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Guanine quadruplex monoclonal antibody 1H6 cross-reacts with restrained thymidine-rich single stranded DNA

Previously we reported the production and characterization of monoclonal antibody 1H6 raised against (T(4)G(4))(2) intermolecular guanine quadruplex (G4) DNA structures (Henderson A. et al. (2014) Nucleic Acids Res., 42, 860–869; Hoffmann R.F. et al. (2016) Nucleic Acids Res., 44, 152–163). It was s...

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Autores principales: Kazemier, Hinke G., Paeschke, Katrin, Lansdorp, Peter M.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5449594/
https://www.ncbi.nlm.nih.gov/pubmed/28449085
http://dx.doi.org/10.1093/nar/gkx245
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author Kazemier, Hinke G.
Paeschke, Katrin
Lansdorp, Peter M.
author_facet Kazemier, Hinke G.
Paeschke, Katrin
Lansdorp, Peter M.
author_sort Kazemier, Hinke G.
collection PubMed
description Previously we reported the production and characterization of monoclonal antibody 1H6 raised against (T(4)G(4))(2) intermolecular guanine quadruplex (G4) DNA structures (Henderson A. et al. (2014) Nucleic Acids Res., 42, 860–869; Hoffmann R.F. et al. (2016) Nucleic Acids Res., 44, 152–163). It was shown that 1H6 strongly stains nuclei and has exquisite specificity for heterochromatin by immuno-electron microscopy. Here we extend our studies of 1H6 reactivity using enzyme-linked immunosorbent assay (ELISA) and microscale thermophoresis (MST). As previously reported, 1H6 was found to strongly bind intermolecular G4 structures with a (T(4)G(4))(2) sequence motif. However, using both methods we did not detect significant binding to G4 structures without thymidines in their sequence motif or to G4 structures made with (T(2)G(4))(2) oligonucleotides. In addition, we observed strong, sequence-specific binding of 1H6 by ELISA to immobilized single stranded poly(T) DNA but not to immobilized poly(C) or poly(A) homo-polymers. Cross-reactivity of 1H6 to poly(T) was not measured in solution using MST. 1H6 was furthermore found to bind to selected areas on DNA fibers but only after DNA denaturation. Based on these observations we propose that 1H6 binds with high affinity to adjacent T's that are restricted in their movement in selected G4 structures and denatured DNA. Cross-reactivity of 1H6 to immobilized single stranded T-rich DNA next to its previously reported specificity for bona fide G4 structures needs to be taken into account in the interpretation of 1H6 binding to (sub-) cellular structures.
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spelling pubmed-54495942017-06-05 Guanine quadruplex monoclonal antibody 1H6 cross-reacts with restrained thymidine-rich single stranded DNA Kazemier, Hinke G. Paeschke, Katrin Lansdorp, Peter M. Nucleic Acids Res Molecular Biology Previously we reported the production and characterization of monoclonal antibody 1H6 raised against (T(4)G(4))(2) intermolecular guanine quadruplex (G4) DNA structures (Henderson A. et al. (2014) Nucleic Acids Res., 42, 860–869; Hoffmann R.F. et al. (2016) Nucleic Acids Res., 44, 152–163). It was shown that 1H6 strongly stains nuclei and has exquisite specificity for heterochromatin by immuno-electron microscopy. Here we extend our studies of 1H6 reactivity using enzyme-linked immunosorbent assay (ELISA) and microscale thermophoresis (MST). As previously reported, 1H6 was found to strongly bind intermolecular G4 structures with a (T(4)G(4))(2) sequence motif. However, using both methods we did not detect significant binding to G4 structures without thymidines in their sequence motif or to G4 structures made with (T(2)G(4))(2) oligonucleotides. In addition, we observed strong, sequence-specific binding of 1H6 by ELISA to immobilized single stranded poly(T) DNA but not to immobilized poly(C) or poly(A) homo-polymers. Cross-reactivity of 1H6 to poly(T) was not measured in solution using MST. 1H6 was furthermore found to bind to selected areas on DNA fibers but only after DNA denaturation. Based on these observations we propose that 1H6 binds with high affinity to adjacent T's that are restricted in their movement in selected G4 structures and denatured DNA. Cross-reactivity of 1H6 to immobilized single stranded T-rich DNA next to its previously reported specificity for bona fide G4 structures needs to be taken into account in the interpretation of 1H6 binding to (sub-) cellular structures. Oxford University Press 2017-06-02 2017-04-25 /pmc/articles/PMC5449594/ /pubmed/28449085 http://dx.doi.org/10.1093/nar/gkx245 Text en © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research. http://creativecommons.org/licenses/by-nc/4.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com
spellingShingle Molecular Biology
Kazemier, Hinke G.
Paeschke, Katrin
Lansdorp, Peter M.
Guanine quadruplex monoclonal antibody 1H6 cross-reacts with restrained thymidine-rich single stranded DNA
title Guanine quadruplex monoclonal antibody 1H6 cross-reacts with restrained thymidine-rich single stranded DNA
title_full Guanine quadruplex monoclonal antibody 1H6 cross-reacts with restrained thymidine-rich single stranded DNA
title_fullStr Guanine quadruplex monoclonal antibody 1H6 cross-reacts with restrained thymidine-rich single stranded DNA
title_full_unstemmed Guanine quadruplex monoclonal antibody 1H6 cross-reacts with restrained thymidine-rich single stranded DNA
title_short Guanine quadruplex monoclonal antibody 1H6 cross-reacts with restrained thymidine-rich single stranded DNA
title_sort guanine quadruplex monoclonal antibody 1h6 cross-reacts with restrained thymidine-rich single stranded dna
topic Molecular Biology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5449594/
https://www.ncbi.nlm.nih.gov/pubmed/28449085
http://dx.doi.org/10.1093/nar/gkx245
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