Rapid purification of recombinant histones.
The development of methods to assemble nucleosomes from recombinant histones decades ago has transformed chromatin research. Nevertheless, nucleosome reconstitution remains time consuming to this day, not least because the four individual histones must be purified first. Here, we present a streamlin...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
Public Library of Science (PLoS)
2014-01-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC4121265?pdf=render |
| _version_ | 1811226871835131904 |
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| author | Henrike Klinker Caroline Haas Nadine Harrer Peter B Becker Felix Mueller-Planitz |
| author_facet | Henrike Klinker Caroline Haas Nadine Harrer Peter B Becker Felix Mueller-Planitz |
| author_sort | Henrike Klinker |
| collection | DOAJ |
| description | The development of methods to assemble nucleosomes from recombinant histones decades ago has transformed chromatin research. Nevertheless, nucleosome reconstitution remains time consuming to this day, not least because the four individual histones must be purified first. Here, we present a streamlined purification protocol of recombinant histones from bacteria. We termed this method "rapid histone purification" (RHP) as it circumvents isolation of inclusion bodies and thereby cuts out the most time-consuming step of traditional purification protocols. Instead of inclusion body isolation, whole cell extracts are prepared under strongly denaturing conditions that directly solubilize inclusion bodies. By ion exchange chromatography, the histones are purified from the extracts. The protocol has been successfully applied to all four canonical Drosophila and human histones. RHP histones and histones that were purified from isolated inclusion bodies had similar purities. The different purification strategies also did not impact the quality of octamers reconstituted from these histones. We expect that the RHP protocol can be readily applied to the purification of canonical histones from other species as well as the numerous histone variants. |
| first_indexed | 2024-04-12T09:32:07Z |
| format | Article |
| id | doaj.art-2b107d34c41844fb94519b511abc8047 |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-04-12T09:32:07Z |
| publishDate | 2014-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj.art-2b107d34c41844fb94519b511abc80472022-12-22T03:38:19ZengPublic Library of Science (PLoS)PLoS ONE1932-62032014-01-0198e10402910.1371/journal.pone.0104029Rapid purification of recombinant histones.Henrike KlinkerCaroline HaasNadine HarrerPeter B BeckerFelix Mueller-PlanitzThe development of methods to assemble nucleosomes from recombinant histones decades ago has transformed chromatin research. Nevertheless, nucleosome reconstitution remains time consuming to this day, not least because the four individual histones must be purified first. Here, we present a streamlined purification protocol of recombinant histones from bacteria. We termed this method "rapid histone purification" (RHP) as it circumvents isolation of inclusion bodies and thereby cuts out the most time-consuming step of traditional purification protocols. Instead of inclusion body isolation, whole cell extracts are prepared under strongly denaturing conditions that directly solubilize inclusion bodies. By ion exchange chromatography, the histones are purified from the extracts. The protocol has been successfully applied to all four canonical Drosophila and human histones. RHP histones and histones that were purified from isolated inclusion bodies had similar purities. The different purification strategies also did not impact the quality of octamers reconstituted from these histones. We expect that the RHP protocol can be readily applied to the purification of canonical histones from other species as well as the numerous histone variants.http://europepmc.org/articles/PMC4121265?pdf=render |
| spellingShingle | Henrike Klinker Caroline Haas Nadine Harrer Peter B Becker Felix Mueller-Planitz Rapid purification of recombinant histones. PLoS ONE |
| title | Rapid purification of recombinant histones. |
| title_full | Rapid purification of recombinant histones. |
| title_fullStr | Rapid purification of recombinant histones. |
| title_full_unstemmed | Rapid purification of recombinant histones. |
| title_short | Rapid purification of recombinant histones. |
| title_sort | rapid purification of recombinant histones |
| url | http://europepmc.org/articles/PMC4121265?pdf=render |
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