Evaluation of Single-Stranded Nucleic Acids as Carriers in the DNA-Directed Assembly of Macromolecules

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Abstract
Current developments in nanosciences indicate that the self-assembly of macromolecules, such as proteins or metallic nanoclusters, can be conveniently achieved by means of nucleic acid hybridization. Within this context, we here report on the evaluation of single-stranded nucleic acids to be utilized as carrier backbones in DNA-directed self-assembly. A microplate solid-phase hybridization assay is described which allows rapid experimental determination of the hybridization efficiencies of various sequence stretches within a given nucleic acid carrier strand. As demonstrated for two DNA fragments of different sequence, the binding efficiencies of several oligonucleotides depend on the formation of specific secondary structure elements within the carrier molecule. A correlation of sequence-specific hybridization capability with modeled secondary structure is also obvious from experiments using the fluorescence gel-shift analysis. Electrophoretic studies on the employment of helper oligonucleotides in the formation of supramolecular conjugates of several oligonucleotide-tagged proteins indicate, that structural constraints can be minimized by disruption of intramolecular secondary structures of the carrier molecule. To estimate the influences of the chemical nature of the carrier, gel-shift experiments are carried out to compare a 170mer RNA molecule with its DNA analogue. Ternary aggregates, containing two protein components bound to the carrier, are formed with a greater efficiency on the DNA instead of the RNA carrier backbone.