SOLUTION STRUCTURE OF THE CPG CONTAINING D(CTTCGAAG)(2) OLIGONUCLEOTIDE - NMR DATA AND ENERGY CALCULATIONS ARE COMPATIBLE WITH A BI BII EQUILIBRIUM AT CPG/

We report the analysis of the solution structure of the DNA duplex d(C TTCGAAG)(2) compared to that of d(CATCGATG)(2), the two oligonucleotid es being related by the permutation of residues 2 and 7. An earlier st udy has demonstrated the malleability of CpG in the tetrad TCGA of d(C ATCGATG)(2) [Lefebvre et al. (1995) Biochemistry 34, 12019-12028]. Con formations of d(CTTCGAAG)(2) were evaluated by (a) two-dimensional NMR , including proton and phosphorus experiments, (b) adiabatic mapping o f the conformational space, (c) restrained molecular mechanics underta ken with sugar phase angle, epsilon-zeta difference angle, and NOE dis tances as input, and (d) back-calculation-refinement against NOE spect ra at various mixing times. d(CTTCGAAG)(2) like d(CATCGATG)(2) exhibit s a B-DNA conformation. However, significant differences are noted bet ween the two oligonucleotides, extending up to the central CpG step, a lthough this step resides in the same TCGA tetrad in both sequences. I n structures obtained with refined NMR data, CpG adopts, for instance, a greater twist and a higher guanine phase within d(CTTCGAAG)(2) comp ared to d(CATCGATG)(2). In the former oligonucleotide, the structure o f CpG resembles strikingly that found in the ACGT tetrad of the cAMP r esponsive element [Mauffret et al. (1992) J. Mol. Biol. 227, 852-875]. Moreover, two conformers with CpG either in the BII state (epsilon, z eta = g(-), t) Or in the BI state (epsilon, zeta = t, g(-)) are found equally stable for d(CTTCGAAG)(2) The energy barrier from BI to BII co mes to only 5.7 kcal/mol, and the path of the transition is very short . When calculations on d(CTTCGAAG)2 are performed taking the BI/BII eq uilibrium into account, the agreement with both the H-1 and P-31 data is found better than in the case with a single conformation taken alon e. The BI/BII equilibrium may also occur in d(CATCGATG)(2), but the am ount of BII conformer is now found weaker compared to its analogue. Th e ability of the CpG phosphate groups to adopt the Bn. conformation co uld provide a satisfying explanation for the high mutation rates obser ved at these sites.