Thiopalmitoylation (i.e., the covalent attachment of
palmitic acid via a thioester linkage to
cysteine residues in the
polypeptide backbone) is a common post-translational modification of
proteins. Several
proteins that have been identified as putative
autoantigens in a variety of T-cell mediated
autoimmune diseases are thiopalmitoylated, and thus, we have hypothesized that endogenous thiopalmitoylated
peptides released during tissue breakdown may play a role in the development and chronicity of
autoimmune diseases. To investigate this, we have studied the effect of thiopalmitoylation on the immunogenic and neuritogenic properties of P0, the major peripheral nervous system (PNS)
myelin protein, which is thiopalmitoylated at
cysteine 153, and described as a candidate
autoantigen in
Guillain-Barre syndrome (GBS), a human inflammatory
demyelinating disease of the PNS. This paper describes the synthesis of palmitoylated
peptide P0(180-199) and P0(152-171) by on-resin acylation using specific
cysteine side-chain protecting groups:
Mmt (labile in diluted
acid) and StBu (labile in the presence of
tributylphosphine). Our results show that the
thiol protecting group had to be adjusted to the
peptide sequence:
Mmt was efficiently used for P0(180-199) thioacylation, but it was not suitable for thiopalmitoylation of P0(152-171) because of a premature deprotection of the Boc protecting group on the epsilon-NH(2) Lys in the presence of 2% TFA, leading to dipalmitoylation. Palmitoylated P0(152-171) was successfully obtained by using StBu as the
thiol protecting group. We could show by circular dichroism that palmitoylation has no influence on the structuration of the
peptide in
solution but palmitoylation increased the stability of the
peptide in the presence of serum. Using
EAN (experimental autoimmune neuritis), the rat model of GBS, we have compared the immunological properties of palm and non-palm P0
peptides and showed that thiopalmitoylation has indeed a great influence on their neuritogenic and immunogenic properties. This study provides further support for our hypothesis concerning the role of thiopalmitoylation in the development and chronicity of inflammatory
demyelinating diseases and confirms that thiopalmitoylation of
peptides may provide a simple means to increase MHC class II restricted responses.