Iodine‐Mediated Peptide Disulfide Bridging is Reversible and Sequence‐Dependent in Solid Support: Use of TEMPO in the Reaction Resolves the Challenge.

Iodine (I2) is an extremely popular reagent for disulfide assembly in solution, solid phase (pseudo‐dilute conditions) peptides, and immobilized materials. However, we noticed that I2‐mediated peptide disulfide assembly often resulted in incomplete reactions due to its redox activity under pseudo‐dilute conditions. The phenomenon intrigued us to conduct an in‐depth analysis of I2‐mediated disulfide assembly with various peptide sequences. We revealed that the redox equilibrium between free thiols and disulfide products was peptide sequence‐dependent and previously unsolicited. Adding TEMPO in the reaction medium attenuates the I2‐mediated redox equilibrium and rapidly (∼5 min) promotes quantitative, clean disulfide assembly. Finally, in silico investigations supported that this chemical equilibrium primarily relies with the increased global free energy upon macrocyclization and the interspace between thiol groups in the linear peptide chain. This fundamental study will leverage multiple advantages in disulfide formation in immobilized materials and peptide modification chemistries with iodine in solid support. [ABSTRACT FROM AUTHOR]