Volume 4 Number 4 April 1977
Nucleic Acids Research
5-Nitrouridine-monohydrate: crystal structure and conformation
Ernst
Egert*, Hans-Jorg Lindner, Wolfgang Hillen +, Hans Gunter Gassen+
_~~~~~~~~~~~~~~
Fachgebiet Organische Chemie 1* and Fachgebiet Biochemie+ Technische Hochschule Darmstadt, Petersenstr. 22, D-6100 Darmstadt, GFR Received 4 February 1977
ABSTRACT The crystal structure of 5-nitrouridine was determined by X-ray analysis. The pyrimidine ring is slightly non-planar, showing a shallow boat conformation. The nitro group has no influence on the C4 - 04 bond length as compared to uridine. The ribose shows the C3'-endo conformation and the base is in the anti orientation to the sugar with a torsion angle of 25.60. This conformation is stabilized by a hydrogen bond from the base to the ribosyl moiety (H6 ... 05'). Stacking interactions between neighboring bases are almost negligible in the crystal. A water molecule is involved in a bifurcated donating hydrogen bond to 04 and to 052 of the nitro group of the one base and an accepting bond from the H3 of the other base. Two more hydrogen bonds are formed between the water molecule and the ribose. The structural aspects of 5-nitrouridine are discussed with respect to the special stacking features found for 5-nitro-1-(S-D-ribosyluronic acid)-uracil monohydrate in the crystal (1). INTRODUCTION The secondary and tertiary structure of nucleic acids containing the common nucleosides is mainly determined by base stacking as well as by hydrogen bonding. Both factors may be influenced when base modified nucleosides are incorporated into nucleic acids. Substituents in the 5-position of pyrimidine nucleosides have no direct effect on the base pairing properties but should have a definite influence on the geometry and stability of the base stacking. Modified pyrimidine ribonucleosides are found mainly in tRNA (2). These include, aside from methylated or thiolated compounds, 5-substituted uridines or 2-thiouridines. The impact of these modifications on the three-dimensional structure of tRNAPhe from yeast has recently been discussed in detail (3). Functional implications seem to be evident if a 5substituted uridine occurs in the 5'-position - the wobble po-
C) Information Retrieval Limited 1 Falconberg Court London Wl V 5FG England
929
Nucleic Acids Research sition - of the anticodon (4). Replacement of uridine by its 5substituted analogue like 5-carboxymethyluridine or 5-carboxymethyluridine methylester in one isoaccepting species of a certain tRNA could block the wobble by pairing only with adenosine but not with guanosine (5). This restriction of codon recognition could be explained by an influence of the substituent in the wobble nucleoside on the geometry of the anticodon stack. Numerous homopolynucleotides containing modified nucleosides were examined by physical techniques in order to correlate the type of substitution to the stacking stability (6). Knowledge concerning the geometry of base stacking, however, can only come from X-ray diffraction techniques. The crystal structures of many natural and synthetic nucleic acid compounds have shown that stacking between the base moieties of nucleosides occurs in crystals as well (7). Naturally, there is no direct correlation between the stacking geometry of a nucleoside in a crystal and nucleosides as components of a nucleic acid. Certain parameters derived from the crystal structure of nucleosides may, however, shed light on our understanding of complicated structures like an anticodon in a tRNA. When uridines are substituted by halogens in the 5-position these polarizable groups often stack on the aromatic ring of neighboring nucleosides (7). Of the uridines substituted in the 5-position whose dipole moments are known, only 5-nitrouridine shows a major difference in the dipole moment compared to uridine (8). Furthermore, the nitro group displays an electron withdrawing effect on the base (-I), which should increase the polarity in the base portion of the nucleoside. We decided to study the conformation of 5-nitrouridine in the crystalline state within a broader perspective in order to clarify the function of pyrimidine nucleosides modified in the 5-position in single stranded stacked nucleic acids. MATERIALS AND METHODS
1-6-D- (2',3 ' ,5'-tribenzoyl)-ribofuranosyl-5-nitrouracil was a generous gift of Dr. Vorbruggen (9). It was debenzoylated by treatment with absolute methanol saturated with NH3 for 24 h at 0OC. The nucleoside was crystallized three times from water and showed the same spectral characteristics as reported by
