ArticleBio-protocol2022
A Direct in vitro Fatty Acylation Assay for Hedgehog Acyltransferase.
Article in Bio-protocol, 2022. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Not yet cited in PubMed.
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2 authors.
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Abstract
Several assays have been developed to monitor the in vitro catalytic activity of Hedgehog acyltransferase (Hhat), an enzyme critical to the Hedgehog signaling pathway in cells. However, the majority of these previously reported assays involve radioactive fatty acyl donor substrates, multiple steps to achieve product readout, or specialized equipment. To increase safety, efficiency, and convenience, we developed a direct, fluorescent in vitro assay to monitor Hhat activity. Our assay utilizes purified Hhat, a fluorescently labeled fatty acyl-CoA donor substrate, and a Sonic hedgehog (Shh) peptide recipient substrate sufficient for fatty acylation. The protocol is a straightforward process that yields direct readout of fatty acylated Shh peptide via fluorescence detection of the transferred fatty acyl group.
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