S.S. Wijmenga, B.N.M. van Buuren/Progress in Nuclear Magnetic Resonance Spectroscopy 32(1998)287-387 also show differences depending on whether they are di(1: 4). Only d(2; 4)differs significantly present in an A- or B-type helix between S-type and N-type conformers, with As can be seen from Table 1 approximately 60 d(2;4 4. 2A for the s-type conformer distances per residue can in principle be measured (pseudorotation angle P= 160)and dH(2: 4)= The number of distances that are constant within 2.8A for the N-type conformer (P=10 +0.2 A is rather high. They represent about 26% Although it is in principle possible to determine of the total number of measurable distances Their the sugar conformation from the di(2": 4")distance percentage is even higher for the intra-nucleotide dis- the accuracy of the determination is limited. The tances, of which they represent about 57%(16 out of di(2: 4) distance is difficult to determine from 28). The distances that convey relevant structural NOE intensities because of spin diffusion effects information (in helices) and are reasonably well due to the close proximity of the H2"and H2 accessible by NMR represent less then half (48%)of protons. Also note that in RNA the H2 proton is the total number of distances, while only 20 are dif- bsent, so that these sugar distances cannot be used ferent between A-and B-type helices(33%). Note at all to determine the puckering. The distance also the small number of structurally very important di(1: 4)is almost identical for N-type and s cross-strand and sequential distances involving pe sugars 3.4 A), but has a lower value for exchanging protons which establish base pairing sugar rings with an intermediate pseudorotation (8%), and the small number of cross-strand distances angle, di(1: 4)=2.6A for P=90. Here again involving non-exchanging protons (3%) On the other spin diffusion can adversely affect the accuracy hand, sequential sugar-to-base and sugar-to-sugar dis- distance of the determination tances, which are so important for establishing base 3. The distances dA(3: 575)depend only weakly on stacking and defining the phosphate backbone, are the sugar ring conformation, but significantly on both relatively large in number(20%). The former the y torsion angle, while the distances di 44; 57 e mostly reasonably well accessible by NMR 6) only depend on the y torsion angle. The whereas the latter are extremely difficult to establish distances di(374; 5/5")therefore allow the deter- Thus, a rather uneven spread in the short distances is mination of the torsion angle [62, 63]. This can be found through the chemical structure. As a conse- done in conjunction with relevant J-couplings(see quence, important structural features such as base Section 5). Given an uncertainty in these distances ing often hinge on the presence of a particular of +0.2 A, they do not discriminate well between NOE contact reflecting one short distance the different ranges of the y torsion angle, in ticular when an equilibrium between g* and g 4.2. Overview of structurally important intra tamers exists. The distances di(2/2 575) nucleotide distances depend on both the sugar puckering and the torsion angle y, but their dependence is weak, and they are The intra-nucleotide distances in dna and rna of the order of 5 to 6 A [62] can conveniently be subdivided according to the 4. The distance between HI and H8/6, d(1: 6/8) categories indicated in Table 1, i.e. (1)conformation depends only on the glycosidic torsion angle x. It independent distances,(2)distances between sugar thus provides a means for determining this torsion protons, (3)distances between H2/2/3/4 and H5/ angle. However, the maximum difference in the 5"(4)distances between HI'through H575"and base values of di(1: 6/8)for x in the syn domain (x= 60°) and in the anti domain(x=240°) is only about 1.2 A. Given that in practice the uncertainty 1. The conformation independent distances are: the in the distance determination from noe data is of geminal proton distances, d 2, 2 )and d 5, 5) the order of±0.2Ato±0.5A. it is to be of 1.8A, d( 5: 6)(= 2.45 A)in Cytosine and expected that the use of d; (1: 6/8)is a rather impre Uracyl, and d 6, M)in Thymidine cise means to determine the x torsion angle. The 2. The distances within the sugar ring are all indepen- other sugar proton to base proton distances, dH(2/ dent of its conformation, except for di(2: 4)and 21314: 6/8), depend on both the sugar puckeringalso show differences depending on whether they are present in an A- or B-type