by introducing the nucleobases purine‐2,6‐diamine (D) and 5‐methylcytosine (M) instead of adenine and cytosine, a substantial increase in stability of corresponding p‐DNA duplexes was observed.     

Synthesis and Hybridization Properties of an Oligonucleotide Consisting of 1′,4′‐Anhydro‐2′,5′‐dideoxy‐2′‐(thymin‐1‐yl)‐D‐altritol

The novel oligonucleotide analogue 7 , consisting of 1′,4′‐anhydro‐2′,5′‐dideoxy‐2′‐(thymin‐1‐yl)‐D ‐altritol ( 4 ), residues was synthesized by the phosphoramidite approach on an automated DNA synthesizer. The phosphoramidite building block 6 was obtained by phosphitylation of the corresponding isonucleoside 5 . Oligoisonucleotide 7 contains an extended phosphodiester linkage with a higher flexibility. Oligoisonucleotide 7 was studied with respect to hybridization properties, enzymatic stability, and CD spectra. It exhibits a high stability towards snake‐venom phosphodiesterase and an acceptable hybridization to complementary single‐stranded DNA and RNA.     

Type I Photosensitization of 2′‐deoxyadenosine 5′‐monophosphate (5′‐dAMP) by Biopterin and its Photoproduct Formylpterin

Biopterin (Bip) and its photoproducts 6‐formylpterin (Fop) and 6‐carboxypterin (Cap) accumulate in the skin of patients suffering from vitiligo, a chronic depigmentation disorder where the protection against UV radiation fails because of the lack of melanin. These compounds absorb in the UV‐A inducing a potential photosensitizing action that can cause damage to DNA and other biomolecules. In this work, we have investigated the capability of these pterin derivatives (Pt) to act as photosensitizers under UV‐A irradiation for the degradation of 2′‐deoxyadenosine 5′‐monophosphate (5′‐dAMP) in aqueous solutions, as model DNA target. Steady‐state and time‐resolved experiments were performed and the effect of pH was evaluated. The results showed that photosensitized degradation of 5′‐dAMP was only observed under acidic conditions, and a mechanistic analysis revealed the participation of the triplet excited state of the pterin derivatives (³Pt*) by electron transfer yielding the corresponding pair of radical ions (Pt^?? and 5′‐dAMP^?+), with successive photosensitizer recovery by electron transfer from Pt^?? to O₂. Finally, 5′‐dAMP^?+ participates in subsequent reactions to yield degradation products.     

Photophysical and Duplex‐DNA‐Binding Properties of Distamycin Dimers Based on 4,4′‐ and 2,2′‐Dialkoxyazobenzenes as the Core

Distamycin‐based tetrapeptide ( 1 ) was covalently tethered to both ends of the central dihydroxyazobenzene moiety at either the 2,2′ or 4,4′ positions. This afforded two isomeric, distamycin–azobenzene–distamycin systems, 2 (para) and 3 (ortho), both of them being photoisomerizable. Illumination of these conjugates in solution at approximately 360 nm induced photoisomerization and the time course of the process was followed by UV/Vis and ¹H NMR spectroscopy. The kinetics of the thermal reversion at various temperatures of cis to trans isomers of the conjugates obtained after photoillumination were also examined. This afforded the respective thermal‐activation parameters. Both the molecular architecture and the location of the substituent around the core azobenzene determined the rate and activation‐energy barrier for the cis‐to‐trans back‐isomerization of these conjugates in solution. Duplex–DNA binding of the conjugates and the changes in DNA‐binding efficiency upon photoisomerization was also examined by CD spectroscopy, thermal denaturation studies, and a Hoechst displacement assay. The conjugate 2 showed higher DNA‐binding affinity and a greater change in the DNA‐binding efficiency upon photoisomerization compared with its 2,2′‐disubstituted counterpart. The experimental findings were substantiated by using molecular‐docking studies involving each conjugate with a model duplex d[(GC(AT)₁₀CG)]₂ DNA molecule.     