930
Nucleic Acids Research Wempen et al. (10). Crystals were grown as light brown monoclinic plates (C H1108N3*H20, M = 307 g-mol 1) from aqueous solution. The cell dimensions were a = 10.569 R, b = 7.144 R, c = 8.471 R and a = 92.86 with two molecules per unit cell (space group P21). Intensity data of 819 symmetry-independent reflections with e < 550 were measured on a Stoe two-circle diffractometer with CuK.-radiation (X = 1.5418 i). 747 normalized structure factors with IEI>3aE were obtained. The data were not corrected for absorption because of the small linear absorption coefficient (i = 11.5 cm 1). The structure was determined by direct methods (11, 12) using the program SHELX-76 (13) and refined by full matrix least squares to a discrepancy factor of R = 0.057. All hydrogen atoms including the two hydrogen atoms of the water molecule could be located in successive difference Fourier syntheses. RESULTS The conformational features of the 5-nitrouridine are not unexpected, especially when compared to the known structures of 5-nitrouracil and 5-nitro-1-(a-D-ribosyluronic acid)uracil but individual details are interesting (14, 1). 1. Structure of the base The pyrimidine ring is slightly non-planar but represents a shallow boat conformation with Ni and C4 above the plane showing a standard deviation of the ring atoms of 0.025 R from the best plane. The nitro group is not co-planar with the pyrimidine ring but forms an angle of 12° with the best plane of the base. This is consistent with the orientation of the nitro group in 5-nitrouracil (14). The bond lengths in the pyrimidine ring are nearly identical in the base and in the nucleoside, with the exception that the C6 - Ni bond is shorter by 0.046 R whereas C6 - C5 and Ni - C2 are longer by 0.030 R in the nucleoside (Fig. 1, 2). The length of the C4 - 04 bond (1.232 A) is the same as it is in uridine and differs from that of 3-deazauridine (1.341 R) (15). 2. Structure of the ribose The ribose in the 5-nitrouridine has a C3'-endo conformation with Cl', C2', C4' and 01' in the plane and C3' out of the
931
Nucleic Acids Research
Fig. 1
Fig. 2
Structural representation of 5-nitrouridine (plot program ORTEP by C.K. Johnson (18)).
Structural formula of 5-nitrouridine, with the determined bond lengths and the bond and torgion angles. Standard deviation:a x=0.08 Al (x=C,N,0)
axxx=0.5
932
Nucleic Acids Research plane by 0.58 A (Fig. 1, 2). The Cl' - 01' bond is shorter by 0.071 i compared to the C4' - 01' bond, as is usually found in nucleoside structures. 3. Conformation about the glycosidic bond The conformation about the glycosidic bond is anti with a torsion angle C6 - Ni - Cl' - 01' of 25.60. The base is held in 05' this conformation by an "intramolecular interaction" H6 (Table 1). Atom 05' is positioned above the ribose and is "gauche" to both C3' - C4' and to 01' - C4' (Fig. 2). The conformation of the ribosyl moiety is similar to uridine, in which case the deviations from the ideal gauche conformation (+600 and -60°) are greater. 4. Packing of the molecules in the crystal The packing of the molecules in the crystal is portrayed in Fig. 3 and 4. If one sights in the direction of the c-axis (Fig. 3), then the bases are located at the left and right side separated by the ribose moieties. The sugars are kept in place by the hydrogen bonds listed in Table 1. In the direction of the a-axis (Fig. 4), one can observe that the base moieties are related by a screw axis and that they form an angle of 480 with respect to each other. The size of this angle indicates that stacking interactions between these bases are almost negligible. Bases within neighboring elementary cells in the b-direction are connected via a water molecule. The bases form no hydrophobic stacking; they are nearly perpendicular to the bc plane and form an angle of 660 with respect to the ac plane. The water is involved in a bifurcated donating hydrogen bond to 04 of the keto group and to 052 of the nitro group. In addition an accepting bond with the amide group of a second base is formed. Furthermore the water interacts with the 01' and 3'-OH group of the ribose moieties (Table 1). The one water molecule forms two donating and two accepting hydrogen bonds which are arranged in a nearly tetrahedral alignment around the oxygen atom. In this respect, the packing of the 5-nitrouridine differs from that of uridine which contains two symmetry-independent molecules in the asymmetric unit. Both pyrimidine rings form a fairly intimate but highly offset stacked pair (16).
