Conjugation of epitope peptides with SH group to branched chain polymeric polypeptides via Cys(Npys)

Since bioconjugates may play an important role as therapeutics in the future, the development of new and effective conjugation strategies is necessary. For the attachment of peptide-like molecules to carriers, there are two main coupling methods involving amide or disulfide bonds. Conjugation through an amide bond can be achieved in several well-defined ways known from peptide chemistry. However, the formation of disulfide bridges between cysteine-containing peptides and carrier molecules still has some problems. In this paper, we describe a novel approach in which the carrier polypeptide is modified by 3-nitro-2-pyridinesulfenyl (Npys)-protected cysteine and this derivative has been applied for conjugation of Cys-containing epitope peptides with poly(L-lysine)-based branched polypeptides. Considering the stability of Npys group in the presence of pentafluorophenol, Boc-Cys(Npys)-OPfp dervivative was selected for introduction to the N-terminal of branches of polypeptides backbone. The branches of the polymers were built up from oligo(DL-alanine) (poly[Lys(DL-Ala(m))], AK) and elongated by an optically active amino acid [poly[Lys(X(i)-DL-Ala(m))], XAK]. We found that the nature of X (Glu, Ser, Thr) has great influence on the incorporation of the protected cysteine residue. Herpes simplex virus and adenovirus epitope peptides were conjugated to Boc-Cys(Npys)-modified polypeptides. Results indicate that the incorporation of epitope peptides depends on the number of Npys group on the polymers as well as on the presence/absence of Boc-protecting group on the Cys residue. This new class of Cys(Npys)-derivatized branched polypeptides is stable for a couple of months and suitable for effective preparation of epitope peptide conjugates possessing increased water solubility.