Photoprotected Peptide α-Ketoacids and Hydroxylamines for Iterative and One-Pot KAHA Ligations: Synthesis of NEDD8

The convergent synthesis of proteins by multiple ligations requires segments protected at the N- and/or C-terminus with masking groups that are orthogonal to the acid-and base-labile protecting groups used in Fmoc-SPPS. They must be stable to solid-phase peptide synthesis, HPLC purification, and ligation conditions and easily removed in the presence of unprotected side chains. In this report, we document photolabile protecting groups for both alpha-ketoacids and hydroxylamines, the key functional groups employed in the alpha-ketoacid-hydroxylamine (KAHA) ligation. The novel photoprotected alpha-ketoacid is easily installed onto numerous different C-terminal peptide alpha-ketoacids and removed by UV light under aqueous conditions. These advances were applied to the one-pot synthesis of NEDD8, an important modifier protein, by three different convergent routes. These new protecting groups provide greater flexibility on the order of fragment assembly and reduce the number of reaction and purification steps needed for protein synthesis with the KAHA ligation.