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Abstract
AMP-activated protein kinase (AMPK) is extremely sensitive to cellular stress, so that nonphysiological activation of the kinase can readily occur during harvesting of cells or tissues. In this chapter we describe methods to harvest cells and tissues, and for kinase assays, that preserve the physiological activation status of AMPK as far as possible. Note that similar care with methods of cell or tissue harvesting is required when AMPK function is monitored by Western blotting, rather than by kinase assays. We also describe methods to determine whether compounds that activate AMPK in intact cells do so indirectly by interfering with cellular ATP synthesis or directly by binding to AMPK and, if the latter, whether this occurs by binding at the AMP-binding sites on the γ subunit or at the ADaM site located between the α and β subunits.
Original language | English |
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Title of host publication | AMPK |
Subtitle of host publication | Methods and Protocols |
Editors | Dietbert Neumann, Benoit Viollet |
Place of Publication | New York |
Publisher | Humana Press |
Pages | 239-253 |
Number of pages | 15 |
ISBN (Electronic) | 9781493975983 |
ISBN (Print) | 9781493975976 |
DOIs | |
Publication status | Published - 2018 |
Publication series
Name | Methods in Molecular Biology |
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Publisher | Humana Press |
Volume | 1732 |
ISSN (Print) | 1064-3745 |
ISSN (Electronic) | 1940-6029 |
Keywords
- Allosteric activation
- AMP-activated protein kinase
- AMPK
- Dephosphorylation
- Kinase assay
- Phosphorylation
ASJC Scopus subject areas
- Molecular Biology
- Genetics
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Role of AMPK in Nutrient Sensing and in Cancer (Investigator Award Renewal)
1/10/17 → 31/03/24
Project: Research