In the equilibrium between B-DNA and Z-DNA in poly(dC-dG), the [Co(NH3)6]3+ ion stabilizes the Z form 4 orders of magnitude more effectively than the Mg2+ ion. The structural basis of this difference is revealed in Z-DNA crystal structures of d(CpGpCpGpCpG) stabilized by either Na+/Mg2+ or Na+/Mg2+ plus [Co(NH3)6]3+. The crystals diffract X-rays to high resolution, and the structures were refined at 1.25 A. The [Co(NH3)6]3+ ion forms five hydrogen bonds onto the surface of Z-DNA, bonding to a guanine O6 and N7 as well as to a phosphate group in the ZII conformation. The Mg2+ ion binds through its hydration shell with up to three hydrogen bonds to guanine N7 and O6. Higher charge, specific fitting of more hydrogen bonds, and a more stable complex all contribute to the great effectiveness of [Co(NH3)6]3+ in stabilizing Z-DNA.

In left-handed Z-DNA, consecutive nucleotides along the chain alternate in the syn and anti conformations. Purine residues form the syn conformation readily and up to now all Z-DNA crystal structures have sequences of alternating purines and pyrimidines. However, we find that d(C-G-A-T-C-G) with the cytosines brominated or methylated on C-5 crystallizes as Z-DNA. The structure reveals thymines in syn and adenines in anti conformations. This suggests that Z-DNA may occur in sequences other than those with alternating purine-pyrimidine sequence. Double-helical DNA can exist both in right-handed and lefthanded forms. The structure of right-handed B-DNA has been known since 1953 and an atomic resolution single-crystal x-ray diffraction analysis in 1979 showed that the DNA hexamer d (CpGpCpGpCpG) [(dC-dG)31 forms a left-handed helix called Z-DNA (1). A variety of studies carried out on oligodeoxynucleotide single crystals (2-6) and on polynucleotides has shown that Z-DNA can form in sequences with alternations ofpurines and pyrimidines (see review in ref. 7). In the Z-DNA molecule, every other base adopts the syn conformation relative to the sugar, in contrast to B-DNA, where all of the bases are found in the anti conformation. Purine nucleosides can adopt the syn conformation as easily as they can adopt the anti conformation, while pyrimidine nucleosides do this less readily (8, 9). However, we do not know whether pyrimidines can adopt the syn conformation in DNA. It has been shown that negative supercoiling can stabilize Z-DNA formation in plasmids (10, 11). Studies of supercoiled plasmids suggested that segments can form Z-DNA without a strict alternation of purines and pyrimidines (12). Here we report that a DNA fragment with the sequence d(CpGpApTpCpG), where the cytosine residues are modified either by methylation or bromination on the C-5 position, crystallizes in the form of left-handed Z-DNA. Examination of the crystal structure reveals that the two central thymine residues adopt the syn conformation with intramolecular distances that are only slightly shorter than those seen with purine residues in the syn conformation. Two of the six residues in this molecule no longer maintain an alternation of purines and pyrimidines and still it forms Z-DNA. This suggests that segments of DNA may form left-handed Z-DNA without a strict adherence to the alternation of purines and pyrimidines. In addition, the presence of syn pyrimidines and anti purines in Z-DNA changes the external shape of the molecule. Crystallization and Structure SolutionThe oligonucleotides were synthesized by an improved phosphate triester method in which either 5-bromo or 5-methyl deoxycytidine nucleosides were used as the starting material (13). The purity of the oligomers was found to be >95% as judged by HPLC analysis. The crystallization mixture contained 2.3 mM oligonucleotide, 33 mM sodium cacodylate buffer (pH 7.0), 25 mM magnesium chloride, 25 mM cobalt hexamine trichloride, and 25 mM calcium chloride. Cobalt hexamine was used a...