Visualization of drug-nucleic acid interactions at atomic resolution: I. Structure of an ethidium/dinucleoside monophosphate crystalline complex,ethidium:5-Iodouridylyl (3′–5′) adenosine

Ethidium forms a crystalline complex with the dinucleoside monophosphate 5-iodouridylyl(3′–5′)adenosine (iodoUpA). These crystals are monoclinic, space group C2, with unit cell dimensions, $\text{a = 28.45}\text{A}\text{?}\text{, b = 13.54}\text{A}\text{?}\text{, c = 34.13}\text{A}\text{?}\text{, β = 98.6 °}$. The structure has been solved to atomic resolution by Patterson and Fourier methods, and refined by full matrix least-squares to a residual of 0.20 on 2017 observed reflections. The asymmetric unit contains two ethidium molecules, two iodoUpA molecules and 27 water molecules, a total of 155 atoms excluding hydrogens. The two iodoUpA molecules are held together by adenine · uracil Watson-Crick-type base-pairing. Adjacent base-pairs within this paired iodoUpA structure and between neighboring iodoUpA molecules in adjoining unit cells are separated by about 6.7 Å; this separation results from intercalative binding by one ethidium molecule and stacking by the other ethidium molecule above and below the base-pairs. Non-crystallographic 2-fold symmetry is utilized in this model drug-nucleic acid interaction, the intercalated ethidium molecule being oriented such that its phenyl and ethyl groups lie in the narrow groove of the miniature nucleic acid double-helix. Base-pairs within the paired nucleotide units are related by a twist of 8 °. The magnitude of this angular twist is related to conformational changes in the sugar-phosphate chains that accompany drug intercalation. These changes partly reflect the differences in ribose sugar ring puckering that are observed (both iodouridine residues have C3′ endo sugar conformations, whereas both adenosine residues have C2′ endo sugar conformations), and alterations in the glycosidic torsional angles describing the base-sugar orientations. Additional small but systematic changes occur in torsional angles that involve the phosphodiester linkages and the C4′C5′ bond. Solution studies have indicated a marked sequence-specific binding preference in ethidium-dinucleotide interactions, and a probable structural explanation for this is provided by this study.This structure and the accompanying one described in the second paper [ethidium:5-idocytidylyl(3′–5′)guanosine] are examples of model drug-nucleic acid intercalative complexes, and the information provided by their structure analyses has led to a general understanding of intercalative drug binding to DNA. This is described in the third paper of this series.     

Visualization of drug-nucleic acid interactions at atomic resolution. VII. Structure of an ethidium/dinucleoside monophosphate crystalline complex, ethidium: uridylyl(3'-5') adenosine

Ethidium forms a crystalline complex with the dinucleoside monophosphate, uridylyl (3'-5') adenosine (UpA). The complex crystallizes in the monoclinic space group P2l with unit cell dimensions, a = 13.704 A, b = 31.674 A, c = 15.131 A, beta = 113.9 degrees. This light atom structure has been solved to atomic resolution and refined by full matrix least squares to a residual of 0.12, using 3,034 observed reflections. The asymmetric unit consists of two ethidium molecules, two UpA molecules and 19 solvent molecules, a total of 145 non-hydrogen atoms. The two UpA molecules are hydrogen-bonded together by Watson-Crick type base pairing. Base-pairs in this duplex are separated by 6.7 A; this reflects intercalative binding by one of the ethidium molecules. The other ethidium molecule stacks on either side of the intercalated base-paired dinucleoside monophosphate, being related by a unit cell translation along the a axis. The conformation of the sugar-phosphate backbone accompanying intercalation has been accurately determined in this analysis, and contains the mixed sugar-puckering pattern: C3' endo (3'-5') C2' endo. This same structural feature has been observed in the ethidium-iodoUpA and ethidium-iodoCpG complexes, and exists in two additional structures containing ethidium-CpG. Taken together, these studies confirm our earlier sugar-puckering assignments and demonstrate that iodine covalently bound to the C5 position on uridine or cytosine does not alter the basic sugar-phosphate geometry or the mode of ethidium intercalation in these model studies. We have proposed this stereochemistry to explain the intercalation of ethidium (as well as other simple intercalators) into both DNA and into double-helical RNA, and discuss this aspect of our work further in this paper and in the accompanying papers.     