933
Nucleic Acids Research X-H ---Y
H --- Y
[Al
X --- Y Li
lX-H-Y
C6 -H
---05'
2.446
3.333
159.7
C2'-H
---051
2.200
3.281
161.4
02'-H 05'-H
---05' ---03'
1.501 1.807
2.672 2.712
165.6 156.2
N3 -H ---OW 03'-H ---OW OW -HW1---01 ' OW -HW2---04 OW -HW2---052
1.952 1.703 2.176 2.428 2.201
3.025 2.763 2.806 3.432 2.963
173.5 143.2 134.2 161.3 128.3
Table 1
[0]
List of X-H---Y contacts divided into four groups: a) intramolecular "interaction" b) intermolecular "interaction" c) intermolecular hydrogen bonds between the ribose moieties d) hydrogen bonds with participation of the water molecule
DISCUSSION Recently, the structure of 5-nitro-1-(a-D-ribosyl-uronic acid)-uracil monohydrate was reported by Srikrishnan and Parthasarathy (1). The structure of the pyrimidine ring in this compound is practically identical with 5-nitrouridine. The base is, also, in the anti orientation with a torsion angle of 0 53.9°. This conformation is stabilized in both molecules by an intramolecular interaction H6 ... 05' which in the case of 5nitrouridine is increased to 3.333 R compared to 3.104 R in the uronic acid. It is always a matter of opinion whether to designate this interaction a hydrogen bond. The structure of the ribose and especially the packing of the molecules in the crystal turns out to be completely different. In 5-nitrouridine the ribose exists in the C3'-endo conformation whereas the uronic acid displays a C2'-endo - C3'-exo structure. The uronic acid - according to the pictures published by these authors - seems to form a sandwich type structure with a water molecule in between the two bases. The intercalation of the water molecule increases the base-base distance to 6.55 R. In the 5-nitrouridine structure, as determined by us, the
934
Nucleic Acids Research water molecule connects two consecutive bases by forming hydrogen bonds to the polar groups of both (Fig. 3, 4). If one sights perpendicular to the plane of the bases, which are connected by a translation along the c-axis, they are 7.76 R apart. If one draws a hypothetical line connecting the two Ni or C4 atoms, i. e. the outermost ring atoms, then the water molecule is placed outside of that "base stacking" by about 2 i in the direction of the a-axis (Fig. 4). Therefore, an intercalating water molecule was not found in this structure. It should be mentioned that the CD spectrum of Apno5U 5 shows major differences when compared to ApU, Apcl U or
Apbr5U.
Thus the electronic effect of the nitro group seems to influence the stacking of 5-nitrouridine in the crystal as well as the conformation of dinucleoside phosphates in solution (17).
Therefore, if a substituent in the 5-position increases the polarity of the base moiety of the nucleoside, solvent in-
Fig. 3
Packing of 5-nitrouridine within the crystal as viewed in the c-axis. The shading shows the ribosyl moieties. The hatched ellypsoids represent the oxygen atoms of the water molecules.
935
Nucleic Acids Research
Fig. 4
Packing of 5-nitrouridine within the crystal as viewed relative to the a-axis. The ribose moieties are omitted.
