Cysteine 29 is the major palmitoylation site on stomatin

The 31 kDa membrane protein stomatin was metabolically, labeled with tritia ted palmitic acid in the human amniotic cell line UAC and immunoprecipitate d. We show that the incorporated palmitate is sensitive to hydroxylamine, i ndicating the binding to cysteine residues. Stomatin contains three cystein es, By expressing a myc-tagged stomatin and substituting the three cysteine s by serine, individually or in combination, we demonstrate that Cys-29 is the predominant site of palmitoylation and that Cys-86 accounts for the rem aining palmitate labeling. Disruption of Cys-52 alone does not show any det ectable reduction of palmitic acid incorporation. Given the organization of stomatin into homo-oligomers, the presence of multiple palmitate chains is likely to increase greatly the affinity of these oligomers for the membran e and perhaps particular lipid domains within it. (C) 1999 Federation of Eu ropean Biochemical Societies.