Alan M Moses
  • Department of Computer Science, University of Toronto, Canada, Canada,
Research fields
  • Cell biology
A Direct in vitro Fatty Acylation Assay for Hedgehog Acyltransferase
Authors:  Adina R. Schonbrun and Marilyn D. Resh, date: 12/20/2022, view: 424, Q&A: 0

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.


Graphical abstract



Graphical abstract adapted from Schonbrun and Resh (2022)

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