A Robust Method for the Purification and Characterization of Recombinant Human Histone H1 Variants
| Autor: | Yael David, Devin M Ray, Ivo C. Lorenz, Jakob M Hebert, Adewola Osunsade, Yazen Jmeian, Nicholas A Prescott |
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| Rok vydání: | 2018 |
| Předmět: |
SUMO-1 Protein
Computational biology Biology Protein Engineering Biochemistry Article Germline law.invention Histones 03 medical and health sciences Histone H1 Peptide Library law Transcription (biology) Escherichia coli Humans Micrococcal Nuclease Peptide library 0303 health sciences Nucleosome binding Circular Dichroism 030302 biochemistry & molecular biology Recombinant Proteins Nucleosomes Chromatin Recombinant DNA Electrophoresis Polyacrylamide Gel Linker |
| Zdroj: | Biochemistry. 58:171-176 |
| ISSN: | 1520-4995 0006-2960 |
| Popis: | Higher order compaction of the eukaryotic genome is key to the regulation of all DNA-templated processes, including transcription. This tightly controlled process involves the formation of mononucleosomes, the fundamental unit of chromatin, packaged into higher order architectures in an H1 linker histone-dependent process. While much work has been done to delineate the precise mechanism of this event in vitro and in vivo, major gaps still exist, primarily due to a lack of molecular tools. Specifically, there has never been a successful purification and biochemical characterization of all human H1 variants. Here we present a robust method to purify H1 and illustrate its utility in the purification of all somatic variants and one germline variant. In addition, we performed a first ever side-by-side biochemical comparison, which revealed a gradient of nucleosome binding affinities and compaction capabilities. These data provide new insight into H1 redundancy and lay the groundwork for the mechanistic investigation of disease-driving mutations. |
| Databáze: | OpenAIRE |
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