2,3,3′,6‐Tetra‐O‐acetyl‐4,1′,6′‐trichloro‐4,1′,4′,6′‐tetradeoxygalactosucrose

At 160 K, one of the Cl atoms in the furanoid moiety of 3‐O‐acetyl‐1,6‐di­chloro‐1,4,6‐tri­deoxy‐β‐d ‐fructo­furan­osyl 2,3,6‐tri‐O‐acetyl‐4‐chloro‐4‐deoxy‐α‐d ‐galacto­pyran­oside, C₂₀H₂₇­Cl₃O₁₁, is disordered over two orientations, which differ by a rotation of about 107° about the parent C—C bond. The conformation of the core of the mol­ecule is very similar to that of 3‐O‐acetyl‐1,4,6‐tri­chloro‐1,4,6‐tri­deoxy‐β‐d ‐tagato­furanos­yl 2,3,6‐tri‐O‐acetyl‐4‐chloro‐4‐deoxy‐α‐d ‐galacto­pyran­oside, particularly with regard to the conformation about the glycosidic linkage.     

Stereoselective Synthesis of [5‐[4,4,4,4′,4′,4′‐Hexafluoro‐N‐(2‐hydroxyethoxy)‐D‐valine]]‐ and [5‐[4,4,4,4′,4′,4′‐Hexafluoro‐N‐(2‐hydroxyethoxy)‐L‐valine]cyclosporin A

Addition of various amines to the 3,3‐bis(trifluoromethyl)acrylamides 10a and 10b gave the tripeptides 11a – 11f , mostly as mixtures of epimers (Scheme 3). The crystalline tripeptide 11f ₂ was found to be the N‐terminal (2‐hydroxyethoxy)‐substituted (R,S,S)‐ester HOCH₂CH₂O‐D ‐Val(F₆)‐MeLeu‐Ala‐O^tBu by X‐ray crystallography. The C‐terminal‐protected tripeptide 11f ₂ was condensed with the N‐terminus octapeptide 2b to the depsipeptide 12a which was thermally rearranged to the undecapeptide 13a (Scheme 4). The condensation of the epimeric tripeptide 11f ₁ with the octapeptide 2b gave the undecapeptide 13b directly. The undecapeptides 13a and 13b were fully deprotected and cyclized to the [5‐[4,4,4,4′,4′,4′‐hexafluoro‐N‐(2‐hydroxyethoxy)‐D ‐valine]]‐ and [5‐[4,4,4,4′,4′,4′‐hexafluoro‐N‐(2‐hydroxyethoxy)‐L ‐valine]]cyclosporins 14a and 14b , respectively (Scheme 5). Rate differences observed for the thermal rearrangements of 12a to 13a and of 12b to 13b are discussed.     

Ultra‐fast adenosine 5′‐triphosphate,adenosine 5′‐diphosphate and adenosine 5′‐monophosphate detection by pressure‐assisted capillary electrophoresis UV detection

Herein, we report a new CE method to measure adenine nucleotides adenosine 5′‐triphosphate, adenosine 5′‐diphosphate, and adenosine 5′‐monophosphate in red blood cells. For this purpose, 20 mmol/L sodium acetate buffer at pH 3.80 was used as running electrolyte, and the separation was performed by the simultaneous application of a CE voltage of 25 kV and an overimposed pressure of 0.2 psi from inlet to outlet. A rapid separation of these analytes in less than 1.5 min was obtained with a good reproducibility for intra‐ and inter‐assay (CV<4 and 8%, respectively) and an excellent analytical recovery (from 98.3 to 99%). The applicability of our method was proved by measuring adenine nucleotides in red blood cells.     

Synthesis and Base Pairing Properties of 1′,5′‐Anhydro‐L‐Hexitol Nucleic Acids (L‐HNA)

Oligonucleotides composed of 1′,5′‐anhydro‐arabino‐hexitol nucleosides belonging to the L series (L ‐HNA) were prepared and preliminarily studied as a novel potential base‐pairing system. Synthesis of enantiopure L ‐hexitol nucleotide monomers equipped with a 2′‐(N⁶‐benzoyladenin‐9‐yl) or a 2′‐(thymin‐1‐yl) moiety was carried out by a de novo approach based on a domino reaction as key step. The L oligonucleotide analogues were evaluated in duplex formation with natural complements as well as with unnatural sugar‐modified oligonucleotides. In many cases stable homo‐ and heterochiral associations were found. Besides T_m measurements, detection of heterochiral complexes was unambiguously confirmed by LC‐MS studies. Interestingly, circular dichroism measurements of the most stable duplexes suggested that L ‐HNA form left‐handed helices with both D and L oligonucleotides.     

