Precisely patterning proteins and other molecules at the nanoscale is crucial to future biosensing and optoelectronic applications. One- and two-dimensional DNA nanoconstructs have proven to be useful scaffolds for nanopatterning. This paper demonstrates the application of nitrilotriacetic acid (NTA) forming chelate complexes to localize histidine (His) tagged proteins via Ni(2+) ions onto DNA based structures. Particularly, enhanced green fluorescent protein (EGFP) was directed to specific surface locations on a designed DNA Origami nanoconstruct, and the resulting EGFP nanopattern was visualized using atomic force microscopy (AFM).
Nanomechanical devices hold the promise of controlling structure and performing exquisitely fine measurements on the molecular scale. We previously have reported three nanoscale devices that were built from DNA. The first device [1] extrudes or withdraws a portion of a DNA cruciform from a cyclic molecule in response to the presence of an intercalator. The second device is powered by the B-Z transition: [2] it switches a DNA domain from one side of a double-helical shaft to the other in response to the presence or absence of [Co(NH 3 ) 6 ] 3+ in its environment. The third device is sequence-dependent; [3] it changes structure as a function of DNA strands that are in solution with it. All of these devices change their states in response to an external stimulus, but they do not report new information about any other molecular system. Here we describe a new DNA device that changes shape when a DNA-distorting protein is added to the system. In this case, we used E. coli integration host factor (IHF), which bends DNA significantly when it binds to an appropriate recognition site.[4] Thus, we have developed a nanomechanical device for which the stimulus is a protein. However, this device does not merely have a different kind of activation signal. A second part of the device contains a series of nucleotide pairs that must be disrupted when the device is distorted by IHF. Consequently, this is a measuring device that enables us to estimate the amount of additional work that the protein can do when it binds to DNA.Our IHF-activated device is illustrated schematically in Figure 1. This diagram does not illustrate the part of the device involved in measuring the work, but, for simplicity, shows just the way in which the binding of IHF to its target site can distort the relationship between two DNA triple crossover (TX) motifs that flank the binding site. The upper panel shows the DNA molecule in the absence of IHF: two previously characterized TX molecules [5] are connected by a DNA double-helical shaft; each TX molecule creates a stiff domain that rigidly transmits motions of the shaft on one side to the dyes on the other side. The shaft contains the lH1
The synthesis of multithiolated DNA molecules that can be used to produce self-assembled monolayers of single-stranded DNA oligonucleotides on gold substrates is described. Generation 3 polyamidoamine (PAMAM) dendrimers were conjugated to DNA oligomers and functionalized with ~30 protected thiol groups. The protected thiol groups-thioacetate groups-allowed the dendrimer-DNA constructs to be stored in a buffer solution for at least 2 months before deprotection without any observable decrease in their ability to assemble into functional layers. The monolayers formed using these multithiolated DNA probe strands demonstrate target capture efficiencies comparable to those of analogous monolayers assembled with DNA functionalized with single thiol groups. A functional advantage of using dendrimer headgroups is the resistance to probe strand loss in prolonged exposure to buffer solutions at a high temperature (95 °C).
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