Stacking interactions of ApA analogues with modified backbones

Abstract

CD spectra have been measured as a function of temperature for a number of ApA analogues with modified backbones. Oligonucleotides with these modified backbones are being used as antisense agents having potential as viral therapeutics. Results of these studies show that when a carbonyl is substituted for the phosphate to produce an uncharged backbone, the analogues that have either sugar or morpholino substitution do not stack. In contrast, when a morpholino group is substituted for the sugar and the phosphate is modified so as to be uncharged, there is strong base stacking. Stacking interactions in the phosphorus-linked morpholino analogues are at least as strong as those found in d(ApA). The stacking interactions in ApA are weak by comparison. Singular value decomposition demonstrates that the stacking is two state, and Taylor series decomposition yields a coefficient that measures base stacking interactions. The van't Hoff equation is applied to the base stacking coefficient from the Taylor series fitting to give thermodynamic parameters.

Morpholino, siRNA, and S-DNA Compared: Impact of Structure and Mechanism of Action on Off-Target Effects and Sequence Specificity

“…2) Relative to S-DNAs, Morpholinos have a much higher affinity for their complementary RNA sequences, and in fact Morpholinos bind RNA with a higher affinity than DNA binds RNA and much higher affinity than S-DNA for RNA [21]. 3) Probably because of their exceptional base stacking properties [24] Morpholinos show excellent solubility in aqueous solution (typically in excess of 100 mg/ml), in sharp contrast to other non-ionic structural types which are generally plagued by poor aqueous solubilities (typically several hundred fold lower than for Morpholinos) [14]. 4) Probably because of their highly unnatural backbone structure and lack of charge on the backbone, Morpholinos appear not to interact to any significant extent with proteins.…”

Morpholino Antisense Oligomers: Design, Preparation, and Properties

“…Conventional wisdom in the antisense field is that non-ionic antisense oligos invariably show poor water solubility. In this regard, it is interesting that a Morpholino dimer containing a rigid carbamate linkage shows little or no base stacking (Kang et al, 1992), and in the absence of special solubilizing groups, Morpholino oligomers containing such carbamate linkages are quite insoluble in aqueous solutions (Stirchak et al, 1989). In contrast, phosphorodiamidate-linked Morpholino oligos of the type shown in Figure 2 show excellent base stacking (Kang et al, 1992) and are several orders of magnitude more soluble in aqueous solutions.…”

Using Morpholinos to Control Gene Expression

“…No crystal structure or high-resolution NMR structural analysis of phosphorodiamidate Morpholinos has been published. However, the study of a Morpholino phosphorodiamidate ApA dimer using circular dichroic spectroscopy showed stacking of bases in aqueous phosphate buffer (Kang et al, 1992). On the basis of molecular modeling, the bases of Morpholinos should stack in a fashion analogous to those of natural nucleic acids, allowing strong interactions with complementary nucleic acid sequences by Watson-Crick base pairing.…”