The 5‐Methylisocytosine β‐D‐Ribopyranosyl (4′ → 2′)‐Oligonucleotide

In the context of Eschenmoser's work on pyranosyl‐RNA (‘p‐RNA’), we investigated the synthesis and base‐pairing properties of the 5‐methylisocytidine derivative. The previously determined clear‐cut restrictions of base‐pairing modes of p‐RNA had led to the expectation that a 5‐methylisocytosine β‐D ‐ribopyranosyl (= D ‐pr(^MeisoC)) based (4′ → 2′)‐oligonucleotide would pair inter alia with D ‐pr(isoG) and L ‐pr(G) based oligonucleotides (D ‐pr and L ‐pr = pyranose form of D ‐ and L ‐ribose, resp.). Remarkably, we could not observe pairing with the D ‐pr(isoG) oligonucleotide but only with the L ‐pr(G) oligonucleotide. Our interpretation concludes that this – at first hand surprising – observation is caused by a change in the nucleosidic torsion angle specific for isoC.     

A Direct Synthesis of Nucleoside Analogs Homologated at the 3′‐ and 5′‐Positions

A new route is presented to prepare analogs of nucleosides homologated at the 3′‐ and 5′‐positions. This route, applicable to both the D ‐ and L ‐enantiomeric forms, is suitable for the preparation of monomeric bis‐homonucleosides needed for the synthesis of oligonucleotide analogs. It begins with the known monobenzyl ether 3 of pent‐2‐yne‐1,5‐diol, which is reduced to alkenol 4 . Sharpless asymmetric epoxidation of 4 , followed by opening of the epoxide 5 with allylmagnesium bromide, gives a mixture of diols 6 and 7 . Protection of the primary alcohol as a silyl ether followed by treatment with OsO₄, NaIO₄, and mild acid in MeOH, followed by reduction, yields (2R,3R) {{[(tert‐butyl)diphenylsilyl]oxy}methyl}tetrahydro‐2‐(2‐hydroxyethyl)‐5‐methoxyfuran (=methyl 3‐{{[(tert‐butyl)diphenylsilyl]oxy}methyl}‐2,3,5‐trideoxy‐α/β‐D ‐erythro‐hexafuranoside; 10 ) (Scheme 1). Protected nucleobases are added to this skeleton with the aid of trimethylsilyl triflate (Scheme 2). The o‐toluoyl (2‐MeC₆H₄CO) and p‐anisoyl (4‐MeOC₆H₄CO) groups were used to protect the exocyclic amino group of cytosine. The bis‐homonucleoside analogs 11 and 14a are then converted to monothiol derivatives suitable for coupling (Schemes 3 and 4) to oligonucleotide analogs with bridging S‐atoms. This synthesis replaces a much longer synthesis for analogous nucleoside analogs that begins with diacetoneglucose (=1,2 : 5,6‐di‐O‐isopropylideneglucose), with the stereogenic centers in the final products derived from the Sharpless asymmetric epoxidation. The new route is useful for large‐scale synthesis of these building blocks for the synthesis of oligonucleotide analogs.     

Synthesis of (5′S)‐5′‐C‐Alkyl‐2′‐deoxynucleosides

We describe the synthesis of (5′S)‐5′‐C‐butylthymidine ( 5a ), of the (5′S)‐5′‐C‐butyl‐ and the (5′S)‐5′‐C‐isopentyl derivatives 16a and 16b of 2′‐deoxy‐5‐methylcytidine, as well as of the corresponding cyanoethyl phosphoramidites 9a , b and 14a , b , respectively. Starting from thymidin‐5′‐al 1 , the alkyl chain at C(5′) is introduced via Wittig chemistry to selectively yield the (Z)‐olefin derivatives 3a and 3b (Scheme 2). The secondary OH function at C(5′) is then introduced by epoxidation followed by regioselective reduction of the epoxy derivatives 4a and 4b with diisobutylaluminium hydride. In the latter step, a kinetic resolution of the diastereoisomer mixture 4a and 4b occurs, yielding the alkylated nucleoside 2a and 2b , respectively, with (5′S)‐configuration in high diastereoisomer purity (de=94%). The corresponding 2′‐deoxy‐5‐methylcytidine derivatives are obtained from the protected 5′‐alkylated thymidine derivatives 7a and 7b via known base interconversion processes in excellent yields (Scheme 3). Application of the same strategy to the purine nucleoside 2′‐deoxyadenine to obtain 5′‐C‐butyl‐2′‐deoxyadenosine 25 proved to be difficult due to the sensitivity of the purine base to hydride‐based reducing agents (Scheme 4).     

