Direct and Efficient Conjugation of Quantum Dots to DNA Nanostructures with Peptide-PNA

Abstract

DNA nanotechnology has proven to be a powerful strategy for the bottom-up preparation of colloidal nanoparticle (NP) superstructures, enabling the coordination of multiple NPs with orientation and separation approaching nanometer precision. To do this, NPs are often conjugated with chemically modified, single-stranded (ss) DNA that can recognize complementary ssDNA on the DNA nanostructure. The limitation is that many NPs cannot be easily conjugated with ssDNA, and other conjugation strategies are expensive, inefficient, or reduce the specificity and/or precision with which NPs can be placed. As an alternative, the conjugation of nanoparticle-binding peptides and peptide nucleic acids (PNA) can produce peptide-PNA with distinct NP-binding and DNA-binding domains. Here, we demonstrate a simple application of this method to conjugate semiconductor quantum dots (QDs) directly to DNA nanostructures by means of a peptide-PNA with a six-histidine peptide motif that binds to the QD surface. With this method, we achieved greater than 90% capture efficiency for multiple QDs on a single DNA nanostructure while preserving both site specificity and precise spatial control of QD placement. Additionally, we investigated the effects of peptide-PNA charge on the efficacy of QD immobilization in suboptimal conditions. The results validate peptide-PNA as a viable alternative to ssDNA conjugation of NPs and warrant studies of other NP-binding peptides for peptide-PNA conjugation.

Benzaldehyde Lyase Kinetic Improvements, Potential Channeling to Alcohol Dehydrogenase, and Substrate Scope when Immobilized on Semiconductor Quantum Dots

“…Understanding how QD–enzyme nanoaggregates form such that the size can be predicted and controlled is clearly important in this context and is the subject for future studies incorporating both experiments and simulations. Protein or nucleic acid materials that can link QDs and other NPs into heteroaggregate structures with predetermined interparticle spacing may be very useful in this regard. , Indeed, it may be possible to scale up to larger size reactions where the NP–enzyme clusters are immobilized on packed columns or in other compartments with porous small diameter frits and the substrates/product-containing media are continuously flowed through and even replenished if needed. , This latter approach would prevent the washing out of conjugates. Although only preliminary in nature, these results indicate the continued potential for a broad range of reaction improvements and benefits using a NP display of enzymes.…”

“…Protein or nucleic acid materials that can link QDs and other NPs into heteroaggregate structures with predetermined interparticle spacing may be very useful in this regard. 63,64 Indeed, it may be possible to scale up to larger size reactions where the NP−enzyme clusters are immobilized on packed columns or in other compartments with porous small diameter frits and the substrates/product-containing media are continuously flowed through and even replenished if needed. 9,65 This latter approach would prevent the washing out of conjugates.…”

Understanding Förster Resonance Energy Transfer in the Sheet Regime with DNA Brick-Based Dye Networks

“…AFM imaging was performed using a Multimode AFM (Bruker) in tapping in air mode. A 10 μL amount of purified DNA block sample (at ∼5 nM) was deposited on a freshly cleaved mica disc and allowed to adsorb for 30–60 s. After that, the mica was rinsed with molecular biology grade water and dried using N 2 gas. , High-resolution transmission electron microscopy (HRTEM) was performed on a JEOL TEM 2010F electron microscope operating at 200 kV. For TEM imaging, the samples were adsorbed on glow-discharged carbon-coated TEM grids (400 mesh, Ted Pella) .…”

Nanoscale 3D spatial addressing and valence control of quantum dots using wireframe DNA origami

“…In another example, Liedl et al fabricated “planet–satellite” nanoclusters with controlled sizes up to 500 nm using DNA-origami-guided self-assembly 37 . And Diaz et al recently demonstrated the efficient conjugation of QDs to DNA nanostructures using QDs with a peptide nucleic acid (PNA) containing a hexahistidine peptide motif 40 . However, the preceding strategies still suffer from multiple ssDNAs and PNAs that are typically bound non-specifically to the QD surface, which increases the probability that a QD binds to more than one DNA nanostructure and consequently reduces the purity of target assemblies.…”