A palmitoyl transferase chemical genetic system to map ZDHHC-specific S-acylation

Abstract

The 23 human ZDHHC S-acyltransferases catalyze long-chain S-acylation at cysteine residues across an extensive network of hundreds of proteins important for normal physiology or dysregulated in disease. Here we present a technology platform to directly map the protein substrates of a specific ZDHHC for the first time at the whole proteome level, in intact cells. Structure-guided engineering of paired ZDHHC hole mutants and bumped chemically tagged fatty acid probes enabled probe transfer to specific protein substrates with excellent selectivity over wild type ZDHHCs. Chemical genetic systems were exemplified for five ZDHHCs (3, 7, 11, 15 and 20), and applied to generate the first de novo ZDHHC substrate profiles, identifying >300 unique and shared substrates across multiple cell lines and S-acylation sites for novel functionally diverse substrates. We expect that this powerful and versatile platform will open a new window on S-acylation biology for a wide range of models and organisms.