2,6-Bis(functionalized) purines as metal-ion-binding surrogate nucleobases that enhance hybridization with unmodified 2′-O-methyl oligoribonucleotides

The capacity of three different purine bases, viz. 2,6-bis(3,5-dimethylpyrazol-1-yl)purine, 2-(3,5-dimethylpyrazol-1-yl)adenine and 2,6-bis(2-acetyl-1-methylhydrazino)purine, to form metal-ion mediated base pairs with the native nucleobases has been examined. For this purpose, ribonucleosides derived from these bases were incorporated into an intrastrand or a 3'-terminal position of short 2'-O-methyl oligoribonucleotides and the hybridization properties of these base modified oligomers in the absence and presence of three different metal ions (Cu2+, Zn2+ and Pd2+) were studied by UV- and CD-spectrometry. The first two bases were found to stabilize short oligonucleotide duplexes when incorporated into the 3'-termini of both strands, even in the absence of divalent metal ions but especially in the presence of Cu2+. The highest melting temperature determined for such a duplex was 71.8 degrees C, nearly 30 degrees C higher than the T-m of the respective solely Watson-Crick paired duplex. Despite the dramatic stabilizing effect of the terminal metallo-base pairs, these short modified oligonucleotides retained sequence-selectivity for the internal Watson-Crick base pairs: two internal mismatches dropped the melting temperature to 10-11 degrees C. In an internal position, only 2,6-bis(3,5-dimethylpyrazol-1-yl)purine, which in the absence of metal ions was destabilizing, exhibited metal-ion-dependent stabilization of duplex formation with unmodified 2'-O-methyl oligoribonucleotides. The melting temperature in the presence of Cu2+ was increased from 6 to 14 degrees C, depending on the identity of the opposite base.