Oligonucleotides Containing 7‐Deaza‐2′‐deoxyxanthosine: Synthesis,Base Protection,and Base‐Pair Stability

Oligonucleotides incorporating 7‐deaza‐2′‐deoxyxanthosine ( 3 ) and 2′‐deoxyxanthosine ( 1 ) were prepared by solid‐phase synthesis using the phosphoramidites 6 – 9 and 16 which were protected with allyl, diphenylcarbamoyl, or 2‐(4‐nitrophenyl)ethyl groups. Among the various groups, only the 2‐(4‐nitrophenyl)ethyl group was applicable to 7‐deazaxanthine protection being removed with 1,8‐diazabicyclo[5.4.0]undec‐7‐ene (DBU) by β‐elimination, while the deprotection of the allyl residue with Pd⁰ catalyst or the diphenylcarbamoyl group with ammonia failed. Contrarily, the allyl group was found to be an excellent protecting group for 2′‐deoxyxanthosine ( 1 ). The base pairing of nucleoside 3 with the four canonical DNA constituents as well as with 3‐bromo‐1‐(2‐deoxy‐β‐D ‐erythro‐pentofuranosyl)‐1H‐pyrazolo[3,4‐d]pyrimidine‐4,6‐diamine ( 4 ) within the 12‐mer duplexes was studied, showing that 7‐deaza‐2′‐deoxyxanthosine ( 3 ) has the same universal base‐pairing properties as 2′‐deoxyxanthosine ( 1 ). Contrary to the latter, it is extremely stable at the N‐glycosylic bond, while compound 1 is easily hydrolyzed under slightly acidic conditions. Due to the pK_a values 5.7 ( 1 ) and 6.7 ( 3 ), both compounds form monoanions under neutral conditions (95% for 1 ; 65% for 3 ). Although both compounds form monoanions at pH 7.0, pH‐dependent T_m measurements showed that the base‐pair stability of 7‐deaza‐2′‐deoxyxanthosine ( 3 ) with dT is pH‐independent. This indicates that the 2‐oxo group is not involved in base‐pair formation.     

Nucleotides,Part LXV,Synthesis of 2′‐Deoxyribonucleoside 5′‐Phosphoramidites: New Building Blocks for the Inverse (5′‐3′)‐Oligonucleotide Approach

The syntheses of the 3′‐O‐(4,4′‐dimethoxytrityl)‐protected 5′‐phosphoramidites 25 – 28 and 5′‐(hydrogen succinates) 29 – 32 , which can be used as monomeric building blocks for the inverse (5′‐3′)‐oligodeoxyribonucleotide synthesis are described (Scheme). These activated nucleosides and nucleotides were obtained by two slightly different four‐step syntheses starting with the base‐protected nucleosides 13 – 20 . For the protection of the aglycon residues, the well‐established 2‐(4‐nitrophenyl)ethyl (npe) and [2‐(4‐nitrophenyl)ethoxy]carbonyl (npeoc) groups were used. The assembly of the oligonucleotides required a slightly increased coupling time of 3 min in application of the common protocol (see Table 1). The use of pyridinium hydrochloride as an activator (instead of 1H‐tetrazole) resulted in an extremely shorter activation time of 30 seconds. We established the efficiency of this inverse strategy by the synthesis of the oligonucleotide 3′‐conjugates 33 and 34 which carry lipophilic caps derived from cholesterol and vitamin E, respectively, as well as by the formation of (3′‐3′)‐ and (5′‐5′)‐internucleotide linkages (see Table 2).     