Visualization of drug-nucleic acid interactions at atomic resolution: II. Structure of an ethidium/dinucleoside monophosphate crystalline complex,ethidium:5-iodocytidylyl (3′–5′) guanosine

Ethidium forms a second crystalline complex with the dinucleoside monophosphate 5-iodocytidylyl(3′–5′)guanosine (iodoCpG). These crystals are monoclinic, P2₁, with $\text{a = 14.06}\text{A}\text{?}\text{, b = 32.34}\text{A}\text{?}\text{, c = 16.53}\text{A}\text{?}\text{, β = 117.8 °}$. The structure has been solved to atomic resolution using rigid-body Patterson vector search and Fourier methods, and refined by full matrix least-squares to a residual of 0.16 on 3180 observed reflections. The structure consists of two ethidium molecules, two iodoCpG molecules, 27 water molecules and four methanol molecules, a total of 165 atoms (excluding hydrogens) in the asymmetric unit. Both iodoCpG molecules are hydrogen-bonded together by guanine · cytosine Watson-Crick base-pairing. Adjacent base-pairs within this paired iodoCpG structure and between neighboring iodoCpG molecules in adjoining unit cells are separated by 6.7 Å. This distance reflects the presence of an ethidium molecule intercalated between base-paired iodoCpG molecules and another ethidium molecule stacked above (and below) the dinucleotide. Approximate 2-fold symmetry is used in the interaction; this reflects the pseudo-2-fold symmetry axis of the phenanthridinium ring system in ethidium coinciding with the approximate 2-fold axis relating base-paired iodoCpG molecules. The phenyl and ethyl groups of the intercalated ethidium molecule lie in the narrow groove of the miniature iodoCpG double-helix. The stacked ethidium, however, lies in the opposite direction, its phenyl and ethyl groups neighboring iodine atoms on cytosine residues. Base-pairs within the paired nucleotide units are related by a twist of about 8 °. The magnitude of this angular twist reflects conformational changes in the sugar-phosphate chains accompanying intercalation. These primarily reflect the differences in ribose sugar ring puckering that are observed (i.e. both iodocytidine residues have C3′ endo sugar conformations, while both guanosine residues have C2′ endo sugar conformations), and alterations in the glycosidic torsional angles that describe the base-sugar orientation.The information provided by this structure analysis (along with the accompanying one (ethidium:iodoUpA), described in the previous paper) has led to an understanding of the general nature of intercalative drug binding to DNA. This is described in the third paper of this series.     

Visualization of drug-nucleic acid interactions at atomic resolution. VIII. Structures of two ethidium/dinucleoside monophosphate crystalline complexes containing ethidium: cytidylyl(3'-5') guanosine

This paper describes two complexes containing ethidium and the dinucleoside monophosphate, cytidylyl(3'-5')guanosine (CpG). Both crystals are monoclinic, space group P2l, with unit cell dimensions as follows: modification 1: a = 13.64 A, b = 32.16 A, c = 14.93 A, beta = 114.8 degrees and modification 2: a = 13.79 A, b = 31.94 A, c = 15.66 A, beta = 117.5 degrees. Each structure has been solved to atomic resolution and refined by Fourier and least squares methods; the first has been refined to a residual of 0.187 on 1,903 reflections, while the second has been refined to a residual of 0.187 on 1,001 reflections. The asymmetric unit in both structures contains two ethidium molecules and two CpG molecules; the first structure has 30 water molecules (a total of 158 non-hydrogen atoms), while the second structure has 19 water molecules (a total of 147 non-hydrogen atoms). Both structures demonstrate intercalation of ethidium between base-paired CpG dimers. In addition, ethidium molecules stack on either side of the intercalated duplex, being related by a unit cell translation along the a axis. The basic feature of the sugar-phosphate chains accompanying ethidium intercalation in both structures is: C3' endo (3'-5') C2' endo. This mixed sugar-puckering pattern has been observed in all previous studies of ethidium intercalation and is a feature common to other drug-nucleic acid structural studies carried out in our laboratory. We discuss this further in this paper and in the accompanying papers.     

Visualization of Drug-Nucleic Acid Interactions at Atomic Resolution. VII. Structure of an Ethidium/Dinucleoside Monophosphate Crystalline Complex,Ethidium: Uridylyl(3′-5′)Adenosine

Abstract

Ethidium forms a crystalline complex with the dinucleoside monophosphate, uridylyl (3′-5′) adenosine (UpA). The complex crystallizes in the monoclinic space group P2, with unit cell dimensions, a = 13.704 Å, b = 31.674 Å, c = 15.131 Å β = 113.9°. This light atom structure has been solved to atomic resolution and refined by full matrix least squares to a residual of 0.12, using 3,034 observed-reflections. The asymmetric unit consists of two ethidium molecules, two UpA molecules and 19 solvent molecules, a total of 145 non-hydrogen atoms. The two UpA molecules are hydrogen-bonded together by Watson-Crick type base pairing. Base-pairs in this duplex are separated by 6.7 Å; this reflects intercalative binding by one of the ethidium molecules. The other ethidium molecule stacks on either side of the intercalated base-paired dinucleoside monophosphate, being related by a unit cell translation along the a axis.

