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. Author manuscript; available in PMC: 2014 Dec 8.
Published in final edited form as: J Proteomics Bioinform. 2014 Aug 25;7(9):256–263. doi: 10.4172/jpb.1000327

Figure 2. Studying S-acylation changes at the protein level has its inherent limitations for proteins with two or more S-acylation sites.

Figure 2

The mutation or reduced expression of a DHHC-PAT results in decreased S-acylation level of certain substrate S-acylation sites. In the currently dominant strategy (left panel), S-acylated proteins are purified using the MLCC method and digested into tryptic peptides for LC-MS analysis. However, for a dually or multiply S-acylated protein, if the S-acylation level for one site is unchanged, then the same amount of S-acylated proteins will be purified before and after DHHC-PAT loss. Because with the MLCC method almost all S-acylated peptides get lost, it wrongly appears that the S-acylated protein is unchanged and thus not the substrate for the DHHC-PAT. In comparison, the site-specific analysis strategy is more appropriate to determine whether a dually or multiply S-acylated protein is a DHHC-PAT substrate (right panel). This strategy can easily distinguish substrate S-acylation sites from non-substrate S-acylation sites on the same protein.