helix. As can be seen from Table 1 approximately 60 distances per residue can in principle be measured. The number of distances that are constant within 6 0:2 A˚ is rather high. They represent about 26% of the total number of measurable distances. Their percentage is even higher for the intra-nucleotide dis￾tances, of which they represent about 57% (16 out of 28). The distances that convey relevant structural information (in helices) and are reasonably well accessible by NMR represent less then half (48%) of the total number of distances, while only 20 are dif￾ferent between A- and B-type helices (33%). Note also the small number of structurally very important cross-strand and sequential distances involving exchanging protons which establish base pairing (8%), and the small number of cross-strand distances involving non-exchanging protons (3%). On the other hand, sequential sugar-to-base and sugar-to-sugar dis￾tances, which are so important for establishing base stacking and defining the phosphate backbone, are both relatively large in number (20%). The former are mostly reasonably well accessible by NMR, whereas the latter are extremely difficult to establish. Thus, a rather uneven spread in the short distances is found through the chemical structure. As a conse￾quence, important structural features such as base pairing often hinge on the presence of a particular NOE contact reflecting one short distance. 4.2. Overview of structurally important intra￾nucleotide distances The intra-nucleotide distances in DNA and RNA can conveniently be subdivided according to the categories indicated in Table 1, i.e. (1) conformation independent distances, (2) distances between sugar protons, (3) distances between H29/20/39/49 and H59/ 50, (4) distances between H19 through H59/50 and base protons. 1. The conformation independent distances are: the geminal proton distances, di(29;20) and di(59;50), of 1.8 A˚ , di(5;6) ( ¼ 2.45 A˚ ) in Cytosine and Uracyl, and di(6,M) in Thymidine. 2. The distances within the sugar ring are all indepen￾dent of its conformation, except for di(20;49) and di(19;49). Only di(20;49) differs significantly between S-type and N-type conformers, with di(20;49) ¼ 4.2 A˚ for the S-type conformer (pseudorotation angle P ¼ 1608) and di(20;49) ¼ 2.8 A˚ for the N-type conformer (P ¼ 108). Although it is in principle possible to determine the sugar conformation from the di(20;49) distance, the accuracy of the determination is limited. The di(20;49) distance is difficult to determine from NOE intensities because of spin diffusion effects, due to the close proximity of the H29 and H20 protons. Also note that in RNA the H20 proton is absent, so that these sugar distances cannot be used at all to determine the puckering. The distance di(19;49) is almost identical for N-type and S￾type sugars (3.4 A˚ ), but has a lower value for sugar rings with an intermediate pseudorotation angle, di(19;49) ¼ 2.6 A˚ for P ¼ 908. Here again spin diffusion can adversely affect the accuracy distance of the determination. 3. The distances di(39; 59/50) depend only weakly on the sugar ring conformation, but significantly on the g torsion angle, while the distances di(49; 59/ 50) only depend on the g torsion angle. The distances di(39/49; 59/50) therefore allow the deter￾mination of the torsion angle [62,63]. This can be done in conjunction with relevant J-couplings (see Section 5). Given an uncertainty in these distances of 6 0.2 A˚ , they do not discriminate well between the different ranges of the g torsion angle, in par￾ticular when an equilibrium between gþ and gt rotamers exists. The distances di(29/20; 59/50) depend on both the sugar puckering and the torsion angle g, but their dependence is weak, and they are of the order of 5 to 6 A˚ [62]. 4. The distance between H19 and H8/6, di(19;6/8), depends only on the glycosidic torsion angle x. It thus provides a means for determining this torsion angle. However, the maximum difference in the values of di(19;6/8) for x in the syn domain (x ¼ 608) and in the anti domain (x ¼ 2408) is only about 1.2 A˚ . Given that in practice the uncertainty in the distance determination from NOE data is of the order of 6 0.2 A˚ to 6 0.5 A˚ , it is to be expected that the use of di(19;6/8) is a rather impre￾cise means to determine the x torsion angle. The other sugar proton to base proton distances, di(29/ 20/39/49;6/8), depend on both the sugar puckering 294 S.S. Wijmenga, B.N.M. van Buuren/Progress in Nuclear Magnetic Resonance Spectroscopy 32 (1998) 287–387