Semi‐Synthesis of Immunogenic Mycobacterial Lipoproteins via Aryl Selenoester‐Mediated Expressed Protein Ligation

Tuberculosis (TB), caused by infection with the bacterium Mycobacterium tuberculosis ( Mtb ), remains one of the most prevalent infectious diseases worldwide. Despite decades of dedicated efforts to develop effective prevention strategies, including vaccines, TB continues to cause significant morbidity and mortality globally. Beyond the Bacille Calmette–Guérin (BCG) vaccine, limited progress has been made to develop more effective TB vaccines. A better understanding of the immunomodulatory roles of key Mtb protein virulence factors is therefore needed for the development of more efficacious protein‐based vaccine candidates. Herein, we report a highly efficient method for the semi‐synthesis of two native mycobacterial lipoproteins and glycolipoproteins, LprA and Mpt83, respectively. Capitalising on the enhanced reactivity of peptide selenoesters as acyl donors, the 21–23 kDa homogeneously lipidated proteins could be generated in a single ligation step. The homogeneous mycobacterial lipoprotein and glycolipoprotein molecules were shown to be agonists of Toll‐like receptor 2 (TLR2) and led to the potent induction of pro‐inflammatory cytokines and chemokines in cells. Taken together, this work presents a robust semi‐synthetic platform for accessing lipoproteins involved in host–pathogen interactions that can help guide future TB vaccine design.