Abstract
Tyrosine kinases are important for many cellular processes and disruption of their regulation is a factor in diseases like cancer, therefore they are a major target of anticancer drugs. There are many ways to measure tyrosine kinase activity in cells by monitoring endogenous substrate phosphorylation, or by using peptide substrates and incubating them with cell lysates containing active kinases. However, most of these strategies rely on antibodies and/or are limited in how accurately they model the intracellular environment. In cases in which activity needs to be measured in cells, but endogenous substrates are not known and/or suitable phosphospecific antibodies are not available, cell-deliverable peptide substrates can be an alternative and can provide information on activation and inhibition of kinases in intact, live cells. In this chapter, we review this methodology and provide a protocol for measuring Abl kinase activity in human cells using enzyme-linked immunosorbent assay (ELISA) with a generic antiphosphotyrosine antibody for detection.
Original language | English (US) |
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Title of host publication | Post-translational Modifications That Modulate Enzyme Activity |
Editors | Benjamin A. Garcia |
Publisher | Academic Press Inc. |
Pages | 375-406 |
Number of pages | 32 |
Volume | 626 |
ISBN (Print) | 9780128186695 |
DOIs | |
State | Published - 2019 |
Publication series
Name | Methods in enzymology |
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Publisher | Academic Press Inc. |
ISSN (Print) | 0076-6879 |
Bibliographical note
Publisher Copyright:© 2019 Elsevier Inc.
Keywords
- Abl kinase
- Cell penetrating peptide
- K562 cells
- Kinase substrate
- Tyrosine kinase assay
PubMed: MeSH publication types
- Journal Article
- Research Support, N.I.H., Extramural