| Chapter title |
Purification of 26S Proteasomes and Their Subcomplexes from Plants.
|
|---|---|
| Chapter number | 24 |
| Book title |
Isolation of Plant Organelles and Structures
|
| Published in |
Methods in molecular biology, January 2017
|
| DOI | 10.1007/978-1-4939-6533-5_24 |
| Pubmed ID | |
| Book ISBNs |
978-1-4939-6531-1, 978-1-4939-6533-5
|
| Authors |
Richard S. Marshall, David C. Gemperline, Richard D. Vierstra |
| Editors |
Nicolas L. Taylor, A. Harvey Millar |
| Abstract |
The 26S proteasome is a highly dynamic, multisubunit, ATP-dependent protease that plays a central role in cellular housekeeping and many aspects of plant growth and development by degrading aberrant polypeptides and key cellular regulators that are first modified by ubiquitin. Although the 26S proteasome was originally enriched from plants over 30 years ago, only recently have significant advances been made in our ability to isolate and study the plant particle. Here, we describe two robust methods for purifying the 26S proteasome and its subcomplexes from Arabidopsis thaliana; one that involves conventional chromatography techniques to isolate the complex from wild-type plants, and another that employs the genetic replacement of individual subunits with epitope-tagged variants combined with affinity purification. In addition to these purification protocols, we describe methods commonly used to analyze the activity and composition of the complex. |
Mendeley readers
Geographical breakdown
| Country | Count | As % |
|---|---|---|
| Unknown | 7 | 100% |
Demographic breakdown
| Readers by professional status | Count | As % |
|---|---|---|
| Researcher | 4 | 57% |
| Student > Master | 2 | 29% |
| Student > Ph. D. Student | 1 | 14% |
| Readers by discipline | Count | As % |
|---|---|---|
| Biochemistry, Genetics and Molecular Biology | 3 | 43% |
| Agricultural and Biological Sciences | 3 | 43% |
| Unknown | 1 | 14% |