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Base Dynamics in the HhaI Protein Binding Site

[Image: see text] Protein–DNA interactions play an important role in numerous biological functions within the living cell. In many of these interactions, the DNA helix is significantly distorted upon protein–DNA complex formation. The HhaI restriction-modification system is one such system, where th...

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Autores principales: Pederson, Kari, Meints, Gary A., Drobny, Gary P.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: American Chemical Society 2023
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10461302/
https://www.ncbi.nlm.nih.gov/pubmed/37561575
http://dx.doi.org/10.1021/acs.jpcb.3c03687
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author Pederson, Kari
Meints, Gary A.
Drobny, Gary P.
author_facet Pederson, Kari
Meints, Gary A.
Drobny, Gary P.
author_sort Pederson, Kari
collection PubMed
description [Image: see text] Protein–DNA interactions play an important role in numerous biological functions within the living cell. In many of these interactions, the DNA helix is significantly distorted upon protein–DNA complex formation. The HhaI restriction-modification system is one such system, where the methylation target is flipped out of the helix when bound to the methyltransferase. However, the base flipping mechanism is not well understood. The dynamics of the binding site of the HhaI methyltransferase and endonuclease (underlined) within the DNA oligomer [d(G(1)A(2)T(3)A(4)G(5)C(6)G(7)C(8)T(9)A(10)T(11)C(12))](2) are studied using deuterium solid-state NMR (SSNMR). SSNMR spectra obtained from DNAs deuterated on the base of nucleotides within and flanking the [5′-GCGC-3′](2) sequence indicate that all of these positions are structurally flexible. Previously, conformational flexibility within the phosphodiester backbone and furanose ring within the target sequence has been observed and hypothesized to play a role in the distortion mechanism. However, whether that distortion was occurring through an active or passive mechanism remained unclear. These NMR data demonstrate that although the [5′-GCGC-3′](2) sequence is dynamic, the target cytosine is not passively flipping out of the double-helix on the millisecond–picosecond time scale. Additionally, although previous studies have shown that both the furanose ring and phosphodiester backbone experience a change in dynamics upon methylation, which may play a role in recognition and cleavage by the endonuclease, our observations here indicate that methylation has no effect on the dynamics of the base itself.
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spelling pubmed-104613022023-08-29 Base Dynamics in the HhaI Protein Binding Site Pederson, Kari Meints, Gary A. Drobny, Gary P. J Phys Chem B [Image: see text] Protein–DNA interactions play an important role in numerous biological functions within the living cell. In many of these interactions, the DNA helix is significantly distorted upon protein–DNA complex formation. The HhaI restriction-modification system is one such system, where the methylation target is flipped out of the helix when bound to the methyltransferase. However, the base flipping mechanism is not well understood. The dynamics of the binding site of the HhaI methyltransferase and endonuclease (underlined) within the DNA oligomer [d(G(1)A(2)T(3)A(4)G(5)C(6)G(7)C(8)T(9)A(10)T(11)C(12))](2) are studied using deuterium solid-state NMR (SSNMR). SSNMR spectra obtained from DNAs deuterated on the base of nucleotides within and flanking the [5′-GCGC-3′](2) sequence indicate that all of these positions are structurally flexible. Previously, conformational flexibility within the phosphodiester backbone and furanose ring within the target sequence has been observed and hypothesized to play a role in the distortion mechanism. However, whether that distortion was occurring through an active or passive mechanism remained unclear. These NMR data demonstrate that although the [5′-GCGC-3′](2) sequence is dynamic, the target cytosine is not passively flipping out of the double-helix on the millisecond–picosecond time scale. Additionally, although previous studies have shown that both the furanose ring and phosphodiester backbone experience a change in dynamics upon methylation, which may play a role in recognition and cleavage by the endonuclease, our observations here indicate that methylation has no effect on the dynamics of the base itself. American Chemical Society 2023-08-10 /pmc/articles/PMC10461302/ /pubmed/37561575 http://dx.doi.org/10.1021/acs.jpcb.3c03687 Text en © 2023 The Authors. Published by American Chemical Society https://creativecommons.org/licenses/by/4.0/Permits the broadest form of re-use including for commercial purposes, provided that author attribution and integrity are maintained (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Pederson, Kari
Meints, Gary A.
Drobny, Gary P.
Base Dynamics in the HhaI Protein Binding Site
title Base Dynamics in the HhaI Protein Binding Site
title_full Base Dynamics in the HhaI Protein Binding Site
title_fullStr Base Dynamics in the HhaI Protein Binding Site
title_full_unstemmed Base Dynamics in the HhaI Protein Binding Site
title_short Base Dynamics in the HhaI Protein Binding Site
title_sort base dynamics in the hhai protein binding site
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10461302/
https://www.ncbi.nlm.nih.gov/pubmed/37561575
http://dx.doi.org/10.1021/acs.jpcb.3c03687
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