SIMULTANEOUS MULTIPLE SYNTHESIS AND SELECTIVE CONJUGATION OF CYCLIZEDPEPTIDES DERIVED FROM A SURFACE LOOP OF A MENINGOCOCCAL CLASS-1 OUTER-MEMBRANE PROTEIN

Starting from the alpha-(2,4-dimethoxybenzyl) ester of N-(9-fluorenylm ethoxycarbonyl)aspartic acid [Fmoc-Asp-ODmb], side-chain-protected res in-bound Fmoc-peptides containing an lon-1-(4,4-dimethyl-2,6-dioxocycl ohexylidene)ethyl lysyl [Lys(Dde)l residue were prepared. The C-termin al dimethoxybenzyl esters of aspartic acid were removed with 1% triflu oroacetic acid and 10% anisole in dichloromethane, followed by Fmoc-cl eavage in the usual manner. The resin-bound peptides were then cyclize d using -benzotriazolyloxy-tris-[N-pyrrolidino]phosphonium hexafluorop hosphate (PyBOP) in the presence of N-methylmorpholine. The (dimethyld ioxocyclohexylidene)ethyl groups of lysine were removed with 1% hydraz ine hydrate in N,N-dimethylacetamide, and the liberated side-chain ami no functions were modified by reaction with pentafluorophenyl S-acetyl mercaptoacetate (SAMA-OPfp). Finally, the peptides were side-chain dep rotected, with exception of the Lys(SAMA) residue, and cleaved from th e solid support with trifluoroacetic acid/anisole/ water, 95/2.5/2.5. Cyclic peptides comprising 7-14 amino acid residues were obtained empl oying this procedure. As a model conjugation, cyclo[Thr-Asn-Asn-Asn-Le u-Lys(SAMA)-Thr-Lys-Asp] was coupled with bromoacetamide. The same pep tide was also coupled with a bromoacetylpeptide to give a well defined peptide/peptide conjugate. Ail peptides were conjugated to bromoacety lated tetanus toroid for immunization purposes. (C) Munksgaard 1994.