Mono- and bilayer smectic liquid crystal ordering in dense solutions of “gapped” DNA duplexes

Abstract

Although its mesomorphic properties have been studied for many years, only recently has the molecule of life begun to reveal the true range of its rich liquid crystalline behavior. End-to-end interactions between concentrated, ultrashort DNA duplexes—driving the self-assembly of aggregates that organize into liquid crystal phases—and the incorporation of flexible single-stranded “gaps” in otherwise fully paired duplexes—producing clear evidence of an elementary lamellar (smectic-A) phase in DNA solutions—are two exciting developments that have opened avenues for discovery. Here, we report on a wider investigation of the nature and temperature dependence of smectic ordering in concentrated solutions of various “gapped” DNA (GDNA) constructs. We examine symmetric GDNA constructs consisting of two 48-base pair duplex segments bridged by a single-stranded sequence of 2 to 20 thymine bases. Two distinct smectic layer structures are observed for DNA concentration in the range ∼230to∼280 mg/mL. One exhibits an interlayer periodicity comparable with two-duplex lengths (“bilayer” structure), and the other has a period similar to a single-duplex length (“monolayer” structure). The bilayer structure is observed for gap length ≳10 bases and melts into the cholesteric phase at a temperature between 30 °C and 35 °C. The monolayer structure predominates for gap length ≲10 bases and persists to >40 °C. We discuss models for the two layer structures and mechanisms for their stability. We also report results for asymmetric gapped constructs and for constructs with terminal overhangs, which further support the model layer structures.

Stability of End-to-End Base Stacking Interactions in Highly Concentrated DNA Solutions

“…For The GDNA synthesis process, sample loading procedure into the borosilicate X-ray capillaries, and details of the SAXS measurements are described in the Supporting Information and in previous publications. 10,11 Nomenclature for Stacking Energies. The "AT−AT" construct in Figure 1A has AT as the terminal base pair on both duplex arms, which results in AT−AT type stacking between neighboring GDNA constructs.…”

“…The introduction of a flexible single-stranded DNA spacer (“gap”) between two rod-like duplexes, producing a “gapped” DNA (GDNA) construct, has been the key for observing elementary layered (smectic) phases in concentrated DNA solutions. , In addition to the DNA concentration (cDNA), the layer structure is sensitive to gap length and temperature. − When the gap length is ≥10 nucleotides (nt), a bilayer structure composed of stacked duplexes prevails at ambient temperatures. The spacing between the bilayers slightly exceeds two duplex lengths, suggesting that attractive enthalpic interactions between blunt duplex ends and a net conformational entropy gain from segregating flexible “gap” segments act together to stabilize the bilayer stacking.…”

Stability of End-to-End Base Stacking Interactions in Highly Concentrated DNA Solutions

“…The values of 𝑑𝑑 are slightly greater than the 32.6 nm length of two 48-bp duplexes, and are consistent with a bilayer smectic structure. 10,11 In addition, a sharp peak is recorded at wider angle. The position of this peak is consistent with the first order diffraction (wavenumber qw) from hexagonal lateral packing of the duplexes in the smectic-B phase of GDNA constructs previously reported at similar concentration and temperature.…”

“…1D-F have an effective end-end interaction that is a hybrid of the component end-end stackings. If, on the other hand, the bilayer structure is formed from stacking extended (“unfolded”) constructs, or if the folded pairs are not discrete ( e.g ., single ends of one folded construct paired with single ends of two others) 10 , this assumption is questionable, and the bilayer melting may instead be determined by some other combination(s) of the non-degenerate end-end interaction energies. In any case, given their limited resolution, the present data are adequately described by an analysis based on a two-step model.…”

Liquid Crystalline Systems from Nature and Interaction of Living Organisms with Liquid Crystals

“…[ 61,62 ] This “hidden” smectic phase of DNA is finally realized by using “Gapped” DNA duplex (G‐duplex) in which single‐stranded DNA (ssDNA) joins double‐stranded DNA (dsDNA) regions, and thus increases molecular flexibility (Figure 4c). [ 63,64 ] It is an interesting result because the increased flexibility suppresses formation of a smectic layer. [ 65 ] In this phenomenon, however, the end‐to‐end adhesion also has an important influence.…”