Conformational analysis of the B and Z forms of the d(m⁵C‐G)₃ and d(br⁵C‐G)₃ hexamers in solution

Abstract

The B and the Z forms of the DNA hexamers d(m5C-G)3 and d(b~-%-G)~ were investigated by means of NMR spectroscopy. It is demonstrated that the low-salt form of d(m5C-G)3 is a B DNA structure. The form, which becomes increasingly predominant when increasing amounts of MgC12 and/or methanol are added to the solution, has Z DNA characteristics. It is shown that the major geometrical features of the Z form of d(mSC-G)3 in the crystal structure are maintained in solution, with the dC residues S sugar conformation, y ' and the base in the anti orientation and the dG residues N (except the 3'-terminal residue), yf and syn. Neither the Z form of the methylated nor that of the brominated compound resembles the Z form, in which the deoxy guanosine sugar rings adopt a Cl'-exo conformation. Substitution of m5C by br5C causes no perceptible conformational changes in either the B or in the Z forms.The polymorphism exhibited by the DNA double helix [I, 21 has fascinated investigators from many fields of research for a long time. It is becoming clear that the character of the bases (purine R or pyrimidine Y) and their sequence determine the conformational possibilities of a given DNA species. For example, the peculiar inversion of the CD spectrum of poly d(G,C) . poly d(G,C) upon addition of salt [3] and/or alcohols [4,5] appeared limited to polymer duplex species that contain alternating dC-dG sequences and few others (dA-dC, dT-dG). This inversion was later shown to be a consequence of the conversion of the right-handed B DNA form into the lefthanded Z DNA species 16, 71. Detailed NMR and CD studies have revealed that Z DNA can be stabilized chemically in several ways. For instance, methylation [8,9] or bromination [lo-121 at the C-5 position of cytosine, or bromination at the C-8 position of guanine [13, 141 shifts the equilibrium between the B and Z forms towards the latter DNA species.Up till now information concerning the detailed geometry of Z DNA has come almost exclusively from X-ray crystallography 1151. In contrast to the classical B form, Z DNA has only a minor groove, which resembles the minor groove of B DNA. The major groove of B DNA constitutes an outer surface of Z DNA and exposes the cytosine C-5 and guanine N-7 and C-8 atoms. The major geometrical characteristics of Z DNA in the crystal can be summarized as follows: (a) the pyrimidines ( Y ) adopt a standard conformation with S-type sugar pucker, base anti (Fig. l ) and g + orientation about torsion angle y (Fig. 2); (b) the purines (R) prefer This paper is number 46 in a series Nucleic Acid Constituents from this laboratory; for part 45 see the preceding paper in this journal.Abbreviations. Me,NCI, tetramethylammonium chloride; NOE, nuclear Overhauser effect; NOESY, two-dimensional nuclear Overhauser spectroscopy; COSY, correlated spectroscopy. N-type sugars, base syn (Fig. I) and a g' rotamer about y ; (c) the phosphodiester bonds adopt the (+, a+ rotamer combination for the Y-R step and usually l -, a' for the R-Y step. It should be mentioned here that spe...

The B and Z forms of the d(m5C‐G)3 and d(br5C‐G)3 hexamers in solution

“…The conformational analysis of the respective B and Z forms will be reported in a companion paper [15]. There it will be shown that the interpretation of the NMR data indeed strongly supports these designations.…”

“…The COSY spectrum of the BjZ mixture was analysed as [15], it should be noted that these upfield shifts accord with the known crystal structure [27] of Z DNA, Fig.8. It is seen that the (Fig.…”

“…The line-broadening, as observed in Fig.2B and D, appears to be caused by the aggregation of the DNA fragment. The effect of the solvent (i. e. the addition of methanol) on the conformation of the B species is discussed in a companion paper [15].…”

“…It should be noted that the proton chemical shifts of both B and Z forms of the methylated and brominated analogues are virtually equal, except for the H-6 resonances, which are shifted downfield in the bromine compound (0.6 ppm and 0.4 ppm in the B and the Z forms respectively). More detailed chemical shift considerations will be given in a companion paper [15].…”

“…d(C-G), type necessitates a careful assignment and analysis of the NMR spectra of each stable molecular species. In the present paper we will loosely designate the low-salt form in aqueous solution as B DNA and the structure which becomes increasingly predominant when increasing amounts of C2H302H and/or MgC12 are added to the solution will be designated Z DNA.The conformational analysis of the respective B and Z forms will be reported in a companion paper [15]. There it will be shown that the interpretation of the NMR data indeed strongly supports these designations.…”