In the 1980s, it was found that addition of N-terminal Arg to proteins induces their ubiquitination and degradation by the N-end rule pathway. While this mechanism applies only to the proteins which also have other features of the N-degron (including a closely adjacent Lys that is accessible for ubiquitination), several test substrates have been found to follow this mechanism very efficiently after ATE1-dependent arginylation. Such property enabled researchers to test ATE1 activity in cells indirectly by assaying for the degradation of such arginylation-dependent substrates. The most commonly used substrate for this assay is E. coli beta-galactosidase (beta-Gal) because its level can be easily measured using standardized colorimetric assays. Here, we describe this method, which has served as a quick and easy way to characterize ATE1 activity during identification of arginyltransferases in different species.
CITATION STYLE
Kashina, A. S. (2023). Assaying ATE1 Activity in Yeast by β-Gal Degradation. In Methods in Molecular Biology (Vol. 2620, pp. 63–70). Humana Press Inc. https://doi.org/10.1007/978-1-0716-2942-0_8
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