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A Polypeptide-DNA Hybrid with Selective Linking Capability Applied to Single Molecule Nano-Mechanical Measurements Using Optical Tweezers

Many applications in biosensing, biomaterial engineering and single molecule biophysics require multiple non-covalent linkages between DNA, protein molecules, and surfaces that are specific yet strong. Here, we present a novel method to join proteins and dsDNA molecule at their ends, in an efficient...

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Detalles Bibliográficos
Autores principales: Moayed, Fatemeh, Mashaghi, Alireza, Tans, Sander J.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2013
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3545873/
https://www.ncbi.nlm.nih.gov/pubmed/23336001
http://dx.doi.org/10.1371/journal.pone.0054440
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author Moayed, Fatemeh
Mashaghi, Alireza
Tans, Sander J.
author_facet Moayed, Fatemeh
Mashaghi, Alireza
Tans, Sander J.
author_sort Moayed, Fatemeh
collection PubMed
description Many applications in biosensing, biomaterial engineering and single molecule biophysics require multiple non-covalent linkages between DNA, protein molecules, and surfaces that are specific yet strong. Here, we present a novel method to join proteins and dsDNA molecule at their ends, in an efficient, rapid and specific manner, based on the recently developed linkage between the protein StrepTactin (STN) and the peptide StrepTag II (ST). We introduce a two-step approach, in which we first construct a hybrid between DNA and a tandem of two STs peptides (tST). In a second step, this hybrid is linked to polystyrene bead surfaces and Maltose Binding Protein (MBP) using STN. Furthermore, we show the STN-tST linkage is more stable against forces applied by optical tweezers than the commonly used biotin-Streptavidin (STV) linkage. It can be used in conjunction with Neutravidin (NTV)-biotin linkages to form DNA tethers that can sustain applied forces above 65 pN for tens of minutes in a quarter of the cases. The method is general and can be applied to construct other surface-DNA and protein-DNA hybrids. The reversibility, high mechanical stability and specificity provided by this linking procedure make it highly suitable for single molecule mechanical studies, as well as biosensing and lab on chip applications.
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spelling pubmed-35458732013-01-18 A Polypeptide-DNA Hybrid with Selective Linking Capability Applied to Single Molecule Nano-Mechanical Measurements Using Optical Tweezers Moayed, Fatemeh Mashaghi, Alireza Tans, Sander J. PLoS One Research Article Many applications in biosensing, biomaterial engineering and single molecule biophysics require multiple non-covalent linkages between DNA, protein molecules, and surfaces that are specific yet strong. Here, we present a novel method to join proteins and dsDNA molecule at their ends, in an efficient, rapid and specific manner, based on the recently developed linkage between the protein StrepTactin (STN) and the peptide StrepTag II (ST). We introduce a two-step approach, in which we first construct a hybrid between DNA and a tandem of two STs peptides (tST). In a second step, this hybrid is linked to polystyrene bead surfaces and Maltose Binding Protein (MBP) using STN. Furthermore, we show the STN-tST linkage is more stable against forces applied by optical tweezers than the commonly used biotin-Streptavidin (STV) linkage. It can be used in conjunction with Neutravidin (NTV)-biotin linkages to form DNA tethers that can sustain applied forces above 65 pN for tens of minutes in a quarter of the cases. The method is general and can be applied to construct other surface-DNA and protein-DNA hybrids. The reversibility, high mechanical stability and specificity provided by this linking procedure make it highly suitable for single molecule mechanical studies, as well as biosensing and lab on chip applications. Public Library of Science 2013-01-15 /pmc/articles/PMC3545873/ /pubmed/23336001 http://dx.doi.org/10.1371/journal.pone.0054440 Text en © 2013 Moayed et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Moayed, Fatemeh
Mashaghi, Alireza
Tans, Sander J.
A Polypeptide-DNA Hybrid with Selective Linking Capability Applied to Single Molecule Nano-Mechanical Measurements Using Optical Tweezers
title A Polypeptide-DNA Hybrid with Selective Linking Capability Applied to Single Molecule Nano-Mechanical Measurements Using Optical Tweezers
title_full A Polypeptide-DNA Hybrid with Selective Linking Capability Applied to Single Molecule Nano-Mechanical Measurements Using Optical Tweezers
title_fullStr A Polypeptide-DNA Hybrid with Selective Linking Capability Applied to Single Molecule Nano-Mechanical Measurements Using Optical Tweezers
title_full_unstemmed A Polypeptide-DNA Hybrid with Selective Linking Capability Applied to Single Molecule Nano-Mechanical Measurements Using Optical Tweezers
title_short A Polypeptide-DNA Hybrid with Selective Linking Capability Applied to Single Molecule Nano-Mechanical Measurements Using Optical Tweezers
title_sort polypeptide-dna hybrid with selective linking capability applied to single molecule nano-mechanical measurements using optical tweezers
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3545873/
https://www.ncbi.nlm.nih.gov/pubmed/23336001
http://dx.doi.org/10.1371/journal.pone.0054440
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