Open AccessLow-Background Acyl-Biotinyl Exchange Largely Eliminates the Coisolation of Non-S-Acylated Proteins and Enables Deep S-Acylproteomic Analysis
Author(s) -
Bo Zhou,
Yang Wang,
Yongnian Yan,
Javier Mariscal,
Dolores Di Vizio,
Michael R. Freeman,
Wei Yang
Publication year - 2019
Publication title -
analytical chemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.117
H-Index - 332
eISSN - 1520-6882
pISSN - 0003-2700
DOI - 10.1021/acs.analchem.9b01520
Subject(s) - acylation , chemistry , palmitoylation , lncap , cysteine , proteomics , biotinylation , biotin , myristoylation , quantitative proteomics , biochemistry , cancer , prostate cancer , gene , phosphorylation , medicine , enzyme , catalysis
Protein S -acylation (also called palmitoylation) is a common post-translational modification whose deregulation plays a key role in the pathogenesis of many diseases. Acyl-biotinyl exchange (ABE), a widely used method for the enrichment of S -acylated proteins, has the potential of capturing the entire S -acylproteome in any type of biological sample. Here, we showed that current ABE methods suffer from a high background arising from the coisolation of non- S -acylated proteins. The background can be substantially reduced by an additional blockage of residual free cysteine residues with 2,2'-dithiodipyridine prior to the biotin-HPDP reaction. Coupling the low-background ABE (LB-ABE) method with label-free proteomics, 2 895 high-confidence candidate S -acylated proteins (including 1 591 known S -acylated proteins) were identified from human prostate cancer LNCaP cells, representing so-far the largest S -acylproteome data set identified in a single study. Immunoblotting analysis confirmed the S -acylation of five known and five novel prostate cancer-related S -acylated proteins in LNCaP cells and suggested that their S -acylation levels were about 0.6-1.8%. In summary, the LB-ABE method largely eliminates the coisolation of non- S -acylated proteins and enables deep S -acylproteomic analysis. It is expected to facilitate a much more comprehensive and accurate quantification of S -acylproteomes than previous ABE methods.