Preparation and Properties of an Analogue of Poly(A) and Poly(G): Poly(isoguanylic acid)

Abstract

Isoguanosine-5''-pyrophosphate, in the presence of an oligonucleotide primer, was polymerized by Escherichia coli polynucleotide phosphorylase (EC 2.7.7.8) under conditions analogous to those required for polymerization of 5''-GMP. The resulting poly(isoguanylic acid), poly(isoG), was a multistranded helix with a stability considerably higher than that of poly(G), and fully resistant to various nucleolytic enzymes. The polymer exhibited a 2-step temperature transition profile in moderately alkaline propylene glycol. Alkaline titration in aqueous medium, by UV and circular dichroism spectroscopy, showed 2 clearly defined transitions, the 2nd of which was fully cooperative. The accompanying changes in sedimentation constants were consistent with a structure for poly(isoG) of a 4-stranded helix, like neutral poly(G). In acid medium, spectral and potentiometric titrations demonstrated the existence of more than 1 transition in the pH range 6.1.2, with accompanying protonation of the isoguanosine residues. In neutral medium the polymer formed no complexes with other potentially complementary homopolymers. In acid medium the protonated form of poly(isoG) did form a triple-stranded complex with poly(I), namely, 2poly(I) .cntdot. poly(isoG)+. Possible structures are formulated for the neutral and protonated forms of poly(isoG) which account for the 2-step thermal transition in alkaline propylene glycol and on alkaline titration in aqueous medium. The nature of the protonated form, and its complex with poly(I) is also discussed.