Controlled Assembly of Artificial Protein–Protein Complexes via DNA Duplex Formation

Płoskoń, Eliza; Wagner, Sara C.; Ellington, Andrew D.; Jewett, Michael C.; O’Reilly, Rachel K.; Booth, Paula J.

doi:10.1021/bc500473s

Cited by 2 publications

(3 citation statements)

References 31 publications

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“…Such powerful approaches typically require a sophisticated experimental sample design, which ensures functionality and accurate assembly of the biomolecules of interest. An assembly strategy which has become very popular for single molecule research is the use of synthetic protein–DNA conjugates, combining the advantages of both worlds. , Proteins provide a rich pool of functionalities, such as catalysis of chemical reactions, generation of forces, and receptor–ligand interactions, while DNA can be used to position a molecule of interest with nanometer precision. − Consequently, protein–DNA conjugates have already found widespread applications across many different single-molecule techniques, particularly in single-molecule interaction sequencing, in DNA-based nanotechnology with advanced functionalities, − and in ultrasensitive detection methods , and imaging . Furthermore, protein–DNA conjugates are essential for studies using single-molecule FRET, magnetic tweezers, or optical tweezers. , …”

Section: Introductionmentioning

confidence: 99%

Efficient Formation of Site-Specific Protein–DNA Hybrids Using Copper-Free Click Chemistry

Mukhortava

Schlierf

2016

Bioconjugate Chem.

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Protein-DNA hybrids have become increasingly popular molecular building blocks in bionanotechnology and single-molecule studies to synergistically combine the programmability of DNA with the chemical diversity of proteins. The growing demand for protein-DNA hybrids requires powerful strategies for their conjugation. Here, we present an efficient two-step method for protein-DNA assembly based on copper-free click chemistry. The method allows site-specificity and high coupling efficiency, while maintaining the conservation of protein activity. We compare our method to a commonly used protocol of direct linkage of maleimide-modified oligos. We demonstrate the significantly higher yield with a protein-DNA conjugate, which is analyzed using single-molecule force spectroscopy.

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Section: Introductionmentioning

confidence: 99%

Efficient Formation of Site-Specific Protein–DNA Hybrids Using Copper-Free Click Chemistry

Mukhortava

Schlierf

2016

Bioconjugate Chem.

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“…This subtle difference in the diameter between the complex containing DNase I and the free enzyme mixture was a result of the residual PEG moieties conjugating the enzyme. Additionally, the measurement of the diameter of the complex between EYFP and ECFP via DNAs using DLS was reported . The diameter of the complex of the two proteins located at the end of the dsDNA was approximately 7 nm.…”

Section: Results and Discussionmentioning

confidence: 99%

“…Additionally, the measurement of the diameter of the complex between EYFP and ECFP via DNAs using DLS was reported. 32 The diameter of the complex of the two proteins located at the end of the dsDNA was approximately 7 nm. In contrast, the diameter of the free monomeric protein was approximately 5 nm.…”

Section: ■ Introductionmentioning

confidence: 99%

Control of Reversible Formation and Dispersion of the Three Enzyme Networks Integrating DNA Computing

2023

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The majority of biological reactions in the cytoplasm of living cells occur via enzymatic cascade reactions. To achieve efficient enzyme cascade reactions mimicking the proximity conditions of enzymes in the cytoplasm, the proximity of each enzyme, creating a high local concentration of proteins, has been recently investigated using the conjugation of synthetic polymer molecules, proteins, and nucleic acids. Although there have been methodologies reported for the complex formation and enhanced activity of cascade reactions due to the proximity of each enzyme using DNA nanotechnology, one pair of the enzyme (GOx and HRP) complex is only assembled by the mutual independence of various shapes of the DNA structure. This study reports the network formation of three enzyme complexes assembled by a triple-branched DNA structure as a unit, thus enabling the reversible formation and dispersion of the three enzyme complex networks using single-stranded DNA, RNA, and enzymes. It was found that the activities of the three enzyme cascade reactions in the enzyme–DNA complex network were controlled by formation and dispersion of the three enzyme complex networks, due to the proximity of each enzyme with the enzyme–DNA complex network. Furthermore, three micro RNA sequences for breast cancer biomarkers were successfully detected using an enzyme–DNA complex network integrated with DNA computing. Overall, the reversible formation and dispersion of the enzyme–DNA complex network through the external stimulation of biomolecules and DNA computing provide a novel platform for controlling the production amount, diagnosis, theranostics, and biological or environmental sensing.

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Controlled Assembly of Artificial Protein–Protein Complexes via DNA Duplex Formation

Cited by 2 publications

References 31 publications

Efficient Formation of Site-Specific Protein–DNA Hybrids Using Copper-Free Click Chemistry

Efficient Formation of Site-Specific Protein–DNA Hybrids Using Copper-Free Click Chemistry

Control of Reversible Formation and Dispersion of the Three Enzyme Networks Integrating DNA Computing

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