Chapter title |
Small-Molecule Screening Assay for Mono-ADP-Ribosyltransferases
|
---|---|
Chapter number | 16 |
Book title |
ADP-ribosylation and NAD+ Utilizing Enzymes
|
Published in |
Methods in molecular biology, August 2018
|
DOI | 10.1007/978-1-4939-8588-3_16 |
Pubmed ID | |
Book ISBNs |
978-1-4939-8587-6, 978-1-4939-8588-3
|
Authors |
Teemu Haikarainen, Sudarshan Murthy, Mirko M. Maksimainen, Lari Lehtiö, Haikarainen, Teemu, Murthy, Sudarshan, Maksimainen, Mirko M., Lehtiö, Lari |
Abstract |
Mono-ADP-ribosyltransferases of the PARP/ARTD enzyme family are enzymes catalyzing the transfer of a single ADP-ribose unit to target proteins. The enzymes have various roles in vital cellular processes such as DNA repair and transcription, and many of the enzymes are linked to cancer-relevant functions. Thus inhibition of the enzymes is a potential way to discover and develop new drugs against cancer. Here we describe an activity-based screening assay for mono-ADP-ribosyltransferases. The assay utilizes the natural substrate of the enzymes, NAD+, and it is based on chemically converting the leftover substrate to a fluorophore and measuring its relative concentration after the enzymatic reaction. The assay is homogenous, robust, and cost-effective and, most importantly, applicable to mono-ADP-ribosyltransferases as well as poly-ADP-ribosyltransferases for screening of small-molecule inhibitors against the enzymes. |
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