The conformation of the sugar-phosphate backbone accompanying intercalation has been accurately determined in this analysis, and contains the mixed sugar-puckering pattern: C3′ endo (3′-5′) C2′ endo. This same structural feature has been observed in the ethidium-iodoUpA and ethidium-iodoCpG complexes, and exists in two additional structures containing ethidium-CpG. Taken together, these studies confirm our earlier sugar-puckering assignments and demonstrate that iodine covalently bound to the C5 position on uridine or cytosine does not alter the basic sugar-phosphate geometry or the mode of ethidium intercalation in these model studies. We have proposed this stereochemistry to explain the intercalation of ethidium (as well as other simple intercalators) into both DNA and into double-helical RNA, and discuss this aspect of our work further in this paper and in the accompanying papers.     

Visualization of Drug-Nucleic Acid Interactions At Atomic Resolution. VIII. Structures of Two Ethidium/Dinucleoside Monophosphate Crystalline Complexes Containing Ethidium: Cytidylyl(3′-5′) Guanosine

Abstract

This paper describes two complexes containing ethidium and the dinucleoside monophosphate, cytidylyl(3′-5′)guanosine (CpG). Both crystals are monoclinic, space group P2₁, with unit cell dimensions as follows: modification 1: a = 13.64 Å, b = 32.16 Å, c - 14.93 Å, β = 114.8° and modification 2: a = 13.79 Å, b = 31.94 Å, c = 15.66 Å, β = 117.5°. Each structure has been solved to atomic resolution and refined by Fourier and least squares methods; the first has been refined to a residual of 0.187 on 1,903 reflections, while the second has been refined to a residual of 0.187 on 1,001 reflections. The asymmetric unit in both structures contains two ethidium molecules and two CpG molecules; the first structure has 30 water molecules (a total of 158 non-hydrogen atoms), while the second structure has 19 water molecules (a total of 147 non-hydrogen atoms). Both structures demonstrate intercalation of ethidium between base-paired CpG dimers. In addition, ethidium molecules stack on either side of the intercalated duplex, being related by a unit cell translation along the a axis.

The basic feature of the sugar-phosphate chains accompanying ethidium intercalation in both structures is: C3′ endo (3′-5′) C2′ endo. This mixed sugar-puckering pattern has been observed in all previous studies of ethidium intercalation and is a feature common to other drug-nucleic acid structural studies carried out in our laboratory. We discuss this further in this paper and in the accompanying papers.     

Visualization of drug-nucleic acid interactions at atomic resolution. X. Structure of a N,N-dimethylproflavine: deoxycytidylyl(3'-5')deoxyguanosine crystalline complex

N,N-dimethylproflavine forms a crystalline complex with deoxycytidylyl(3'-5')deoxyguanosine (d-CpG), space group P2(1)2(1)2, with a = 21.37 A, b = 34.05 A, c = 13.63 A. The structure has been solved to atomic resolution and refined by Fourier and least squares methods to a residual of 0.18 on 2,032 observed reflections. The structure consists of two N,N-dimethylproflavine molecules, two deoxycytidylyl (3'-5')deoxyguanosine molecules and 16 water molecules, a total of 128 nonhydrogen atoms. As with other structures of this type, N,N-dimethylproflavine molecules intercalate between base-paired d-CpG dimers. In addition, dimethylproflavine molecules stack on either side of the intercalated duplex, being related by a unit cell translation along the c axis. Both sugar-phosphate chains demonstrate the mixed sugar puckering geometry: C3' endo (3'-5') C2' endo. This same intercalative geometry has been seen in two other complexes containing N,N-dimethylproflavine and iodoCpG, described in the accompanying paper. Taken together, these studies indicate a common intercalative geometry present in both RNA- and DNA- model systems. Again, N,N-dimethylproflavine behaves as a simple intercalator, intercalating asymmetrically between guanine-cytosine base-pairs. The free amino- group on the intercalated dimethylproflavine molecule does not hydrogen bond directly to the phosphate oxygen. Other aspects of the structure will be presented.     

