Light‐Induced Triggering of Peroxidase Activity Using Quantum Dots

Fruk, Ljiljana; Rajendran, Vidyalakshmi; Spengler, Mark; Niemeyer, Christof M.

doi:10.1002/cbic.200700594

Cited by 61 publications

(61 citation statements)

References 25 publications

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“…To gain access to fusion proteins bearing the complementary Snap-and Halo-protein tags, we constructed expression plasmids by genetic fusion of the genes encoding the protein of interest (POI) and Snap-tag or HaloTag (see the Supporting Information). As model POIs we chose the fluorescent proteins enhanced yellow fluorescent protein (EYFP) [22] and mKate, [23] the enzymes cytochrome C peroxidase (CCP) [24] and esterase 2 from Alicyclobacillus acidocaldarius thermos (EST2), [25] to which the self-labeling tags were fused at the C terminus (POI-Snap or POI-Halo, respectively). In addition, the bispecific Halo-Snap fusion protein "covalin", [26] a chimera which specifically reacts with both BG and CH, as well as monovalent STV (mSTV), [27] were used in this study.…”

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confidence: 99%

Orthogonal Protein Decoration of DNA Origami

Saccá¹,

Meyer²,

Erkelenz³

et al. 2010

Angew Chem Int Ed

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Structural DNA nanotechnology [1,2] and the technique of DNA origami [3] enable the rapid generation of a plethora of complex self-assembled nanostructures. [4][5][6] Since DNA molecules themselves display limited chemical, optical, and electronic functionality, it is of utmost importance to devise methods to decorate DNA scaffolds with functional moieties to realize applications in sensing, catalysis, and device fabrication. Protein functionalization is particulary desirable because it allows exploitation of an almost unlimited variety of functional elements which nature has evolved over billions of years.[7] The delicate architecture of proteins has resulted in no generally applicable method being currently available to selectively couple these components on DNA scaffolds, and thus approaches used so far are based on reversible antibodyantigen interactions, [8,9] aptamer binding, [10,11] nucleic acid hybridization of DNA-tagged proteins, [12,13] or predominantly biotin-streptavidin (STV) interactions. [14][15][16][17][18][19] We demonstrate here that DNA nanostructures can be site-specifically decorated with several different proteins by using coupling systems orthogonal to the biotin-STV system. In particular, benzylguanine (BG) and chlorohexane (CH) groups incorporated in DNA origami have been used as suicide ligands for the site-specific coupling of fusion proteins containing the self-labeling protein tags O 6 -alkylguanine-DNA-alkyltransferase (hAGT), which is often referred to as "Snap-tag", [20] or haloalkane dehalogenase, which is also known as "HaloTag".[21] By using various model proteins we demonstrate the general applicability of this approach for the generation of DNA superstructures that are selectively decorated with multiple different proteins.To realize orthogonal protein immobilization on DNA origami using self-ligating protein tags, we chose the Snap-tag, developed by Johnsson and co-workers, [20] and the commercially available HaloTag [21] system. The respective smallmolecule suicide tags (O 6 -benzylguanine (BG) and 5-chlorohexane (CH)) for both self-labeling protein tags are readily available as amino-reactive N-hydroxysuccinimide (NHS) derivatives (BG-NHS and CH-NHS; Figure 1 a). Complete derivatization of alkylamino-modified oligonucleotides was achieved by coupling with 30 molar equivalents of BG-NHS or CH-NHS, as indicated by electrophoretic analysis (Figure 1 b). To gain access to fusion proteins bearing the complementary Snap-and Halo-protein tags, we constructed expression plasmids by genetic fusion of the genes encoding the protein of interest (POI) and Snap-tag or HaloTag (see the Supporting Information). As model POIs we chose the fluorescent proteins enhanced yellow fluorescent protein (EYFP) [22] and mKate, [23] the enzymes cytochrome C peroxidase (CCP) [24] and esterase 2 from Alicyclobacillus acidocaldarius thermos (EST2), [25] to which the self-labeling tags were fused at the C terminus (POI-Snap or POI-Halo, respectively). In addition, the bispecific Halo-Snap fusion protein...

