BACKBONE PROTECTION AND ITS APPLICATION TO THE SYNTHESIS OF A DIFFICULT PHOSPHOPEPTIDE SEQUENCE

The N-(2-hydroxy-4-methoxybenzyl) (Hmb) backbone amide-protecting grou p has been applied to the synthesis of phosphopeptides via post-assemb ly global phosphorylation. Reversible protection of the Hmb 2-hydroxy moiety was mandatory in order to prevent its phosphorylation and resul tant irreversible stabilisation to acidolysis. This was achieved throu gh the use of either the acetyl (Ac) or allyloxycarbonyl (Alloc) group s, introduced through their respective anhydrides in the presence of t ertiary base. Following global phosphorylation, Ac or Alloc could be r emoved from resin-bound Hmb backbone-substituted fully protected phosp hopeptide by either hydrazinolysis or palladium-catalysed cleavage to re-establish Hmb acid-lability. Similarly, hydrazinolysis in solution of otherwise deprotected, backbone-substituted phosphopeptide was foun d to be efficient and free from phosphoroamino acid beta-elimination s ide-reactions. The optimised protocols were applied in the preparation of peptides from human Tau [390-406; Ser(PO3H)(396)] and the MAP kina se ERK 2 [178-188; Thr(PO3H)(183); Tyr(PO3H)(185)].