teraction could have a major impact on the stacking tendency of nucleosides. This should be kept in mind when the structural and functional consequences of 5-substitutioni in the wobble nucleoside of the anticodon of a tRNA are discussed. ACKNOWLEDGEMENT We wish to thank Dr. H. Vorbruggen, Schering AG. Berlin, for the nucleoside-tribenzoate. Furthermore we express our gratitude to O.E. Beck for his help in preparing the manuscript. The work was supported by the Fonds der Chemischen Industrie. REFERENCES 1 Srikrishnan, T. and Parthasarathy, R. (1976) Nature 264, 379-380 2 Barell, B.G. and Clark, B.F.C. (1974) in Handbook of Nucleic Acid Sequences, Joynson and Bruvvers Ltd., Oxford 3 Quigley, G.J. and Rich, A. (1976) Science 194, 796-806 4 Crick, F.H.C. (1966) J.Mol.Biol. 19, 548-555 5 Gray, M.W. (1976) Biochemistry 15, 3046-3051 6 Borek, J. (1971) in Physicochemical Characterisation of Oligonucleotides and Polynucleotides IFI/Plenum Corp. N.Y., Washington, London 7 Hawkinson, S.W. and Coulter, C.L. (1971) Acta Cryst. B 27, 34-42
936
Nucleic Acids Research 8
9 10 11 12 13 14 15 16
17 18
Kulakowska, I., Geller, M., Lesyng, B. and Wierzchowski, K.L. (1974) Biochim.Biophys. Acta 361, 119-130 Niedballa, U. and Vorbrtggen, H. (1974) J.Org.Chem. 39, 3668-3672 Wempen, I., Doerr, I.L., Kaplan, L. and Fox, J.J. (1960) J.Amer.Chem.Soc. 82, 1624-1629 Karle, J. and Karle, I.L. (1966) Acta Cryst. 21, 849-859 Germain, G. and Woolfson, M.M. (1968) Acta Cryst. B 24, 91-96 Sheldrick, G. not yet published Craven, B.M. (1967) Acta Cryst. 23, 376-383 Schwalbe, C.H., Gassen, H.G. and Saenger, W. (1972) Nature 238, 250 Green, E.A., Rosenstein, R.D., Shiono, R., Abraham, D.J., Trus, B.L. and Marsh, R.E. (1975) Acta Cryst. B 31, 102-107 Hillen, W. and Gassen, H.G. (manuscript in preparation) Johnson, C.K. (1965) ORNL-3794 (revised) Oak Ridge National Laboratory, Oak Ridge
937
Nucleic Acids Research
5-Nitrouridine-monohydrate: crystal structure and conformation
Ernst
Egert*, Hans-Jorg Lindner, Wolfgang Hillen +, Hans Gunter Gassen+
_~~~~~~~~~~~~~~
Fachgebiet Organische Chemie 1* and Fachgebiet Biochemie+ Technische Hochschule Darmstadt, Petersenstr. 22, D-6100 Darmstadt, GFR Received 4 February 1977
ABSTRACT The crystal structure of 5-nitrouridine was determined by X-ray analysis. The pyrimidine ring is slightly non-planar, showing a shallow boat conformation. The nitro group has no influence on the C4 - 04 bond length as compared to uridine. The ribose shows the C3'-endo conformation and the base is in the anti orientation to the sugar with a torsion angle of 25.60. This conformation is stabilized by a hydrogen bond from the base to the ribosyl moiety (H6 ... 05'). Stacking interactions between neighboring bases are almost negligible in the crystal. A water molecule is involved in a bifurcated donating hydrogen bond to 04 and to 052 of the nitro group of the one base and an accepting bond from the H3 of the other base. Two more hydrogen bonds are formed between the water molecule and the ribose. The structural aspects of 5-nitrouridine are discussed with respect to the special stacking features found for 5-nitro-1-(S-D-ribosyluronic acid)-uracil monohydrate in the crystal (1). INTRODUCTION The secondary and tertiary structure of nucleic acids containing the common nucleosides is mainly determined by base stacking as well as by hydrogen bonding. Both factors may be influenced when base modified nucleosides are incorporated into nucleic acids. Substituents in the 5-position of pyrimidine nucleosides have no direct effect on the base pairing properties but should have a definite influence on the geometry and stability of the base stacking. Modified pyrimidine ribonucleosides are found mainly in tRNA (2). These include, aside from methylated or thiolated compounds, 5-substituted uridines or 2-thiouridines. The impact of these modifications on the three-dimensional structure of tRNAPhe from yeast has recently been discussed in detail (3). Functional implications seem to be evident if a 5substituted uridine occurs in the 5'-position - the wobble po-
C) Information Retrieval Limited 1 Falconberg Court London Wl V 5FG England
929
