Application of Cystamine and N,N‘-Bis(glycyl)cystamine as Linkers in Polysaccharide−Protein Conjugation

Abstract

Pneumococcal polysaccharide type 6B, 14, or 23F (35−70 kDa) was activated with cyanogen bromide and modified with cystamine. After reduction of the spacer, the thiol-containing (i.e. cysteamine-modified) polysaccharide obtained was added in a 5−10-fold molar excess to bromoacetylated tetanus toxoid to give thioether-linked polysaccharide−protein conjugates in a yield of 10−20%. This approach failed for preparing a type 19F polysaccharide−protein conjugate, possibly due to intramolecular elimination of cysteamine from the reduced 19F polysaccharide. When N,N‘-bis(glycyl)cystamine was introduced as a spacer molecule, the elimination of the reduced spacer was suppressed, thus allowing preparation of a 19F polysaccharide−tetanus toxoid conjugate (15%).