Visualization of drug-nucleic acid interactions at atomic resolution. IV. Structure of an aminoacridine--dinucleoside monophosphate crystalline complex, 9-aminoacridine--5-iodocytidylyl (3'--5') guanosine

9-Aminoacridine forms a crystalline complex with the dinucleoside monophosphate, 5-iodocytidylyl(3′–5′)guanosine (iodoCpG). These crystals are monoclinic, space group P2₁ with $\text{a = 13.98}\text{A}\text{?}\text{, b = 30.58}\text{A}\text{?}\text{, c = 22.47}\text{A}\text{?}$ and β = 113.9 °. The structure has been solved to atomic resolution by Patterson and Fourier methods, and refined by a combination of Fourier and sum-function Fourier methods. The asymmetric unit contains four 9-aminoacridine molecules, four iodoCpG molecules and 21 water molecules, a total of 245 atoms. 9-Aminoacridine demonstrates two different intercalative binding modes and, along with these, two slightly different intercalative geometries in this model system.The first of these is very nearly symmetric, the 9-amino group lying in the narrow groove of the intercalated base-paired nucleotide structure. The second shows grossly asymmetric binding to the dinucleotide, the 9-amino group lying in the wide groove of the structure. Associated with these two different intercalative binding modes is a difference in geometries in the structures. Although both structures demonstrate C3′ endo (3′–5′) C2′ endo mixed sugar puckering patterns (i.e. both cytidine residues have C3′ endo sugar conformations, while both guanosine residues have C2′ endo sugar conformations), with corresponding twist angles between base-pairs of about 10 °, they differ in the magnitude of the helical screw axis dislocation accompanying intercalation (Sobell et al., 1977a,b). In the pseudosymmetric intercalative structure, this value is about +0.5 Å, whereas in the asymmetric intercalative structure this value is about +2.7 Å. These conformational differences can be best described as a “sliding” of base-pairs on the intercalated acridine molecule.Although the pseudosymmetric intercalative structure can be used in 9-aminoacridine-DNA binding, the asymmetric intercalative structure cannot since this poses stereochemical difficulties in connecting neighboring sugar-phosphate chains to the intercalated dinucleotide. It is possible, however, that the asymmetric binding mode is related to the mechanism of 9-aminoacridine-induced frameshift mutagenesis (Sakore et al., 1977), and we discuss this possibility here in further detail.     

Reporter molecules as probes of DNA conformation: structure of a crystalline complex containing 2-methyl-4-nitroaniline ethylene dimethylammonium hydrobromide--5-iodocytidylyl (3'-5')guanosine

2-Methyl-4-nitroaniline ethylene dimethylammonium hydrobromide forms a crystalline complex with the self-complementary dinucleoside monophosphate, 5- iodocytidylyl (3'-5')guanosine. The crystals are tetragonal, with a = b = 32.192 A and c = 23.964 A, space group P4(3)2(1)2. The structure has been solved to atomic resolution by Patterson and Fourier methods, and refined by full matrix least squares. 5- Iodocytidylyl (3'-5')guanosine molecules are held together in pairs through Watson-Crick base-pairing, forming an antiparallel duplex structure. Nitroaniline molecules stack above and below guanine-cytosine pairs in this duplex structure. In addition, a third nitroaniline molecule stacks on one of the other two nitroaniline molecules. The asymmetric unit contains two 5- iodocytidylyl (3'-5')guanosine molecules, three nitroaniline molecules, one bromide ion and thirty-one water molecules, a total of 160 atoms. Details of the structure are described.     

Visualization of drug-nucleic acid interactions at atomic resolution. IX. Structures of two N,N-dimethylproflavine: 5-iodocytidylyl (3'-5') guanosine crystalline complexes

This paper describes two complexes containing N,N-dimethylproflavine and the dinucleoside monophosphate, 5-iodocytidylyl (3'-5') guanosine (iodoCpG). The first complex is triclinic, space group P1, with unit cell dimensions a = 11.78 A, b = 14.55 A, c = 15.50 A, alpha = 89.2 degrees, beta = 86.2 degrees, gamma = 96.4 degrees. The second complex is monoclinic, space group P21, with a = 14.20 A. b = 19.00 A, c = 20.73 A, beta = 103.6 degrees. Both structures have been solved to atomic resolution and refined by Fourier and least squares methods. The first structure has been refined anisotropically to a residual of 0.09 on 5,025 observed reflections using block diagonal least squares, while the second structure has been refined anisotropically to a residual of 0.13 on 2,888 reflections with full matrix least squares. The asymmetric unit in both structures contains two dimethylproflavine molecules and two iodoCpG molecules; the first structure has 16 water molecules (a total of 134 non-hydrogen atoms), while the second structure has 18 water molecules (a total of 136 non-hydrogen atoms). Both structures demonstrate intercalation of dimethylproflavine between base-paired iodoCpG dimers. In addition, dimethylproflavine molecules stack on either side of the intercalated duplex, being related by a unit cell translation along b and a axes, respectively. The basic structural feature of the sugar-phosphate chains accompanying dimethylproflavine intercalation in both structures is the mixed sugar puckering pattern: C3' endo (3'-5') C2' endo. This same structural information is again demonstrated in the accompanying paper, which describes a complex containing dimethylproflavine with deoxyribo-CpG. Similar information has already appeared for other "simple" intercalators such as ethidium, acridine orange, ellipticine, 9-aminoacridine, N-methyl-tetramethylphenanthrolinium and terpyridine platinum. "Complex" intercalators, however, such as proflavine and daunomycin, have given different structural information in model studies. We discuss the possible reasons for these differences in this paper and in the accompanying paper.     