Nucleic Acids Research sition - of the anticodon (4). Replacement of uridine by its 5substituted analogue like 5-carboxymethyluridine or 5-carboxymethyluridine methylester in one isoaccepting species of a certain tRNA could block the wobble by pairing only with adenosine but not with guanosine (5). This restriction of codon recognition could be explained by an influence of the substituent in the wobble nucleoside on the geometry of the anticodon stack. Numerous homopolynucleotides containing modified nucleosides were examined by physical techniques in order to correlate the type of substitution to the stacking stability (6). Knowledge concerning the geometry of base stacking, however, can only come from X-ray diffraction techniques. The crystal structures of many natural and synthetic nucleic acid compounds have shown that stacking between the base moieties of nucleosides occurs in crystals as well (7). Naturally, there is no direct correlation between the stacking geometry of a nucleoside in a crystal and nucleosides as components of a nucleic acid. Certain parameters derived from the crystal structure of nucleosides may, however, shed light on our understanding of complicated structures like an anticodon in a tRNA. When uridines are substituted by halogens in the 5-position these polarizable groups often stack on the aromatic ring of neighboring nucleosides (7). Of the uridines substituted in the 5-position whose dipole moments are known, only 5-nitrouridine shows a major difference in the dipole moment compared to uridine (8). Furthermore, the nitro group displays an electron withdrawing effect on the base (-I), which should increase the polarity in the base portion of the nucleoside. We decided to study the conformation of 5-nitrouridine in the crystalline state within a broader perspective in order to clarify the function of pyrimidine nucleosides modified in the 5-position in single stranded stacked nucleic acids. MATERIALS AND METHODS
1-6-D- (2',3 ' ,5'-tribenzoyl)-ribofuranosyl-5-nitrouracil was a generous gift of Dr. Vorbruggen (9). It was debenzoylated by treatment with absolute methanol saturated with NH3 for 24 h at 0OC. The nucleoside was crystallized three times from water and showed the same spectral characteristics as reported by
930
Nucleic Acids Research Wempen et al. (10). Crystals were grown as light brown monoclinic plates (C H1108N3*H20, M = 307 g-mol 1) from aqueous solution. The cell dimensions were a = 10.569 R, b = 7.144 R, c = 8.471 R and a = 92.86 with two molecules per unit cell (space group P21). Intensity data of 819 symmetry-independent reflections with e < 550 were measured on a Stoe two-circle diffractometer with CuK.-radiation (X = 1.5418 i). 747 normalized structure factors with IEI>3aE were obtained. The data were not corrected for absorption because of the small linear absorption coefficient (i = 11.5 cm 1). The structure was determined by direct methods (11, 12) using the program SHELX-76 (13) and refined by full matrix least squares to a discrepancy factor of R = 0.057. All hydrogen atoms including the two hydrogen atoms of the water molecule could be located in successive difference Fourier syntheses. RESULTS The conformational features of the 5-nitrouridine are not unexpected, especially when compared to the known structures of 5-nitrouracil and 5-nitro-1-(a-D-ribosyluronic acid)uracil but individual details are interesting (14, 1). 1. Structure of the base The pyrimidine ring is slightly non-planar but represents a shallow boat conformation with Ni and C4 above the plane showing a standard deviation of the ring atoms of 0.025 R from the best plane. The nitro group is not co-planar with the pyrimidine ring but forms an angle of 12° with the best plane of the base. This is consistent with the orientation of the nitro group in 5-nitrouracil (14). The bond lengths in the pyrimidine ring are nearly identical in the base and in the nucleoside, with the exception that the C6 - Ni bond is shorter by 0.046 R whereas C6 - C5 and Ni - C2 are longer by 0.030 R in the nucleoside (Fig. 1, 2). The length of the C4 - 04 bond (1.232 A) is the same as it is in uridine and differs from that of 3-deazauridine (1.341 R) (15). 2. Structure of the ribose The ribose in the 5-nitrouridine has a C3'-endo conformation with Cl', C2', C4' and 01' in the plane and C3' out of the
931
Nucleic Acids Research
Fig. 1
Fig. 2
Structural representation of 5-nitrouridine (plot program ORTEP by C.K. Johnson (18)).
