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

Area of Science:

• Biochemistry
• Molecular Biology
• Proteomics

Background:

• Zinc finger Asp-His-His-Cys motif-containing (ZDHHC) S-acyltransferases regulate protein function through S-acylation.
• Hundreds of proteins are substrates for ZDHHC enzymes, playing roles in physiology and disease.

Purpose of the Study:

• To develop a technology for directly mapping protein substrates of specific ZDHHC S-acyltransferases at the whole-proteome level in intact cells.
• To identify novel substrates and S-acylation sites for various ZDHHC enzymes.

Main Methods:

• Utilized structure-guided engineering of ZDHHC 'hole' mutants and chemically tagged fatty acid probes.
• Employed a chemical-genetic system for selective probe transfer to ZDHHC substrates.
• Applied the technology to five human ZDHHCs (3, 7, 11, 15, and 20).

Main Results:

• Successfully mapped protein substrates for specific ZDHHCs with high selectivity.
• Identified over 300 substrates and S-acylation sites across functionally diverse proteins.
• Generated de novo ZDHHC substrate profiles in multiple cell lines.

Conclusions:

• The developed technology enables direct, whole-proteome mapping of ZDHHC substrates in intact cells.
• This platform significantly expands the understanding of S-acylation biology.
• The findings are expected to facilitate research across various models and organisms.