Visualization of Drug-Nucleic Acid Interactions at Atomic Resolution. X. Structure of a N,N-Dimethylproflavine: Deoxycytidylyl(3′-S′)Deoxyguanosine Crystalline Complex

Abstract

N,N-dimethylproflavine forms a crystalline complex with deoxycytidylyl(3′-5′)deoxyguanosine (d-CpG), space group P2₁,2₁2, with a = 21.37 Å, b = 34.05 Å, c = 13.63 Å. The structure has been solved to atomic resolution and refined by Fourier and least squares methods to a residual of 0.18 on 2,032 observed reflections. The structure consists of two N,N- dimethylproflavine molecules, two deoxycytidylyl (3′-5′)deoxyguanosine molecules and 16 water molecules, a total of 128 nonhydrogen atoms. As with other structures of this type, N,N-dimethylproflavine molecules intercalate between base-paired d-CpG dimers. In addition, dimethylproflavine molecules stack on either side of the intercalated duplex, being related by a unit cell translation along the c axis.

Both sugar-phosphate chains demonstrate the mixed sugar puckering geometry: C3′ endo (3′-5′) C2′ endo. This same intercalative geometry has been seen in two other complexes containing N,N-dimethylproflavine and iodoCpG, described in the accompanying paper. Taken together, these studies indicate a common intercalative geometry present in both RNA- and DNA- model systems. Again, N,N-dimethylproflavine behaves as a simple intercalator, intercalating asymmetrically between guanine-cytosine base-pairs. The free amino- group on the intercalated dimethylproflavine molecule does not hydrogen bond directly to the phosphate oxygen. Other aspects of the structure will be presented.     

Visualization of drug--nucleic acid interactions at atomic resolution. VI. Structure of two drug--dinucleoside monophosphate crystalline complexes, ellipticine--5-iodocytidylyy (3'-5') guanosine and 3,5,6,8-tetramethyl-N-methyl phenanthrolinium--5-iodocytidylyl (3'-5') guanosine

Ellipticine and 3,5,6,8-tetramethyl-N-methyl phenanthrolinium form complexes with the dinucleoside monophosphate, 5-iodocytidylyl(3′–5′)guanosine. These crystals are isomorphous: ellipticine-iodoCpG² crystals are monoclinic, space group P2₁ with $\text{a = 13.88}\text{A}\text{?}\text{, b = 19.11}\text{A}\text{?}\text{, c = 21.42}\text{A}\text{?}$, β = 105.4; TMP-iodoCpG crystals are monoclinic, space group P2₁, with $\text{a = 13.99}\text{A}\text{?}\text{, b = 19.12}\text{A}\text{?}\text{, c = 21.31}\text{A}\text{?}$, β = 104.9 °. Both structures have been solved to atomic resolution by Patterson and Fourier methods, and refined by full matrix least-squares.The asymmetric unit in the ellipticine-iodoCpG structure contains two ellipticine molecules, two iodoCpG molecules, 20 water molecules and 2 methanol molecules, a total of 144 atoms, whereas, in the tetramethyl-N-methyl phenanthrolinium-iodoCpG complex, the asymmetric unit contains two TMP molecules, two iodoCpG molecules, 17 water molecules and 2 methanol molecules, a total of 141 atoms. In both structures, the two iodoCpG molecules are hydrogenbonded together by guanine-cytosine Watson-Crick base-pairing. Adjacent base-pairs within this paired iodoCpG structure are separated by about 6.7 Å; this separation results from intercalative binding by one ellipticine (or TMP) molecule and stacking by the other ellipticine (or TMP) molecule above or below the base-pairs. Base-pairs within the paired nucleotide units are related by a twist of 10 to 12 °. The magnitude of this angular twist is related to conformational changes in the sugar-phosphate chains that accompany drug intercalation. These changes partly reflect the mixed sugar puckering pattern observed: C3′ endo (3′–5′) C2′ endo (i.e. both iodocytidine residues have C3′ endo conformations, whereas both guanosine residues have C2′ endo conformations), and additional small but systematic changes in torsional angles that involve the phosphodiester linkages and the C4′C5′ bond.The stereochemistry observed in these model drug-nucleic acid intercalative complexes is almost identical to that observed in the ethidium-iodoUpA and -iodoCpG complexes determined previously (Tsai et al., 1975a,b,1977; Jain et al., 1977). This stereochemistry is also very similar to that observed in the 9-aminoacridine-iodoCpG and acridine orange-iodoCpG complexes described in the preceding papers (Sakore et al., 1979 Reddy et al., 1979). We have already proposed this stereochemistry to provide a unified understanding of a large number of intercalative drug-DNA (and RNA) interactions (Sobell et al., 1977a,b), and discuss this aspect of our work further in this paper.     

