Selective modification of peptides
and proteins is of foremost
importance for the development of biopharmaceuticals and exploring
biochemical pathways, as well as other applications. Here, we present
a study on the development of a general and easily applicable selective
method for N-terminal acylation of biomolecules,
applying a new type of phenol esters. Key to the success was the development
of highly tunable phenol activators bearing in the ortho-position,
sulfonic acid or sulfonamide, acting as a steric shield for hydrolysis,
and electron-withdrawing groups in the other ortho- and para-position
for controlling the reactivity of the activated phenol esters. A library
of heptapeptides, testing all 20 natural amino acids positioned at
the N-terminal, were acylated in a selective manner
at the N-terminus. The majority showed high conversion
and excellent N
α-selectivity. Several
biologically relevant biomolecules, including DesB30 insulin and human
growth hormone, could also be modified at the N-terminal
in a highly selective way, exemplified by either a fluorophore or
a fatty acid sidechain. Finally, taking advantage of the possibility
to accurately adjust the reactivity of the phenol esters, we present
a potential strategy for the construction of dual active biopharmaceuticals
through the employment of a bifunctional acylation linker and demonstrate
its use in the creation of a GLP-1 insulin analogue, coupled through
the lysine residue of GLP-1 and the N-terminal PheB1 amine of DesB30 insulin.