The solution structure of the circular trinucleotide cr(GpGpGp) determined by NMR and molecular mechanics calculation

Abstract

The 3′-5′ circular trinucleotide cr(GpGpGp) was studied by means of 1D and 2D high resolution NMR techniques and molecular mechanics calculations. Analysis of the J-couplings, obtained from the 1H and 13C-NMR spectra, allowed the determination of the conformation of the sugar rings and of the 'circular' phosphate backbone. In the course of the investigations it was found that the Karplus-equation most recently parametrized for the CCOP J-coupling constants could not account for the measured J(C4′P) of 11.1 Hz and new parametrization for both HCOP and CCOP coupling constants is therefore presented. Subsequent analysis of the coupling constants yielded 'fixed' values for the torsion angles β and δ (with β = 178° and δ = 139°). The value of the latter angle corresponds to an S-type sugar conformation. The torsion angles λ and ε are involved in a rapid equilibrium in which they are converted between the gauche( + ) and trans and between the trans and gauche( - ) domain respectively. We show that the occurrence of e in the gauche(-) domain necessitates S-type sugar conformations. Given the aforementioned values for β, λ, δ and ε the ring closure constraints for the ring, formed by the phosphate backbone can only be fulfilled if α and ζ adopt some special values. After energy minimization with the CHARMm force field only two combinations of α and ζ result in energetically favourable structures, i.e. the combination α(t)/ζ(g-) in case λ is in a gauche( + ) and ε is in a trans conformation, and the combination α(t)/ζ (g +) for the combination λ(t)/ε(g - ) . The results are discussed in relation to earlier findings obtained for cd(ApAp) and cr(GpGp), the latter molecule being a regulator of the synthesis of cellulose in Acetobacter xylinum. © 1994 Oxford University Press.