Georg Becher

STEREOELECTRONIC EFFECTS OF MODIFIED PURINES ON THE SUGAR CONFORMATION OF NUCLEOSIDES AND FLUORESCENCE PROPERTIES

Abstract Conformational analyses of the sugar moieties of a series base-modified purine-2′-deoxynucleosides on the basis of vicinal [1H,1H] coupling constants is presented (PSEUROT 6.2) Fluorescence data of several 7-deaza- and 8-azapurine 2′-deoxynucleosides are given.

The High‐Anti Conformation of 7‐Halogenated 8‐Aza‐7‐deaza‐2′‐deoxy‐guanosines: A Study of the Influence of Modified Bases on the Sugar Structure of Nucleosides

Introduction. – The conformational flexibility of nucleic acids can be more complexthan that of peptides because the sugar-phosphate backbone conformation is descibedby five single-bond rotations in addition to five sugar torsions [1a]. It is expected that inmany cases, the conformational changes among nucleoside or nucleotide structuresobey a certain conformational pathway. This means that the movement of one torsion islinearly coupled with another one [2][3]. In general, the molecular structures ofnucleosides and nucleotides have been regarded as conformationally ˝rigid˛ [4]probably due to the small variances of torsion angles. Besides gauche and anomericeffects [5], important determinants of the overall structure of a nucleoside are the stericand stereoelectronic effects of the nucleobases [6]. In a series of influential reports, atfirst Gassen and coworkers [7], and later Chattopadhyaya and coworkers [8] havedemonstrated the importance of such effects on both the structure of a nucleoside aswell as on the secondary structure of a DNA molecule. Recently, our interest wasfocussed on the stereoelectronic influence of modified nucleobases – and here inparticular of aza- and deazapurines – on the structure and stability of oligonucleotidescontaining such compounds [9–13]. We have observed that the incorporation of 8-aza-7-deazapurine 2’-deoxynucleotides into oligodeoxynucleotides exerts an extraordinaryinfluence on their duplex stability which is significantly enhanced over that of theparent unmodified oligomers [14–16]. This may be traced to an altered secondarystructure of the base-modified oligomers with probably enhanced stacking interactions

8-Aza-7-deazapurine DNA: Synthesis and Duplex Stability of Oligonucleotides Containing 7-Substituted Bases

Abstract The 7-substituted 8-aza-7-deazapurine phosphoramidites 1a – 3c as well as the phosphoramidite 4a were synthesized. In comparison to the parent purine oligonucleotide duplexes, the 7-substituted 8-aza-7-deazapurine residues lead to a significant duplex stabilization.

Synthesis, Base Pairing, and Fluorescence Properties of Oligonucleotides Containing 1H-Pyrazolo[3,4-d]pyrimidin-6-amine (8-Aza-7-deazapurin-2-amine) as an Analogue of Purin-2-amine

The synthesis of the N9- and N8-(β-D-2′-deoxyribonucleosides) 2 and 10, respectively, of 8-aza-7-deazapurin-2-amine (=1H-pyrazolo[3,4-d]pyrimidin-6-amine) is described. The fluorescence properties and the stability of the N-glycosylic bond of 2 were determined and compared with those of the 2′-deoxyribonucleosides 1 and 3 of purin-2-amine and 7-deazapurin-2-amine respectively. From the nucleoside 2, the phosphoramidite 14 was prepared, and oligonucleotides were synthesized. Duplexes containing compound 1 or 2 are slightly less stable than those containing 2′-deoxyadenosine, while their CD spectra are rather different. The fluorescence of the nucleosides is strongly quenched (>95%) in single-stranded as well as in duplex DNA. The residual fluorescence was used to determine the melting profiles, which gave Tm values similar to those determined from the UV melting curves.

Unexpected Dehalogenation of 3-Bromopyrazolo[3,4-d]pyrimidine Nucleosides During Nucleobase-Anion Glycosylation

Abstract The anion-glycosylation (KOH, MeCN, TDA-1) of 3-bromopyrazolo[3,4-d]-pyrimidines 4a and 4b with 2-deoxy-3,5-di-O-(p-toluoyl)-α-D-erythro-pentofuranosyl chloride (5) furnishes the regioisomeric N′-β-D-2′-deoxyribonucleosides 6a and 6b together with the dehalogenated N2-regioisomers 8a and 8b, stereoselectively. The dehalogenation takes place after the glycosylation and results from the sensitivity of the N-2 nucleosides toward aqueous base. An addition/elimination mechanism is suggested for the dehalogenation reaction.

Base-Modified Oligonucleotides With Increased Duplex Stability

: Oligonucleotides incorporating 8-aza-7-dazapurines (pyrazolo[3,4-d]pyrimidines) were synthesized. The corresponding nucleosides were prepared and were converted into phosphoramidites. The oligonucleotide duplex stability was studied and was compared to that of the parent compounds containing the canonical purine nucleosides. The presence of 7-halogeno or 7-alkynyl substituents increases the duplex stability significantly.

Stabilisation of duplex DNA by 7-halogenated 8-aza-7-deazaguanines

Oligonucleotides containing 7-halogenated 8-aza-7-deaza-2′-deoxyguanosine (c7z8Gd) derivatives such as d(Br7c7z8 G-C)4 8 (Tm = 88 °C) and d(I7c7z8 G-C)4 9 (Tm = 84 °C) are significantly more stable than d(G-C)4 5 (Tm = 59 °C).

Pyrazolo[3,4-d]pyrimidine nucleic acids: adjustment of the dA-dT to the dG-dC base pair stability.

Abstract The pyrazolo[3,4-d]pyrimidine-4,6-diamine nucleosides 2b-d stabilize the dA-dT base pair significantly when the dA-residue is replaced. Oligonucleotide duplexes incorporating 2b-d show a 4–6°C T m increase per modification. The 7-bromo compound 2b harmonizes the stability of the dA-dT vs. the dG-dC pair. According to this the stability of such duplexes depends no longer on the base pair composition of a DNA molecule.

Fluorescence properties and base pair stability of oligonucleotides containing 8-aza-7-deaza-2'-deoxyisoinosine or 2'-deoxyisoinosine.

Abstract The fluorescence and the base pairing properties of 8-aza-7-deaza-2′-deoxyisoinosine (1) are described and compared with those of 2′-deoxyisoinosine (2). The corresponding phosphoramidites (11,12) are synthesized using the diphenyl-carbamoyl (DPC) residue for the 2-oxo group protection. The nucleosides 1 and 2 base pair with 2′-deoxy-5-methylisocytidine in DNA duplexes with antiparallel chain orientation and with 2′-deoxycytidine in a parallel DNA. These base pairs are less stable than the canonical dA-dT pair and that of 2′-deoxyinosine (4) with 2′-deoxycytidine. The fluorescence of the nucleosides 1 and 2 is quenched (∼95%) in duplex DNA. The residual fluorescence is used to determine the Tm-values, which are found to be the same as determined UV-spectrophotometrically.