Structural formula of 5-nitrouridine, with the determined bond lengths and the bond and torgion angles. Standard deviation:a x=0.08 Al (x=C,N,0)
axxx=0.5
932
Nucleic Acids Research plane by 0.58 A (Fig. 1, 2). The Cl' - 01' bond is shorter by 0.071 i compared to the C4' - 01' bond, as is usually found in nucleoside structures. 3. Conformation about the glycosidic bond The conformation about the glycosidic bond is anti with a torsion angle C6 - Ni - Cl' - 01' of 25.60. The base is held in 05' this conformation by an "intramolecular interaction" H6 (Table 1). Atom 05' is positioned above the ribose and is "gauche" to both C3' - C4' and to 01' - C4' (Fig. 2). The conformation of the ribosyl moiety is similar to uridine, in which case the deviations from the ideal gauche conformation (+600 and -60°) are greater. 4. Packing of the molecules in the crystal The packing of the molecules in the crystal is portrayed in Fig. 3 and 4. If one sights in the direction of the c-axis (Fig. 3), then the bases are located at the left and right side separated by the ribose moieties. The sugars are kept in place by the hydrogen bonds listed in Table 1. In the direction of the a-axis (Fig. 4), one can observe that the base moieties are related by a screw axis and that they form an angle of 480 with respect to each other. The size of this angle indicates that stacking interactions between these bases are almost negligible. Bases within neighboring elementary cells in the b-direction are connected via a water molecule. The bases form no hydrophobic stacking; they are nearly perpendicular to the bc plane and form an angle of 660 with respect to the ac plane. The water is involved in a bifurcated donating hydrogen bond to 04 of the keto group and to 052 of the nitro group. In addition an accepting bond with the amide group of a second base is formed. Furthermore the water interacts with the 01' and 3'-OH group of the ribose moieties (Table 1). The one water molecule forms two donating and two accepting hydrogen bonds which are arranged in a nearly tetrahedral alignment around the oxygen atom. In this respect, the packing of the 5-nitrouridine differs from that of uridine which contains two symmetry-independent molecules in the asymmetric unit. Both pyrimidine rings form a fairly intimate but highly offset stacked pair (16).
933
Nucleic Acids Research X-H ---Y
H --- Y
[Al
X --- Y Li
lX-H-Y
C6 -H
---05'
2.446
3.333
159.7
C2'-H
---051
2.200
3.281
161.4
02'-H 05'-H
---05' ---03'
1.501 1.807
2.672 2.712
165.6 156.2
N3 -H ---OW 03'-H ---OW OW -HW1---01 ' OW -HW2---04 OW -HW2---052
1.952 1.703 2.176 2.428 2.201
3.025 2.763 2.806 3.432 2.963
173.5 143.2 134.2 161.3 128.3
Table 1
[0]
List of X-H---Y contacts divided into four groups: a) intramolecular "interaction" b) intermolecular "interaction" c) intermolecular hydrogen bonds between the ribose moieties d) hydrogen bonds with participation of the water molecule
DISCUSSION Recently, the structure of 5-nitro-1-(a-D-ribosyl-uronic acid)-uracil monohydrate was reported by Srikrishnan and Parthasarathy (1). The structure of the pyrimidine ring in this compound is practically identical with 5-nitrouridine. The base is, also, in the anti orientation with a torsion angle of 0 53.9°. This conformation is stabilized in both molecules by an intramolecular interaction H6 ... 05' which in the case of 5nitrouridine is increased to 3.333 R compared to 3.104 R in the uronic acid. It is always a matter of opinion whether to designate this interaction a hydrogen bond. The structure of the ribose and especially the packing of the molecules in the crystal turns out to be completely different. In 5-nitrouridine the ribose exists in the C3'-endo conformation whereas the uronic acid displays a C2'-endo - C3'-exo structure. The uronic acid - according to the pictures published by these authors - seems to form a sandwich type structure with a water molecule in between the two bases. The intercalation of the water molecule increases the base-base distance to 6.55 R. In the 5-nitrouridine structure, as determined by us, the
934
Nucleic Acids Research water molecule connects two consecutive bases by forming hydrogen bonds to the polar groups of both (Fig. 3, 4). If one sights perpendicular to the plane of the bases, which are connected by a translation along the c-axis, they are 7.76 R apart. If one draws a hypothetical line connecting the two Ni or C4 atoms, i. e. the outermost ring atoms, then the water molecule is placed outside of that "base stacking" by about 2 i in the direction of the a-axis (Fig. 4). Therefore, an intercalating water molecule was not found in this structure. It should be mentioned that the CD spectrum of Apno5U 5 shows major differences when compared to ApU, Apcl U or
Apbr5U.
