Detection of internal and overall dynamics of a two-atom-tethered spin-labeled DNA

Abstract

DNA motions consist of several components which couple, making their investigation difficult. This study describes an approach for obtaining dynamical information by EPR when spin-labeled nucleic acids are examined. The analysis is accomplished by implementing two motional models. The first model (i.e., dynamic cylinder model) views the spin-labeled helix as a diffusing cylinder containing internal dynamics which are characterized by an order parameter. The second model (i.e., base disk model) provides correlation times describing the diffusion of the spin-labeled base. In each model, the nitroxide motion consists of both global and internal contributions. Dynamic cylinder and base disk simulations of four duplexes containing nitroxides attached to thymidine by a two-atom tether (DUMTA)-(dT)7DUMTA-(dT)7.(dA)15, [(dT)7DUMTA(dT)7]2.(dA)30, [(dT)7DUMTA(dT)7]3.(dA)45, and [(dT)7DUMTA(dT)7]m.-(dA)n--demonstrate the useful application of this approach. From dynamic cylinder simulations, the order parameter for internal motions is found to be independent of the helix length (S = 0.32 +/- 0.01). Previous base disk simulations of a DNA 26mer and polymer labeled with a five-atom-tethered nitroxide seemed to indicate that tau perpendicular was only sensitive to internal dynamics. Results from base disk simulations of DUMTA-labeled DNA indicate that the perpendicular component of the base disk correlation time (tau perpendicular = 1.4-6.2 ns) is sensitive to global dynamics. Thus, tau perpendicular is a quantitative indicator of both internal and global dynamics. Comparison of the two models reveals that tau perpendicular infinity S2 tau rb, where tau rb represents the rigid-body diffusion of the DNA helix. This relationship between S and tau perpendicular provides a framework for studying conformational changes and size-dependent phenomena in spin-labeled nucleic acids. Application of the dynamic cylinder model to a B-Z transition generates distinct values of S for each of the conformations, indicating that Z-DNA is more rigid than B-DNA.

Site-directed Spin Labeling Studies on Nucleic Acid Structure and Dynamics

“…The Bobst group developed a base disk model, in which the spin-labeled base is considered an axially symmetric diffusing system (25,29). The principle motion is characterized by two correlation times: τ ‖ describes rotational diffusion about the principle axis as defined by the average position of the bonds that connect the base to the nitroxide, and τ ⊥ describes the motion about the axis perpendicular to the tether bond.…”

“…The mean-square oscillation amplitude is directly related to the order parameter, but the simulation does not produce any estimation of the correlation time for the internal dynamics. For nitroxides that are more flexibly attached to bases within a DNA duplex (e.g., Table I, entry 5), the Bobst group has taken a similar approach and developed a dynamic cylinder model (29), which yields an order parameter that describes the amplitude of the nitroxide motion. It was suggested that the order parameter reports on both the base dynamics and the motion of the nitroxide with respect to the base.…”

“…Data on B-form DNA duplexes were simulated using a model that characterizes nitroxide motions by two correlation times, τ ‖ and τ ⊥ (Section V.B.2) (25,29,101,109). τ ‖ was found to be tether dependent, was less than 1 ns, and was attributable to nitroxide motion that was independent of base motion (11).…”

“…τ ⊥ was thought to represent motions including twisting and tilting of the individual base, collective bending and twisting of the DNA segment, and global tumbling of DNA (25,29,101,109). Values of τ ⊥ from all double-stranded systems were reported to be in the 1-10 ns range (11,25,29,101,109), leading to the conclusion that there are significant base motions and these motions contribute to the dynamics of the nitroxide. However, questions have been raised as to the degree to which τ ⊥ is influenced by the motion of the tether versus that of the base (see below) (14).…”

“…There has been debate over whether the rigidly coupled nitroxides alter the behavior of the base, and if so, to what degree (11,14,29,101,109). On the other hand, for the flexible nitroxides, questions have been raised regarding whether motions of the flexible linker lead to an overestimation of base dynamics (14).…”