Backbone charge transport in double-stranded DNA.

Understanding charge transport in DNA molecules is a long-standing problem of fundamental importance across disciplines1,2. It is also of great technological interest due to DNA's ability to form versatile and complex programmable structures. Charge transport in DNA-based junctions has been reported using a wide variety of set-ups2-4, but experiments so far have yielded seemingly contradictory results that range from insulating5-8 or semiconducting9,10 to metallic-like behaviour11. As a result, the intrinsic charge transport mechanism in molecular junction set-ups is not well understood, which is mainly due to the lack of techniques to form reproducible and stable contacts with individual long DNA molecules. Here we report charge-transport measurements through single 30-nm-long double-stranded DNA (dsDNA) molecules with an experimental set-up that enables us to address individual molecules repeatedly and to measure the current-voltage characteristics from 5 K up to room temperature. Strikingly, we observed very high currents of tens of nanoamperes, which flowed through both homogeneous and non-homogeneous base-pair sequences. The currents are fairly temperature independent in the range 5-60 K and show a power-law decrease with temperature above 60 K, which is reminiscent of charge transport in organic crystals. Moreover, we show that the presence of even a single discontinuity ('nick') in both strands that compose the dsDNA leads to complete suppression of the current, which suggests that the backbones mediate the long-distance conduction in dsDNA, contrary to the common wisdom in DNA electronics2-4.

Advances in Synthesis and Measurement of Charge Transport in DNA-Based Derivatives.

Charge transport through molecular structures is interesting both scientifically and technologically. To date, DNA is the only type of polymer that transports significant currents over distances of more than a few nanometers in individual molecules. For molecular electronics, DNA derivatives are by far more promising than native DNA due to their improved charge-transport properties. Here, the synthesis of several unique DNA derivatives along with electrical characterization and theoretical models is surveyed. The derivatives include double stranded poly(G)-poly(C) DNA molecules, four stranded G4-DNA, metal-DNA hybrid molecular wires, and other DNA molecules that are modified either at the bases or at the backbone. The electrical characteristics of these nanostructures, studied experimentally by electrostatic force microscopy, conductive atomic force microscopy, and scanning tunneling microscopy and spectroscopy, are reviewed.

Highly Conductive Thin Uniform Gold-Coated DNA Nanowires.

Over the past decades, DNA, the carrier of genetic information, has been used by researchers as a structural template material. Watson-Crick base pairing enables the formation of complex 2D and 3D structures from DNA through self-assembly. Various methods have been developed to functionalize these structures for numerous utilities. Metallization of DNA has attracted much attention as a means of forming conductive nanostructures. Nevertheless, most of the metallized DNA wires reported so far suffer from irregularity and lack of end-to-end electrical connectivity. An effective technique for formation of thin gold-coated DNA wires that overcomes these drawbacks is developed and presented here. A conductive atomic force microscopy setup, which is suitable for measuring tens to thousands of nanometer long molecules and wires, is used to characterize these DNA-based nanowires. The wires reported here are the narrowest gold-coated DNA wires that display long-range conductivity. The measurements presented show that the conductivity is limited by defects, and that thicker gold coating reduces the number of defects and increases the conductive length. This preparation method enables the formation of molecular wires with dimensions and uniformity that are much more suitable for DNA-based molecular electronics.

Atomic force microscopy characterization of kinase-mediated phosphorylation of a peptide monolayer.

We describe the detailed microscopic changes in a peptide monolayer following kinase-mediated phosphorylation. A reversible electrochemical transformation was observed using square wave voltammetry (SWV) in the reversible cycle of peptide phosphorylation by ERK2 followed by dephosphorylation by alkaline phosphatase. A newly developed method for analyzing local roughness, measured by atomic force microscope (AFM), showed a bimodal distribution. This may indicate either a hole-formation mechanism and/or regions on the surface in which the peptide changed its conformation upon phosphorylation, resulting in increased roughness and current. Our results provide the mechanistic basis for developing biosensors for detecting kinase-mediated phosphorylation in disease.

Integrating proteomics with electrochemistry for identifying kinase biomarkers.

We present an integrated approach for highly sensitive identification and validation of substrate-specific kinases as cancer biomarkers. Our approach combines phosphoproteomics for high throughput cancer-related biomarker discovery from patient tissues and an impedimetric kinase activity biosensor for sensitive validation. Using non-small-cell lung cancer (NSCLC) as a proof-of-concept study, label-free quantitative phosphoproteomic analysis of a pair of cancerous and its adjacent normal tissues revealed 198 phosphoproteins that are over-phosphorylated in NSCLC. Among the differentially regulated phosphorylation sites, the most significant alteration was in residue S165 in the Hepatoma Derived Growth Factor (HDGF) protein. Hence, HDGF was selected as a model system for the electrochemical studies. Further motif-based analysis of this altered phosphorylation site revealed that extracellular-signal-regulated kinase 1/2 (ERK1/2) are most likely to be the corresponding kinases. For validation of the kinase-substrate pair, densely packed peptide monolayers corresponding to the HDGF phosphorylation site were coupled to a gold electrode. Phosphorylation of the monolayer by ERK2 and dephosphorylation by alkaline phosphatase (AP) were detected by electrochemical impedance spectroscopy (EIS) and surface roughness analysis. Compared to other methods for quantification of kinase concentration, this label-free electrochemical assay offers the advantages of ultra-sensitivity as well as higher specificity for the detection of cancer-related kinase-substrate pair. With implementation of multiple kinase-substrate biomarker pairs, we expect this integrated approach to become a high throughput platform for discovery and validation of phosphorylation-mediated biomarkers.