Thus the electronic effect of the nitro group seems to influence the stacking of 5-nitrouridine in the crystal as well as the conformation of dinucleoside phosphates in solution (17).
Therefore, if a substituent in the 5-position increases the polarity of the base moiety of the nucleoside, solvent in-
Fig. 3
Packing of 5-nitrouridine within the crystal as viewed in the c-axis. The shading shows the ribosyl moieties. The hatched ellypsoids represent the oxygen atoms of the water molecules.
935
Nucleic Acids Research
Fig. 4
Packing of 5-nitrouridine within the crystal as viewed relative to the a-axis. The ribose moieties are omitted.
teraction could have a major impact on the stacking tendency of nucleosides. This should be kept in mind when the structural and functional consequences of 5-substitutioni in the wobble nucleoside of the anticodon of a tRNA are discussed. ACKNOWLEDGEMENT We wish to thank Dr. H. Vorbruggen, Schering AG. Berlin, for the nucleoside-tribenzoate. Furthermore we express our gratitude to O.E. Beck for his help in preparing the manuscript. The work was supported by the Fonds der Chemischen Industrie. REFERENCES 1 Srikrishnan, T. and Parthasarathy, R. (1976) Nature 264, 379-380 2 Barell, B.G. and Clark, B.F.C. (1974) in Handbook of Nucleic Acid Sequences, Joynson and Bruvvers Ltd., Oxford 3 Quigley, G.J. and Rich, A. (1976) Science 194, 796-806 4 Crick, F.H.C. (1966) J.Mol.Biol. 19, 548-555 5 Gray, M.W. (1976) Biochemistry 15, 3046-3051 6 Borek, J. (1971) in Physicochemical Characterisation of Oligonucleotides and Polynucleotides IFI/Plenum Corp. N.Y., Washington, London 7 Hawkinson, S.W. and Coulter, C.L. (1971) Acta Cryst. B 27, 34-42
936
Nucleic Acids Research 8
9 10 11 12 13 14 15 16
17 18
Kulakowska, I., Geller, M., Lesyng, B. and Wierzchowski, K.L. (1974) Biochim.Biophys. Acta 361, 119-130 Niedballa, U. and Vorbrtggen, H. (1974) J.Org.Chem. 39, 3668-3672 Wempen, I., Doerr, I.L., Kaplan, L. and Fox, J.J. (1960) J.Amer.Chem.Soc. 82, 1624-1629 Karle, J. and Karle, I.L. (1966) Acta Cryst. 21, 849-859 Germain, G. and Woolfson, M.M. (1968) Acta Cryst. B 24, 91-96 Sheldrick, G. not yet published Craven, B.M. (1967) Acta Cryst. 23, 376-383 Schwalbe, C.H., Gassen, H.G. and Saenger, W. (1972) Nature 238, 250 Green, E.A., Rosenstein, R.D., Shiono, R., Abraham, D.J., Trus, B.L. and Marsh, R.E. (1975) Acta Cryst. B 31, 102-107 Hillen, W. and Gassen, H.G. (manuscript in preparation) Johnson, C.K. (1965) ORNL-3794 (revised) Oak Ridge National Laboratory, Oak Ridge
937
