9622 zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA Biochemistry zyxwvu 1992, 31, 9622-9628 Crystal and Molecular Structure of a DNA Fragment Containing a 2-Aminoadenine Modification: The Relationship between Conformation, Packing, and Hydration in Z-DNA Hexamerst Bohdan Schneider, Stephan L. Ginell, Roger Jones, Barbara Gaffney, and Helen M. Berman’ Department of Chemistry, Rutgers, The State University of New Jersey, New Brunswick, New Jersey 08903 Received April 28, 1992; Revised Manuscript Received July 16, 1992 ABSTRACT: The crystal and molecular structure of d(CGUA’CG)z (where A’ is 2-aminoadenine) has been determined and refined to an R factor of 13.8% for data 8.0-1.3 zyxwvu A. The structure is very similar to the original Z-DNA structures with the sequence d(CGCGCG)2 [Gessner, R. zyxw V., Frederick, C. A,, Quigley, G. J., Rich, A,, zyxwvutsrq & Wang, A. H.-J. (1989) J. Biol. Chem. zyxwv 264, 79211 and shows that the substitution of 2-aminoadenine-uracil base pairs in the two central steps is consistent with Z-DNA formation. In addition, we show how waters mediating intermolecular interactions may help to explain the ZI-ZII conformational pattern found in many Z-DNA structures. SinceZ-DNA was first described (Wang et al., 1979;Drew & Dickerson, 1981), it has provided fertile territory for exploring a variety of biological and chemical issues that underlie the diversity and flexibility of nucleic acid structure. The roles of sequence and chemical environment are among the many issues that have been explored in attempts to understand this unusual and unexpected conformation. A survey of the Z-DNA crystal structures (Table I) shows that, for hexamers, sequencescontainingalternating CG form Z-typeconformations. When UA (Geierstangeret al., 199l), TA (Wang et al., 1984), or AT (Wang et al., 1985) are substituted in the central step and are accompanied by 5-methylated cytosines in the flanking regions, Z-DNA structures are also formed. The presence of 2-aminoadenine thymine base pairs in the flanking regions is also consistent with Z-DNA formation (Coll et al., 1986). On the other hand, Raman spectroscopy measurements on crystalline d(CGTACG)2 have shown that this sequence forms B-type structures (Peticolas et al., 1989). In the d(CGUA’CG)2 structure reported here, we examine the question of whether Z-DNA forms when the central region contains 2-aminoad- enine-uracil base pairs. The hydration of Z-DNA structures has been the subject of considerable study (Zhou & Ho, 1990; Chevrier et al., 1986). The structure of d(CGUA’CG)* provides further opportunity to explore the interrelationships between con- formation, packing, and water structure in Z-DNA hexamers. EXPERIMENTALPROCEDURES Purified and lyophilized d(CGUA’CG)z (Gaffney et al., 1984) was dissolved in water to a concentration of approx- imately 6.0 mM. Crystals were grown at 4 OC by vapor diffusion of both sitting and hanging drops against a well consisting of water and 45-50% 2-methyl-2,4-pentanediol (MPD). Drops containing 1.8-2.0 mM DNA, 30 mM sodium cacodylate buffer (pH 7.0), 15 mM MgCl2,5 mM spermine tetrahydrochloride, and 20% MPD grew hexagonal rods in 2-4 weeks. +This work has been supported by the NIH (Grant GM21589 to H.M.B. andGM31483 to R.J.). TheNucleic AcidDatabaseissupported by Grant DIR NO12772 from the NSF. * Author to whom correspondence should be addressed. 0006-2960/92/043 1 -9622$03.00/0 Table I: Structures of Z-DNA Hexamers Compared to d(CGUA’CG)z“ structure description CGCGCG spermine/Mgz+ CGCGCG Mg2+ MeCGMCCGMCCG MCCGTAMFG MeCGUAMcCG/Cu2+ CA’CGTG CGCMCGCG MCCGUAMeCG CGCGCG/Cu2+ BrCGBrCGBrCG 29 1 K BrCGBrCGBrCG 310 K CGCGTJG CGCGCG spermine rms 0.12 0.20 0.35 0.37 0.41 0.34 0.33 0.43 0.24 0.33 0.34 0.50 0.46 refb 1 1 2 3 4 5 6 7 8 9 9 10 11 - code ZDFOOl ZDFOO2 ZDFB03 ZDFB06 ZDFB10 ZDFBl 1 ZDFB2l ZDFB24 ZDF028 ZDFB04 ZDFBO5 ZDFBl2 ZDF029 The column labeled ‘structure description” identifies a structure; ‘rms” lists root mean square deviations of a particular DNA structure with d(CGUA’CG)2. All except the phosphate oxygen atoms were superimposed, “ZI/ZII“ specifies the location of a ZII conformation in a sequence;‘ref‘ is the reference to a structure listed in the next footnote; ‘code” is the Nucleic Acid Database (Berman et al. 1992) code name of a structure. References: (1) ZDFOO1,ZDFOOl(Gwner et al., 1989); (2) ZDFB03 (Fujii et al., 1982); (3) ZDFB06 (Wang et al., 1984); (4) ZDFBlO (Geierstanger et al., 1991); (5) ZDFBll (Collet al., 1986);(6) ZDFB2l (Ginell et al., 1990); (7) ZDFB24 (Zhou & Ho, 1990); (8) ZDF028 (Kagawa et al., 1991); (9) ZDFB04, ZDFBO5 (Chevrier et al., 1986);(10) ZDFBlZ(Colletal., 1989);(11) ZDF029 (Eglietal., 1991). A residual electron density at the backbone between residues 8 and 9 has been described as a partially occupied ZII conformation. The crystal data were obtained at room temperature and showed that this sequence crystallizes in the identical space group and with cell dimensions similar to those found for other Z-DNA hexamers (Table 11). X-ray intensity data were collected using 0/28 scans on an Enraf-Nonius CAD4 diffractometer. The intensities were corrected for Lorentz, polarization, absorption, and decay with the Enraf-Nonius program package MOLEN (Fair, 1990). A total of 13 324 unique reflections were collected to a resolution of 1 .O A. Of those, 5790 were above 2 4 0 . Analysis of the data statistics showed that the effectiveresolution was 1.3Asince a significant drop was observed in the number of observed reflections above this resolution. Although thecrystal data of the structure reported here are similar to those of d(CGCGCG)* (Gessner et al., 1989), rotation and translation searches were conducted using zyxwvutsrqp 0 1992 American Chemical Society