Tissue transglutaminase modulates α‐synuclein oligomerization

We have studied the interaction of the enzyme tissue transglutaminase (tTG), catalyzing cross‐link formation between protein‐bound glutamine residues and primary amines, with Parkinson's disease‐associated α‐synuclein protein variants at physiologically relevant concentrations. We have, for the first time, determined binding affinities of tTG for wild‐type and mutant α‐synucleins using surface plasmon resonance approaches, revealing high‐affinity nanomolar equilibrium dissociation constants. Nanomolar tTG concentrations were sufficient for complete inhibition of fibrillization by effective α‐synuclein cross‐linking, resulting predominantly in intramolecularly cross‐linked monomers accompanied by an oligomeric fraction. Since oligomeric species have a pathophysiological relevance we further investigated the properties of the tTG/α‐synuclein oligomers. Atomic force microscopy revealed morphologically similar structures for oligomers from all α‐synuclein variants; the extent of oligomer formation was found to correlate with tTG concentration. Unlike normal α‐synuclein oligomers the resultant structures were extremely stable and resistant to GdnHCl and SDS. In contrast to normal β‐sheet‐containing oligomers, the tTG/α‐synuclein oligomers appear to be unstructured and are unable to disrupt phospholipid vesicles. These data suggest that tTG binds equally effective to wild‐type and disease mutant α‐synuclein variants. We propose that tTG cross‐linking imposes structural constraints on α‐synuclein, preventing the assembly of structured oligomers required for disruption of membranes and for progression into fibrils. In general, cross‐linking of amyloid forming proteins by tTG may prevent the progression into pathogenic species.