Nerve growth factor is a powerful protein for treating central nervous system diseases, but its therapeutic potential is limited by a severe pain-inducing side effect. This adverse effect arises from insufficient receptor selectivity and can, in principle, be mitigated through side chain mutations or modifications, making it a suitable target for chemical protein synthesis. Despite its small size (13 kDa), the chemical synthesis of NGF has been challenging due to its exceptional hydrophobicity and the requirement for a 104-residue N-terminal chaperone peptide for folding. Here, we present the chemical synthesis of NGF using α-ketoacid-hydroxylamine (KAHA) ligations, featuring recombinant expression of the chaperone peptide and its chemoselective conversion to a C-terminal α-ketoacid.^[1] A novel solubility tag, SOLACE, and ester-forming KAHA ligations enabled the assembly of linear proNGF (25 kDa) from three synthetic and one recombinant segment. Controlled folding and disulfide bond formation mediated by the chaperone peptide followed by proteolytic cleavage yielded biologically active synthetic NGF. This work establishes a platform for engineering NGF variants with tailored receptor selectivity and offers a general strategy for the semisynthesis of neurotrophins and related proteins that also require long chaperone peptides for proper folding. 

[1] N. Y. Nötel, A. E. McMillan, V. R. Pattabiraman, K. Vulić, J. W. Bode, ACS Central Science, 2025, ASAP.