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Application of multinuclear 2D NMR spectroscopy to metal-DNA interactions
NUCLEIC ACIDS
549
APPLICATION OF MULTINUCLEAR 2D NMR SPECTROSCOPY TO METAL-DNA INTERACTIONS
HO36
M. Iwamoto, K. A. Keating, Y. Xu, and L. G. Marzilli Department of Chemistry, Emory University, Atlanta, GA 30322, U.S.A.
A knowledge of the effects of metal species on DNA strncture is important in understanding the natural role of metal species (Mg2+, Zn2+) and in identifying the critical lesions formed by anticancer drugs (Pt(I1) drugs) and by toxic metal species (Cd 2+, Hg(I1) compounds). We have used multinuclear NMR spectroscopy to identify metal binding sites, even for labile metal species, and to assess structural changes [ 1,2]. Binding sites can be identified by studies of oligonucleotides of varying lengths after complete or nearly complete ‘H, l3C and/or 31P NMR signal assignments using COSY, NOESY, HMBC, HMQC, etc. Structural changes can be assessed in part by chemical shift changes in the signals of these nuclei as well as by measurement of the time dependence of the NOE cross-peaks. Such data are converted into solution structural information using distance geometry methods. Insight into the struture of these DNAs can be gained by examining oligonucleotide models by these NMR methods. Platination of oligonucleotides by Pt anticancer drugs can lead to single-stranded species, distorted duplexes, and hairpin structures. Single-stranded species are distorted and reveal characteristic changes in 3 lP and 13C NMR signals indicative of conformational changes in the sugar phosphate backbone of N7,N7 cross-linked Such distortions appear to persist in duplex species such as cis%$&2d(CTCCG*G*CCT)-d(GAGGCCGGA), where the asterisks indicate GN7 platination. A nearly complete assignment of all lH NMR signals has been made from the COSY and NOESY spectra of this non-self-complementary duplex. Few DNA duplexes treated with cis-Pt(NH&C12 have been assigned as completely. The duplex studied here exhibits a kink, bending towards the major groove, consistent with an N7,N7 intrastrand cross-link formed by cisPt(NH3)2. There is also evidence for more than one conformation of this duplex in solution; these conformations interconvert slowly on the NMR time scale. Also presented will be the structure of an unusually stable hairpin adduct formed by Pt(ethylenediamine)C12 with 5’-dAlT2G3G*4G*gTgA7C8C9CloAl lTl2-3’. This structure reveals that nucleotides Al-G4 and Cg-T12 form a double-stranded stem region that is similar to B-DNA. Unusual 1H and 31P chemical shifts gave indication of a grossly distorted loop region consisting of nucleotides G5-C8. The shift of the G4 H8 signal is unusually far upfield. Also, in both the duplex and hairpin structures, the sugars of the 5’ platinated G have been converted from an S to an N conformer. 1. 2.
X. Jia, G. Zon, and L.G. Marzilli, Znorg. Chem, 30,228 (1991). L. G. Marzilli, S. Mukundan, Jr., Y. Xu, G. Zon, A. Bergman, P. Yohannes, and M. D. Reily, in Platinum and Other Metal Coordination Compounds in Cancer Chemotherapy, Howell, S. B., Ed., Plenum Press, New York, 1991, pp. 101-114.
549
APPLICATION OF MULTINUCLEAR 2D NMR SPECTROSCOPY TO METAL-DNA INTERACTIONS
HO36
M. Iwamoto, K. A. Keating, Y. Xu, and L. G. Marzilli Department of Chemistry, Emory University, Atlanta, GA 30322, U.S.A.
A knowledge of the effects of metal species on DNA strncture is important in understanding the natural role of metal species (Mg2+, Zn2+) and in identifying the critical lesions formed by anticancer drugs (Pt(I1) drugs) and by toxic metal species (Cd 2+, Hg(I1) compounds). We have used multinuclear NMR spectroscopy to identify metal binding sites, even for labile metal species, and to assess structural changes [ 1,2]. Binding sites can be identified by studies of oligonucleotides of varying lengths after complete or nearly complete ‘H, l3C and/or 31P NMR signal assignments using COSY, NOESY, HMBC, HMQC, etc. Structural changes can be assessed in part by chemical shift changes in the signals of these nuclei as well as by measurement of the time dependence of the NOE cross-peaks. Such data are converted into solution structural information using distance geometry methods. Insight into the struture of these DNAs can be gained by examining oligonucleotide models by these NMR methods. Platination of oligonucleotides by Pt anticancer drugs can lead to single-stranded species, distorted duplexes, and hairpin structures. Single-stranded species are distorted and reveal characteristic changes in 3 lP and 13C NMR signals indicative of conformational changes in the sugar phosphate backbone of N7,N7 cross-linked Such distortions appear to persist in duplex species such as cis%$&2d(CTCCG*G*CCT)-d(GAGGCCGGA), where the asterisks indicate GN7 platination. A nearly complete assignment of all lH NMR signals has been made from the COSY and NOESY spectra of this non-self-complementary duplex. Few DNA duplexes treated with cis-Pt(NH&C12 have been assigned as completely. The duplex studied here exhibits a kink, bending towards the major groove, consistent with an N7,N7 intrastrand cross-link formed by cisPt(NH3)2. There is also evidence for more than one conformation of this duplex in solution; these conformations interconvert slowly on the NMR time scale. Also presented will be the structure of an unusually stable hairpin adduct formed by Pt(ethylenediamine)C12 with 5’-dAlT2G3G*4G*gTgA7C8C9CloAl lTl2-3’. This structure reveals that nucleotides Al-G4 and Cg-T12 form a double-stranded stem region that is similar to B-DNA. Unusual 1H and 31P chemical shifts gave indication of a grossly distorted loop region consisting of nucleotides G5-C8. The shift of the G4 H8 signal is unusually far upfield. Also, in both the duplex and hairpin structures, the sugars of the 5’ platinated G have been converted from an S to an N conformer. 1. 2.
X. Jia, G. Zon, and L.G. Marzilli, Znorg. Chem, 30,228 (1991). L. G. Marzilli, S. Mukundan, Jr., Y. Xu, G. Zon, A. Bergman, P. Yohannes, and M. D. Reily, in Platinum and Other Metal Coordination Compounds in Cancer Chemotherapy, Howell, S. B., Ed., Plenum Press, New York, 1991, pp. 101-114.