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Volker J.,Rutgers University | Eric Plum G.,Rutgers University | Eric Plum G.,IBET Inc. | Klump H.H.,University of Cape Town | And 2 more authors.
Biopolymers | Year: 2010

Clusters of closely spaced oxidative DNA lesions present challenges to the cellular repair machinery. When located in opposing strands, base excision repair (BER) of such lesions can lead to double strand DNA breaks (DSB). Activation of BER and DSB repair pathways has been implicated in inducing enhanced expansion of triplet repeat sequences. We show here that energy coupling between distal lesions (8oxodG and/or abasic sites) in opposing DNA strands can be modulated by a triplet repeat bulge loop located between the lesion sites. We find this modulation to be dependent on the identity of the lesions (8oxodG vs. abasic site) and the positions of the lesions (upstream vs. downstream) relative to the intervening bulge loop domain. We discuss how such bulge loop-mediated lesion crosstalk might influence repair processes, while favoring DNA expansion, the genotype of triplet repeat diseases. © 2009 Wiley Periodicals, Inc.

Braunlin W.,Rutgers University | Braunlin W.,Rational Affinity Devices LLC | Volker J.,Rutgers University | Plum G.E.,IBET Inc. | And 2 more authors.
Biopolymers | Year: 2013

We describe a novel hybridization assay that employs a unique class of energy tunable, bulge loop-containing competitor strands (C*) that hybridize to a probe strand (P). Such initial «pre-binding» of a probe strand modulates its effective «availability» for hybridizing to a target site (T). More generally, the assay described here is based on competitive binding equilibria for a common probe strand (P) between such tunable competitor strands (C*) and a target strand (T).We demonstrate that loop variable, energy tunable families of C*P complexes exhibit enhanced discrimination between targets and mismatched targets, thereby reducing false positives/negatives. We refer to a C*P complex between a C* competitor single strand and the probe strand as a "tuning fork," since the C* strand exhibits branch points (forks) at the duplex-bulge interfaces within the complex. By varying the loop to create families of such "tuning forks," one can construct C*P "energy ladders" capable of resolving small differences within the target that may be of biological/functional consequence. The methodology further allows quantification of target strand concentrations, a determination heretofore not readily available by conventional hybridization assays. The dual ability of this tunable assay to discriminate and quantitate targets provides the basis for developing a technology we refer to as a "DNA Meter." Here we present data that establish proof-of-principle for an in solution version of such a DNA Meter. We envision future applications of this tunable assay that incorporate surface bound/spatially resolved DNA arrays to yield enhanced discrimination and sensitivity. © 2013 Wiley Periodicals, Inc.

Volker J.,Rutgers University | Gindikin V.,Rutgers University | Klump H.H.,University of Cape Town | Plum G.E.,IBET Inc. | And 2 more authors.
Journal of the American Chemical Society | Year: 2012

DNA repeat domains can form ensembles of canonical and noncanonical states, including stable and metastable DNA secondary structures. Such sequence-induced structural diversity creates complex conformational landscapes for DNA processing pathways, including those triplet expansion events that accompany replication, recombination, and/or repair. Here we demonstrate further levels of conformational complexity within repeat domains. Specifically, we show that bulge loop structures within an extended repeat domain can form dynamic ensembles containing a distribution of loop positions, thereby yielding families of positional loop isomers, which we designate as "rollamers". Our fluorescence, absorbance, and calorimetric data are consistent with loop migration/translocation between sites within the repeat domain ("rollamerization"). We demonstrate that such "rollameric" migration of bulge loops within repeat sequences can invade and disrupt previously formed base-paired domains via an isoenthalpic, entropy-driven process. We further demonstrate that destabilizing abasic lesions alter the loop distributions so as to favor "rollamers" with the lesion positioned at the duplex/loop junction, sites where the flexibility of the abasic "universal hinge" relaxes unfavorable interactions and/or facilitates topological accommodation. Another strategic siting of an abasic site induces directed loop migration toward denaturing domains, a phenomenon that merges destabilizing domains. In the aggregate, our data reveal that dynamic ensembles within repeat domains profoundly impact the overall energetics of such DNA constructs as well as the distribution of states by which they denature/renature. These static and dynamic influences within triplet repeat domains expand the conformational space available for selection and targeting by the DNA processing machinery. We propose that such dynamic ensembles and their associated impact on DNA properties influence pathways that lead to DNA expansion. © 2012 American Chemical Society.

Volker J.,Rutgers University | Plum G.E.,IBET Inc. | Gindikin V.,Rutgers University | Klump H.H.,University of Cape Town | And 2 more authors.
Biopolymers | Year: 2014

Repetitive DNA sequences exhibit complex structural and energy landscapes, populated by metastable, noncanonical states, that favor expansion and deletion events correlated with disease phenotypes. To probe the origins of such genotype-phenotype linkages, we report the impact of sequence and repeat number on properties of (CNG) repeat bulge loops. We find the stability of duplexes with a repeat bulge loop is controlled by two opposing effects; a loop junction-dependent destabilization of the underlying double helix, and a self-structure dependent stabilization of the repeat bulge loop. For small bulge loops, destabilization of the underlying double helix overwhelms any favorable contribution from loop self-structure. As bulge loop size increases, the stabilizing loop structure contribution dominates. The role of sequence on repeat loop stability can be understood in terms of its impact on the opposing influences of junction formation and loop structure. The nature of the bulge loop affects the thermodynamics of these two contributions differently, resulting in unique differences in repeat size-dependent minima in the overall enthalpy, entropy, and free energy changes. Our results define factors that control repeat bulge loop formation; knowledge required to understand how this helix imperfection is linked to DNA expansion, deletion, and disease phenotypes. Copyright © 2013 Wiley Periodicals, Inc.

Volker J.,Rutgers University | Eric Plum G.,Rutgers University | Eric Plum G.,IBET Inc. | Klump H.H.,University of Cape Town | And 2 more authors.
Journal of the American Chemical Society | Year: 2010

Energy coupling between distal DNA domains may have profound regulatory consequences for biological processes, allowing for allosteric control of nucleic acid function. Repair of oxidative lesions at or near triplet repeat domains can enhance DNA expansion events that result in debilitating disease states. We report here position, distance, and lesion-dependent energy crosstalk between pairs of lesions in a triplet repeat bulge loop and an adjacent duplex domain. We discuss the implications of such coupled communication between lesions in distal loop and duplex domains for lesion repair and DNA expansion associated with diseases "Figure Presented". Copyright © 2010 American Chemical Society.

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