Intercalation of water molecules between nucleic acid bases in the crystal structures of 6-azathymine hemihydrate and 5-amino-2-thiocytosine dihydrochloride dihydrate

The crystal structure of 6-azathymine hemihydrate (6AzTH) exhibits a novel intercalation of water molecules interposed half-way between the modified bases 6.3 to 6.7 A apart. The crystal contains four molecules of 6-azathymine (6AzT) and two water molecules as the independent repeating unit. These two water molecules together with the four bases form two separate water sandwiches. In the crystal structure these sandwiches form two sets of local clusters. The anhydrous crystalline form of 6AzT, on the other hand, is stabilized by base stacking interactions. Both the water molecules in 6AzTH that are involved in sandwich formation have trigonal coordination around them. A reexamination of the crystal structure of 5-amino-2-thiocytosine (5A2TC) revealed that one of the water molecules in this structure also forms a water sandwich and has trigonal coordination whereas the other water molecule with tetrahedral coordination does not form a sandwich. The environment and the characteristics of the intercalated water molecule in these structures suggest a possible role for such water intercalations in the dynamics of DNA. Crystals of 6AzTH are monoclinic, space group P21/n, with unit cell parameters a = 8.861 (1), b = 13.177 (3), c = 20.662 (2) A, beta = 93.35 (1) degrees, and Z = 16. From diffractometer data (2503 reflections, greater than or equal to 3 sigma), the crystal structure was solved and refined to an R of 0.056.     

Crystallization of mouse pancreatic ribonuclease

Mouse pancreatic ribonuclease has been crystallized in a form suitable for X-ray structure determination. The crystals grown from solutions of 2-methyl-2,4-pentanediol diffract to high resolution and belong to the hexagonal space group P6(1) (P6(5)) with unit cells dimensions a = b = 64.44 A, c = 53.91 A, y = 120 degrees and V = 1.94 x 10(5) A3 (1 A = 0.1 nm). There are six molecules per unit cell (1 molecule/asymmetric unit), and Vm = 2.3 A3/dalton.     

Visualization of Drug-Nucleic Acid Interactions at Atomic Resolution. IX. Structures of Two N,N-Dimethylproflavine: 5-Iodocytidylyl (3′-5′) Guanosine Crystalline Complexes

Abstract

This paper describes two complexes containing N,N-dimethylproflavine and the dinucleoside monophosphate, 5-iodocytidylyl(3′-5′)guanosine (iodoCpG). The first complex is triclinic, space group PI, with unit cell dimensions a = 11.78 Å, b = 14.55 Å, c = 15.50 Å, a = 89.2°, β = 86.2°, γ = 96.4°. The second complex is monoclinic, space group P2₁, with a = 14.20 Å, b = 19.00 Å, c = 20.73 Å, β = 103.6°. Both structures have been solved to atomic resolution and refined by Fourier and least squares methods. The first structure has been refined anisotropically to a residual of 0.09 on 5,025 observed reflections using block diagonal least squares, while the second structure has been refined isotropically to a residual of 0.13 on 2,888 reflections with full matrix least squares. The asymmetric unit in both structures contains two dimethylproflavine molecules and two iodoCpG molecules; the first structure has 16 water molecules (a total of 134 non-hydrogen atoms), while the second structure has 18 water molecules (a total of 136 non-hydrogen atoms). Both structures demonstrate intercalation of dimethylproflavine between base-paired iodoCpG dimers. In addition, dimethylproflavine molecules stack on either side of the intercalated duplex, being related by a unit cell translation along b and a axes, respectively.

