The Canonical DHHC Motif Is Not Absolutely Required for the Activity of the Yeast S-acyltransferases Swf1 and Pfa4

Abstract

Background: Mutations in the DHHC motif of S-acyltransferases are thought to result in lack of activity. Results: The yeast S-acyltransferases Swf1 and Pfa4 are partially active in the absence of an intact DHHC motif. Conclusion: S-acylation may occur by alternative mechanisms. Significance: These results contribute to understanding the mechanism of protein S-acylation and suggest that proteins with divergent DHHC motifs might possess S-acyltransferase activity.

Protein palmitoylation: Palmitoyltransferases and their specificity

“…lysine, arginine, or protonated histidine side chains. 44 Indeed, the studies of Mitchell et al 41 and Gonzalez Montoro et al 12 suggest that the first histidine within the DHHC motif plays a significant role in the second step of the palmitoylation mechanism, involving the transfer of palmitoyl residue from PATs to the target protein. This might be due to histidine-aided deprotonation of the cysteine residues of the acceptor protein.…”

“…3,11,16 This is not the case, however, for two yeast PATs, Pfa4, and Swf1, because palmitoylation still occurred when their DHHC cysteine residue was mutated into arginine or alanine. 12 In view of the recent work of Hemsley and Grierson, 17 it might be explained that it is possible for PATs to be acylated in trans when the cysteine of the DHHC motif is mutated. Moreover, at least in the case of human DHHCs 5, 6, and 8, autoplamitoylation was detected in the C-terminal region defined as CCX7-13C(S/T) motif, where three cysteine residues are present.…”

“…11 On the contrary, a DQHC mutant of Swf1 undergoes autoacylation and has partial PAT activity. 12 Moreover, the human DHHC13 protein contains a natural DQHC motif and actively palmitoylates huntingtin protein and ClipR-59. 45,46 It is not only intramolecular factors that can modulate the mechanism of palmitoylation, but intermolecular factors can also be involved.…”

“…33 Most probably, the cysteine of the DHHC motif is the primary one which is autoacylated, as shown in many studies, although secondary sites of autoacylation might also exist. 12,18,42 From in vitro kinetics studies, the stoichiometry is one acyl chain per DHHC motif. 42 In order for autoacylation to occur, the target cysteine residue has to be in a deprotonated state as a thiolate anion that acts as a nucleophile for palmitoyl-CoA.…”

“…Recent findings, however, show that, despite the mutations in the DHHC motif of yeast S-acyltransferases Swf1 and Pfa4, their activity is partially preserved. 12 The labile character of the palmitic acid attachment suggests that acylation/deacylation cycles must be tightly regulated. Although it is not known how the process is regulated, it is possible that it might occur on different levels, e.g.…”