Thiazolidine formation as a general and site-specific conjugation method for synthetic peptides and proteins

A general and mild method for the conjugation of synthetic peptides and proteins through site-specific thiazolidine formation was developed. This method is based on the specific reaction of the 1,2-aminothiol of cysteine with an aldehyde under acidic conditions to form a stable thiazolidine product. The carbonyl precursors are derived from the 1,2-aminoalcohols of Ser or Thr or the vicinal cis-diols of glycoproteins, which can be selectively and rapidly converted to the aldehyde form by periodate oxidation. Since the position where the aldehyde is generated can be manipulated or predetermined, the thiazolidine formation is essentially site-specific. The potential of this method is demonstrated by the biotinylation of three synthetic peptides at the N-terminal Ser or Thr and at the carbohydrate sites of alpha 1-acid glycoprotein. 1-Biotinyl-6-cysteinyl-hexadiamide, a new, water-soluble, long-chained, 1,2-aminothiol derivate containing biotin, was prepared as a capture device. This reagent reacted rapidly with periodate-oxidized N-terminal Ser or Thr peptides and with glycoproteins under very mild conditions to form the thiazolidine conjugates. Based on our results, the thiazolidine conjugation method would be useful for the attachment of radiolabels, reporter groups, imaging agents, and cytotoxic drugs to peptides and proteins.