Search

The microsecond dynamics of the sugar moiety of A and T residues in DNA duplexes, ${\hbox{d}^{5'}}({}^{1}\hbox{C}^{2}\hbox{G}^{3}{\bf A}^{4}{\bf T}^{5}{\bf T}\break {}^{6}{\bf A}^{7}{\bf A}^{8}{\bf T}^{9}\hbox{C}^{10}\hbox{G})_{2}^{3'}$ (1b) and ${\hbox{d}^{5'}}({}^{1}\hbox{C}^{2}\hbox{C}^{3}{\bf A}^{4}{\bf T}^{5}{\bf T}^{6}{\bf A}^{7}{\bf A}^{8}{\bf T}^{9}\hbox{G}^{10}\hbox{G})_{2}^{3'}$ (2b), containing ¹³C/²H double-labelled 2'(R/S),5'(R/S)-²H2-1',2',3',4',5'-¹³C5-2'-deoxyribofuranose moieties (all labelled A and T are shown in bold), were studied using ¹³C nuclear spin relaxation measurements. An exchange contribution was detected in the transverse relaxation rates (R1ρ) of ¹³C of the labelled nucleotides. The comparison of the dynamics of various nucleotide residues of duplex 1b with those of the duplex 2b demonstrated that the replacement of the ²G·⁹C base pair in the former by a ²C·⁹G base pair in the latter alters the time-scale of motions in the AT tract. Moreover, the T residues show different microsecond dynamic behaviour to the A residues in the A·T base pairs. Since ²H nuclear magnetic spin relaxation (T1ρ) measurements of the same nucleotides show no dependence on the spin lock strength, it was concluded that the main mechanism of ²H T1ρ relaxation is quadrupolar. Although we observed a clear difference in the dynamic characteristics of the AT tract of the duplexes 1b and 2b [as evident from distinct differences in both spin lock dependent ¹³C relaxation (T1ρ) and in the amplitude of the exchange parameter amongst all deoxyadenosine nucleotide residues in both duplexes], we failed, however, to observe any difference in hydration behaviour in solution, thereby suggesting that there is no straightforward correlation in these two intrinsic dynamic properties of DNA duplex. It is noteworthy, however, that as the flexibility of the minor groove increases in both duplexes, we observe more long-lived water molecules around. Copyright © 2000 John Wiley & Sons, Ltd.

The T1 and T1ρ of deuterium in ¹³C/²H double-labeled 2'(R/S),5'(R/ S)-²H2 -1',2',3',4',5'- ¹³C5 -2'-deoxyadenosine and the corresponding thymidine derivative as well as in the non-uniformly labeled (shown in bold and underlined) DNA duplex, d^5'(¹C²G³A⁴T⁵T⁶A⁷A⁸T⁹C¹⁰G)2^3', have been determined for the first time. These double-labelled nucleoside blocks have a special feature in that the geminal 2'–2" and 5'–5" proton–proton couplings are eliminated by replacement with diastereomeric deuterium at C-2' and C-5' centers. This uniquely enables us to perform deuterium relaxation measurement experiments through selective polarization transfer, ¹H–¹³C– ²H–¹³C–¹H at C-2' and C-5' centers, thereby allowing filtration of all other naturally abundant methylene- and methine-¹³C as well as enriched methine- ¹³C fragments. Comparison of T1 and T 1ρ of ²H in double-labeled ( ¹³C/²H) 2'-deoxyadenosine and thymidine with that of the non-uniformly labeled DNA duplex, d^5'(¹C²G³ A⁴T⁵T⁶A⁷A⁸T⁹C¹⁰G)2^3', shows that the dynamics of various nucleotide residues are indeed non-uniform. Copyright © 1999 John Wiley & Sons, Ltd.

The effect of ²H on ¹³C longitudinal (T1) and transverse (T2) relaxation parameters was determined for the first time for diastereospecifically deuterium-labeled nucleosides, which are used as the building blocks for non-uniform isotope labeling for the solution NMR structure determination of the large biologically functional oligo-DNA and -RNA (‘NMR window’ approach, ref. 7). It emerged that the T1 and T2 of the deuterated methine carbon in the diastereospecifically deuterium-labeled nucleoside 9 could be used as the correction term to give the monoexponential decay of ¹³C longitudinal and transverse magnetization of the constituent ¹H–¹³C–²H group. The correlation time derived from this corrected T1 of the methylene carbon corresponds well with the correlation time obtained from deuterium relaxation study. The extreme narrowing limit (ωτc≪1) where dipole–dipole (DD) relaxation of ¹³C and quadrupole (Q) relaxation of ²H are related by T1^DD/T2^DD≈1 and T1^Q/T2^Q≈1 was used to demonstrate the above conclusion. The difference in the observable T1 and T2 in various methylene and methine-type carbons with either fully protonated or diastereospecifically deuterated nucleosides 1–14 allowed the estimation of the contribution of the alternative relaxation pathways other than DD relaxation. It was found by comparison of the T1 relaxation of the quaternary carbon with the methine carbon (¹³C–²H) or (¹³C–¹H) in compound 2 that the contribution of the intermolecular and intramolecular relaxations of ¹³C with protons that are two bonds away is larger than DD(¹³C–²H), and the sum of all these contributions define the T1 of the methine carbon (¹³C–²H). The observed difference between the experimental T1 and T2 of the methine carbon is attributed to the cross-correlation between DD(¹³C–²H) and Q(²H) relaxation, which is consistent with recent theoretical predictions. For T2 measurement, the decoupling of deuterium with 0.6–2.5 kHz power during the echo period by WALTZ does not effectively eliminate the DD(¹³C–²H)–Q(²H) cross-correlation for the methine carbon. The suppression of this DD(¹³C–²H)–Q(²H) cross-correlation was, however, more effective by applying a 180° deuterium pulse in the middle of the short (0.5 ms) echo period (compare T2 of 3.91s and 0.3s, respectively, at 294 K using these two different decoupling procedures). The comparison of the observed T1 and T2 relaxations of the methylene carbon shows that they are indeed very close. The various contributions of the methine carbon relaxation such as DD(¹³C–²H), intermolecular and cross-correlation, DD(¹³C–¹H)–Q(²H), to the relaxation of the methylene carbon were ca. 15% in T1 and ca. 25% in T2. © 1998 John Wiley & Sons, Ltd.