Total Synthesis of the Bicyclic Depsipeptide HDAC Inhibitors Spiruchostatins A and B, 5′′‐epi‐Spiruchostatin B,FK228 (FR901228) and Preliminary Evaluation of Their Biological Activity

The bicyclic depsipeptide histone deacetylase (HDAC) inhibitors spiruchostatins A and B, 5′′‐epi‐spiruchostatin B and FK228 were efficiently synthesized in a convergent and unified manner. The synthetic method involved the following crucial steps: i) a Julia–Kocienski olefination of a 1,3‐propanediol‐derived sulfone and a L ‐ or D ‐malic acid‐derived aldehyde to access the most synthetically challenging unit, (3S or 3R,4E)‐3‐hydroxy‐7‐mercaptohept‐4‐enoic acid, present in a D ‐alanine‐ or D ‐valine‐containing segment; ii) a condensation of a D ‐valine‐D ‐cysteine‐ or D ‐allo‐isoleucine‐D ‐cysteine‐containing segment with a D ‐alanine‐ or D ‐valine‐containing segment to directly assemble the corresponding seco‐acids; and iii) a macrocyclization of a seco‐acid using the Shiina method or the Mitsunobu method to construct the requisite 15‐ or 16‐membered macrolactone. The present synthesis has established the C5′′ stereochemistry of spiruchostatin B. In addition, HDAC inhibitory assay and the cell‐growth inhibition analysis of the synthesized depsipeptides determined the order of their potency and revealed some novel aspects of structure–activity relationships. It was also found that unnatural 5′′‐epi‐spiruchostatin B shows extremely high selectivity (ca. 1600‐fold) for class I HDAC1 (IC₅₀=2.4 nM ) over class II HDAC6 (IC₅₀=3900 nM ) with potent cell‐growth‐inhibitory activity at nanomolar levels of IC₅₀ values.     

Cu(I)‐Catalyzed Efficient Synthesis of 2′‐Triazolo‐nucleoside Conjugates

A small library of thirty‐two 2′‐triazolyl uridine and 2′‐triazolyl‐5‐methyluridine has been synthesized by Cu(I)‐catalyzed condensation of 2′‐azido‐2′‐deoxyuridine and 2′‐azido‐2′‐deoxy‐5‐methyluridine with different alkynes and aryl propargyl ethers in almost quantitative yields. Triazolo‐nucleoside conjugates, which can be evaluated for different biological activity for suitable drug development, were unambiguously identified on the basis of ¹H NMR, ¹³C NMR, IR, and HRMS data analysis. These compounds have been synthesized for the first time and have not been reported in the literature earlier.     

The Synthesis of Methylated,Phosphorylated, and Phosphonated 3′‐Aminoacyl‐tRNASec Mimics

The twenty first amino acid, selenocysteine (Sec), is the only amino acid that is synthesized on its cognate transfer RNA (tRNA^Sec) in all domains of life. The multistep pathway involves O‐phosphoseryl‐tRNA:selenocysteinyl‐tRNA synthase (SepSecS), an enzyme that catalyzes the terminal chemical reaction during which the phosphoseryl–tRNA^Sec intermediate is converted into selenocysteinyl‐tRNA^Sec. The SepSecS architecture and the mode of tRNA^Sec recognition have been recently determined at atomic resolution. The crystal structure provided valuable insights that gave rise to mechanistic proposals that could not be validated because of the lack of appropriate molecular probes. To further improve our understanding of the mechanism of the biosynthesis of selenocysteine in general and the mechanism of SepSecS in particular, stable tRNA^Sec substrates carrying aminoacyl moieties that mimic particular reaction intermediates are needed. Here, we report on the accurate synthesis of methylated, phosphorylated, and phosphonated serinyl‐derived tRNA^Sec mimics that contain a hydrolysis‐resistant ribose 3′‐amide linkage instead of the natural ester bond. The procedures introduced allow for efficient site‐specific methylation and/or phosphorylation directly on the solid support utilized in the automated RNA synthesis. For the preparation of (S)‐2‐amino‐4‐phosphonobutyric acid–oligoribonucleotide conjugates, a separate solid support was generated. Furthermore, we developed a three‐strand enzymatic ligation protocol to obtain the corresponding full‐length tRNA^Sec derivatives. Finally, we developed an electrophoretic mobility shift assay (EMSA) for rapid, qualitative characterization of the SepSecS‐tRNA interactions. The novel tRNA^Sec mimics are promising candidates for further elucidation of the biosynthesis of selenocysteine by X‐ray crystallography and other biochemical approaches, and could be attractive for similar studies on other tRNA‐dependent enzymes.     