The basic structural feature of the sugar-phosphate chains accompanying dimethylproflavine intercalation in both structures is the mixed sugar puckering pattern: C3′ endo (3′-5′) C2′ endo. This same structural information is again demonstrated in the accompanying paper, which describes a complex containing dimethylproflavine with deoxyribo-CpG.

Similar information has already appeared for other “simple” intercalators such as ethidium, acridine orange, ellipticine, 9-aminoacridine, N-methyl-tetramethylphenanthrolinium and terpyridine platinum. “Complex” intercalators, however, such as proflavine and daunomycin, have given different structural information in model studies. We discuss the possible reasons for these differences in this paper and in the accompanying paper.     

Visualization of drug--nucleic acid interactions at atomic resolution. V. Structure of two aminoacridine--dinucleoside monophosphate crystalline complexes, proflavine--5-iodocytidylyl (3'-5') guanosine and acridine orange--5-iodocytidylyl (3'-5') guanosine

Acridine orange and proflavine form complexes with the dinucleoside monophosphate, 5-iodocytidylyl(3′–5′)guanosine. The acridine orange-iodoCpG² crystals are monoclinic, space group P2₁, with unit cell dimensions $\text{a = 14.36}\text{A}\text{?}\text{, b = 19.64}\text{A}\text{?}\text{, c = 20.67}\text{A}\text{?}$, β = 102.5 °. The proflavine-iodoCpG crystals are monoclinic, space group C2, with unit cell dimensions $\text{a = 32.14}\text{A}\text{?}\text{, b = 22.23}\text{A}\text{?}\text{, c = 18.42}\text{A}\text{?}$, β = 123.3 °. Both structures have been solved to atomic resolution by Patterson and Fourier methods, and refined by full matrix least-squares.Acridine orange forms an intercalative structure with iodoCpG in much the same manner as ethidium, ellipticine and 3,5,6,8-tetramethyl-N-methyl phenanthrolinium (Jain et al., 1977, Jain et al., 1979), except that the acridine nucleus lies asymmetrically in the intercalation site. This asymmetric intercalation is accompanied by a sliding of base-pairs upon the acridine nucleus and is similar to that observed with the 9-aminoacridine-iodoCpG asymmetric intercalative binding mode described in the previous papers (Sakore et al., 1977, Sakore et al., 1979). Basepairs above and below the drug are separated by about 6.8 Å and are twisted about 10 °; this reflects the mixed sugar puckering pattern observed in the sugar-phospate chains: C3′ endo (3′–5′) C2′ endo (i.e. each cytidine residue has a C3′ endo sugar comformation, while each guanosine residue has a C2′ endo sugar conformation), alterations in glycosidic torsional angles and other small but significant conformational changes in the sugar-phosphate backbone.Proflavine, on the other hand, demonstrates symmetric intercalation with iodoCpG. Hydrogen bonds connect amino groups on proflavine with phosphate oxygen atoms on the dinucleotide. In contrast to the acridine orange structure, base-pairs above and below the intercalative proflavine molecule are twisted about 36 °. The altered magnitude of this angular twist reflects the sugar puckering pattern that is observed: C3′ endo (3′–5′) C3′ endo. Since proflavine is known to unwind DNA in much the same manner as ethidium and acridine orange (Waring, 1970), one cannot use the information from this model system to understand how proflavine binds to DNA (it is possible, for example, that hydrogen bonding observed between proflavine and iodoCpG alters the intercalative geometry in this model system).Instead, we propose a model for proflavine-DNA binding in which proflavine lies asymmetrically in the intercalation site (characterized by the C3′ endo (3′–5′) C2′ endo mixed sugar puckering pattern) and forms only one hydrogen bond to a neighboring phosphate oxygen atom. Our model for proflavine-DNA binding, therefore, is very similar to our acridine orange-DNA binding model. We will describe these models in detail in this paper.     

A novel end-to-end binding of two netropsins to the DNA decamers d(CCCCCIIIII)2, d(CCCBr5CCIIIII)2and d(CBr5CCCCIIIII)2.