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mentioning

confidence: 99%

Orthogonal Protein Decoration of DNA Origami

Saccá¹,

Meyer²,

Erkelenz³

et al. 2010

Angew Chem Int Ed

Self Cite

266

183

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“…Consequently, work was started on the synthesis and in-depth evaluation of a novel class of redox-enzymes, recently described by Fruk et al (25,27,28), who described the synthesis of such conjugates by means of the reconstitution of apoenzymes with covalent DNA-hemin adducts. It was also evident from other projects in our group, that the class of cytochrome P450 enzymes are tunable by synthetic cofactors, such as semiconductor nanoparticles (29)(30)(31). Given the fact that P450 enzymes have extensively been used for the biofunctionalization of electronic devices in the area of biosensing (26), we reasoned that future nanobioelectromc systems might benefit from the development of semisynthetic DNA-P450 conjugates.…”

Section: Development Of Novel Semisynthetic Redox Enzymesmentioning

confidence: 78%

Semisynthetic DNA-protein conjugates for fabrication of nucleic acid based nanostructures

Rabe¹,

Feldkamp²,

Niemeyer³

2008

AIP Conference Proceedings

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We here report on the developments of semisynthetic DNA-protein conjugates and their assembly into multi-component nanostructures. We describe the improvement of the DNA sequences embedded in such nanostructures by computational and analytical methods. Moreover, we report on the exploration of novel DNA conjugates of streptavidin or redox proteins with improved properties for the assembly of nucleic acid based nanostructures.

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“…Although we used a limited set of only three different capture oligomers in this model study, the complexity of the DNA array can be further increased using a larger number of designed orthogonal oligonucleotide pairs 42. We therefore anticipate that the self‐assembly and regeneration capabilities implemented in this device will open the door to a number of applications such as the investigation of coupled redox enzymes and their effectors,40, 43–46 or other applications in enzymology, biomedical diagnostics, and nanobiotechnology.…”

Section: Discussionmentioning

confidence: 99%

Addressable Microfluidic Polymer Chip for DNA‐Directed Immobilization of Oligonucleotide‐Tagged Compounds

Schröder

Hoffmann

Müller

et al. 2009

Small

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A microfluidic polymer chip for the self-assembly of DNA conjugates through DNA-directed immobilization is developed. The chip is fabricated from two parts, one of which contains a microfluidic channel produced from poly(dimethylsiloxane) (PDMS) by replica-casting technique using a mold prepared by photolithographic techniques. The microfluidic part is sealed by covalent bonding with a chemically activated glass slide containing a DNA oligonucleotide microarray. The dimension of the PDMS-glass microfluidic chip is equivalent to standard microscope slides (76 x 26 mm(2)). The DNA microarray surface inside the microfluidic channels is configured through conventional spotting, and the resulting DNA patches can be conveniently addressed with compounds containing complementary DNA tags. To demonstrate the utility of the addressable surface within the microfluidic channel, DNA-directed immobilization (DDI) of DNA-modified gold nanoparticles (AuNPs) and DNA-conjugates of the enzymes glucose oxidase (GOx) and horseradish peroxidase (HRP) are carried out. DDI of AuNPs is used to demonstrate site selectivity and reversibility of the surface-modification process. In the case of the DNA-enzyme conjugates, the patterned assembly of the two enzymes allows the establishment and investigation of the coupled reaction of GOx and HRP, with particular emphasis on surface coverage and lateral flow rates. The results demonstrate that this addressable chip is well suited for the generation of fluidically coupled multi-enzyme microreactors.

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Light‐Induced Triggering of Peroxidase Activity Using Quantum Dots

Cited by 61 publications

References 25 publications

Orthogonal Protein Decoration of DNA Origami

Orthogonal Protein Decoration of DNA Origami

Semisynthetic DNA-protein conjugates for fabrication of nucleic acid based nanostructures

Addressable Microfluidic Polymer Chip for DNA‐Directed Immobilization of Oligonucleotide‐Tagged Compounds

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