Flanking A·T basepairs destabilize the B(∗) conformation of DNA A-tracts

Abstract

Capillary electrophoresis has been used to characterize the interaction of monovalent cations with 26-basepair DNA oligomers containing A-tracts embedded in flanking sequences with different basepair compositions. A 26-basepair random-sequence oligomer was used as the reference; lithium and tetrabutylammonium (TBA(+)) ions were used as the probe ions. The free solution mobilities of the A-tract and random-sequence oligomers were identical in solutions containing <∼ 100 mM cation. At higher cation concentrations, the A-tract oligomers migrated faster than the reference oligomer in TBA(+) and slower than the reference in Li(+). Hence, cations of different sizes can interact very differently with DNA A-tracts. The increased mobilities observed in TBA(+) suggest that the large hydrophobic TBA(+) ions are preferentially excluded from the vicinity of the A-tract minor groove, increasing the effective net charge of the A-tract oligomers and increasing the mobility. By contrast, Li(+) ions decrease the mobility of A-tract oligomers because of the preferential localization of Li(+) ions in the narrow A-tract minor groove. Embedding the A-tracts in AT-rich flanking sequences markedly alters preferential interactions of monovalent cations with the B(∗) conformation. Hence, A-tracts embedded in genomic DNA may or may not interact preferentially with monovalent cations, depending on the relative number of A · T basepairs in the flanking sequences.

Figure 1

(A) Dependence of the mobility ratios, μ(A-tract)/μ(random), observed for: (•,○), A4T1in(0); (▲,Δ), A4T1in(6); and (♦,⋄), A4T1in(10) as a function of cation concentration in BGEs containing: TBA⁺ (solid symbols) or Li⁺ (open symbols). The error bars correspond to the standard deviation of replicate measurements in each BGE. (B) Fractional change of the mobility ratios observed for A4T1in(0) and A4T1in(6) as a function of cation concentration; the transition midpoint occurs at 150 ± 10 mM cation.

Figure 2

Dependence of the mobility ratios observed for oligomer A4T1in(6) on temperature in: (○), 300 mM Li⁺; and (•), 300 mM TBA⁺. The midpoints of the thermal transitions are 39 ± 1°C in Li⁺ and 27 ± 1°C in TBA⁺.

Figure 3

Dependence of the average mobility differences observed per A-tract, in m.u., on A-tract length. The BGEs contained 300 mM TBA⁺ (solid symbols); or 300 mM Li⁺ (open symbols). The error bars represent the standard deviation of the mobility differences from the average; the standard deviations of the individual mobility differences were smaller than the sizes of the symbols. The straight line through the TBA⁺ data was drawn by linear regression (r² = 0.985); the Li⁺ data were fitted to a three-parameter exponential decay (r² = 0.839).

Figure 4

Mobility differences, in m.u., observed for oligomers A4T1in(0) to A4T1in(11) as a function of the number of flanking A·T basepairs, in BGEs containing: (•), 300 mM TBA⁺; or (○), 300 mM Li⁺. Error bars corresponding to the reproducibility of the measurements are smaller than the sizes of the symbols.