Netropsin is bound to the DNA decamer d(CCCCCIIIII)2, the C-4 bromo derivative d(CCCBr5CCIIIII)2and the C-2 bromo derivative d(CBr5CCCCIIIII)2in a novel 2:1 mode. Complexes of the native decamer and the C-4 bromo derivative are isomorphous, space group P1, unit cell dimensions a = 32.56 A (32.66), b = 32.59 A (32.77), c = 37.64 A (37.71), alpha = 86.30 degrees (86.01 degrees), beta = 84.50 degrees (84.37 degrees), gamma = 68.58 degrees (68.90 degrees) with two independent molecules (A and B) in the asymmetric unit (values in parentheses are for the derivative). The C-2 bromo derivative is hexagonal P61, unit cell dimensions a = b = 32.13 A, c = 143.92, gamma = 120 degrees with one molecule in the asymmetric unit. The structures were solved by the molecular replacement method. The novelty of the structures is that there are two netropsins bound end-to-end in the minor groove of each B-DNA decamer which has nearly a complete turn. The netropsins are held by hydrogen bonding interactions to the base atoms and by sandwiching van der Waal's interactions from the sugar-phosphate backbones of the double helix similar to every other drug.DNA complex. Each netropsin molecule spans approximately 5 bp. The netropsins refined with their guanidinium heads facing each other at the center, although an orientational disorder for the netropsins cannot be excluded. The amidinium ends stretch out toward the junctions and bind to the adjacent duplexes in the columns of stacked symmetry-related complexes. Both cationic ends of netropsin are bridged by water molecules in one of the independent molecules (molecule A) of the triclinic structures and also the hexagonal structure to form pseudo-continuous drug.decamer helices.     

Comparison of the structures of the native form of rat liver 5S rRNA and yeast tRNAphe: small-angle and wide-angle X-ray scattering study

The small-angle and wide-angle X-ray scattering of tRNA^phe (yeast) and ribosomal 5S RNA (rat liver) in solution have been analysed and compared. tRNA^phe in solution is folded into a compact L-shaped structure similar to its structure in crystals. The geometry of the secondary structure of the double helical regions is also equivalent to the A-form in the crystalline state. Despite differences between the molar mosses of 5S rRNA (40 000 g mol^?1) and tRNA^phe (25 000 g mol^?1), and the fact that the 5S rRNA molecule is more anisometric than the tRNA^phe molecule, there are many structural similarities. The geometrical parameters of the secondary structure of double helical regions in both RNA molecules are almost identical; the mean rise per base pair is about 0.253–0.28 nm and the mean turn angle is about 32.5–33.5. Identical cross-sectional radii of gyration, R_sq,1 ≈ 1.16 nm and R_sq,2 = 0.92 nm, identical molar mass per unit length, , and a mean thickness of the molecules D ≈ 1.65 nm suggest a similar, nearly coplanar organization of isolated, double helical arms. Furthermore, there are compact regions in the central parts of both molecules, which are the sites of tertiary interactions in the tRNA^phe molecule and are a potential site of tertiary interactions in the SS rRNA molecule for stabilization of the complicated L-shape of the two molecules. Both molecules have a pseudo-twofold axis,w hich may play a role in recognition for binding of specific proteins.     

The crystal and molecular structure of 2'-deoxy-2'-fluoroinosine monohydrate.

The structure of the hydrate of 2'-deoxy-2'-fluoroinosine has been determined by single-crystal x-ray diffraction. The nucleoside crystallizes in space group P2(1)2(1)2(1) with unit cell dimensions, a = 33.291, b = 10. 871, c = 6.897A. There are two nucleosides and two water molecules in the asymmetric unit. The structure was solved by direct methods and refined to a residual R = 0.095. The two independent nucleosides in the asymmetric unit show different conformations about the glycosidic bond, while other structural details are similar. The base orientation to the sugar is syn in molecule A, whereas anti in molecule B. The exocyclic C(4')-C(5') bond conformation defined with respect to C(3')-C(4')-C(5')-O(5') is gauche+ in both molecules A and B. The sugar ring pucker defined by the pseudorotation phase angle P is a twisted conformation in both, C(3')-endo-C(4')-exo with P = 29 degrees in molecule A and C(4')-exo-C(3')-endo with P = 41 degrees in molecule B. It is shown by comparison with x-ray results of other 2'-fluoronucleosides and unmodified nucleosides including inosines that, in addition to a strong preference of the C(3')-endo type pucker, twisted conformations involving C(4')-exo puckering may be one of characteristic features of 2'-fluoronucleosides.     

The synthetic DNA duplex of poly d(Abr5U).poly d(Abr5U) adopts an A-DNA-like structure

An X-ray fiber diffraction study of the synthetic DNA duplex poly d(Abr5U).poly d(Abr5U) shows that its sodium salt adopts an unexceptional A-DNA-like structure. Similar to A-DNA, two molecules are packed in a monoclinic unit cell (a = 2.23 nm, b = 4.14 nm, c = 5.61 nm and alpha = beta = gamma = 90 degrees) of space group C2. Because of its dinucleotide chemical motif, the c-repeat is twice that in A-DNA but, notably, corresponding backbone conformation angles of adjacent nucleotides are almost identical. This is in marked contrast to many B-like conformations of polydinucleotides.