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Analysis of the HindIII-catalyzed reaction by time-resolved crystallography
In order to investigate the mechanism of the reaction catalyzed by HindIII, structures of HindIII–DNA complexes with varying durations of soaking time in cryoprotectant buffer containing manganese ions were determined by the freeze-trap method. In the crystal structures of the complexes obtained aft...
Autores principales: | , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
International Union of Crystallography
2015
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4321485/ https://www.ncbi.nlm.nih.gov/pubmed/25664735 http://dx.doi.org/10.1107/S1399004714025188 |
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author | Kawamura, Takashi Kobayashi, Tomoki Watanabe, Nobuhisa |
author_facet | Kawamura, Takashi Kobayashi, Tomoki Watanabe, Nobuhisa |
author_sort | Kawamura, Takashi |
collection | PubMed |
description | In order to investigate the mechanism of the reaction catalyzed by HindIII, structures of HindIII–DNA complexes with varying durations of soaking time in cryoprotectant buffer containing manganese ions were determined by the freeze-trap method. In the crystal structures of the complexes obtained after soaking for a longer duration, two manganese ions, indicated by relatively higher electron density, are clearly observed at the two metal ion-binding sites in the active site of HindIII. The increase in the electron density of the two metal-ion peaks followed distinct pathways with increasing soaking times, suggesting variation in the binding rate constant for the two metal sites. DNA cleavage is observed when the second manganese ion appears, suggesting that HindIII uses the two-metal-ion mechanism, or alternatively that its reactivity is enhanced by the binding of the second metal ion. In addition, conformational change in a loop near the active site accompanies the catalytic reaction. |
format | Online Article Text |
id | pubmed-4321485 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2015 |
publisher | International Union of Crystallography |
record_format | MEDLINE/PubMed |
spelling | pubmed-43214852015-02-20 Analysis of the HindIII-catalyzed reaction by time-resolved crystallography Kawamura, Takashi Kobayashi, Tomoki Watanabe, Nobuhisa Acta Crystallogr D Biol Crystallogr Research Papers In order to investigate the mechanism of the reaction catalyzed by HindIII, structures of HindIII–DNA complexes with varying durations of soaking time in cryoprotectant buffer containing manganese ions were determined by the freeze-trap method. In the crystal structures of the complexes obtained after soaking for a longer duration, two manganese ions, indicated by relatively higher electron density, are clearly observed at the two metal ion-binding sites in the active site of HindIII. The increase in the electron density of the two metal-ion peaks followed distinct pathways with increasing soaking times, suggesting variation in the binding rate constant for the two metal sites. DNA cleavage is observed when the second manganese ion appears, suggesting that HindIII uses the two-metal-ion mechanism, or alternatively that its reactivity is enhanced by the binding of the second metal ion. In addition, conformational change in a loop near the active site accompanies the catalytic reaction. International Union of Crystallography 2015-01-23 /pmc/articles/PMC4321485/ /pubmed/25664735 http://dx.doi.org/10.1107/S1399004714025188 Text en © Kawamura et al. 2015 http://creativecommons.org/licenses/by/2.0/uk/ This is an open-access article distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original authors and source are cited. |
spellingShingle | Research Papers Kawamura, Takashi Kobayashi, Tomoki Watanabe, Nobuhisa Analysis of the HindIII-catalyzed reaction by time-resolved crystallography |
title | Analysis of the HindIII-catalyzed reaction by time-resolved crystallography |
title_full | Analysis of the HindIII-catalyzed reaction by time-resolved crystallography |
title_fullStr | Analysis of the HindIII-catalyzed reaction by time-resolved crystallography |
title_full_unstemmed | Analysis of the HindIII-catalyzed reaction by time-resolved crystallography |
title_short | Analysis of the HindIII-catalyzed reaction by time-resolved crystallography |
title_sort | analysis of the hindiii-catalyzed reaction by time-resolved crystallography |
topic | Research Papers |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4321485/ https://www.ncbi.nlm.nih.gov/pubmed/25664735 http://dx.doi.org/10.1107/S1399004714025188 |
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