Synthesis and Anti‐HIV Activity of Triazolo‐Fused 3′,5′‐Cyclic Nucleoside Analogues Derived from an Intramolecular Huisgen 1,3‐Dipolar Cycloaddition

Triazolo‐fused 3′,5′‐cyclic nucleoside analogues were synthesized by an intramolecular 1,3‐dipolar cycloaddition of nucleoside‐derived azido‐alkynes in a regio‐ and stereospecific manner. The thymine nucleoside base in these target compounds was transformed successfully into the corresponding 5‐methylcytosine component. The synthesized compounds were examined in a MAGI assay for exploring the anti‐HIV activity and in a H9 T lymphocytes assay for measuring the cell toxicity.     

Pentopyranosyl Oligonucleotide Systems,Communication No. 10 , The α‐L‐Lyxopyranosyl‐(4′→2′)‐oligonucleotide System

To determine whether the remarkable chemical properties of the pyranosyl isomer of RNA as an informational Watson‐Crick base‐pairing system are unique to the pentopyranosyl‐(4′→2′)‐oligonucleotide isomer derived from the RNA‐building block D ‐ribose, studies on the entire family of diastereoisomeric pyranosyl‐(4′→2′)‐oligonucleotide systems deriving from D ‐ribose, L ‐lyxose, D ‐xylose, and L ‐arabinose were carried out. The result of these extended studies is unambiguous: not only pyranosyl‐RNA, but all members of the pentopyranosyl‐(4′→2′)‐oligonucleotide family are highly efficient Watson‐Crick base‐pairing systems. Their synthesis and pairing properties will be described in a series of publications in this journal. The present paper describes the α‐L ‐lyxopyranosyl‐(4′→2′)‐system.     

Enzymatic Synthesis of Nucleoside‐5′‐O‐(1‐thiophosphates) and (SP)‐Adenosine‐5′‐O‐(1‐thiotriphosphate)

Treatment of adenosine with PSCl₃ in trimethyl phosphate gave, after ion‐exchange chromatography, adenosine‐5′‐O‐monophosphate (AMP; 28%) and adenosine‐5′‐O‐monothiophosphate (AMPS; 48%). AMPS was studied as a thiophosphate residue donor in an enzymatic transphosphorylation with nucleoside phosphotransferase (NPase) of the whole cells of Erwinia herbicola. As exemplified by a number of natural and sugar‐ and base‐modified nucleosides, it was demonstrated that NPase of the whole cells of Erwinia herbicola catalyzes the transfer of both thiophosphate and phosphate residues with a similar efficiency. An incubation of AMPS in a phosphorylating extract of Saccharomyces cerevisiae (K‐phosphate buffer (0.3 M , pH 7.0); 3% glucose; 15 mM MgCl₂; 28°, 8 h), followed by ion‐exchange column chromatography afforded AMP (8%), AMPS (recovered, 23%), ATP (11%), and (S_P)‐adenosine‐5′‐O‐(1‐thiotriphosphate) ((S_P)‐ATPαS); (total yield 37%; 48% based on the consumed AMPS). For comparison of physicochemical properties, adenosine was chemically transformed into ATPαS as a mixture of the (S_P) (53%) and (R_P) (44%) diastereoisomers.     

Synthesis of Potentially Antiviral 2′,3′‐Dideoxy‐2′‐fluoro‐3′‐(hydroxyamino)nucleosides

A series of novel 3′‐(alkyl(hydroxy)amino)‐2′‐fluoronucleoside analogs were prepared via conjugate addition of N‐methylhydroxylamine to various 2‐fluorobutenolides. The adducts 13a and 16 were obtained as single isomers under absolute control of stereochemistry. The crucial N‐demethylation of 23 – 25 was readily achieved by means of DDQ oxidation, followed by nitrone/oxime exchange reaction. By this procedure, a variety of alkyl groups could be efficiently introduced at the 3′‐N‐atom of the nucleoside analogs, some of which might display potentially interesting anti‐HIV properties.