Tissue transglutaminase modulates alpha-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 alpha-synuclein protein variants at physiologically relevant concentrations. We have, for the first time, determined binding affinities of tTG for wild-type and mutant alpha-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 alpha-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/alpha-synuclein oligomers. Atomic force microscopy revealed morphologically similar structures for oligomers from all alpha-synuclein variants; the extent of oligomer formation was found to correlate with tTG concentration. Unlike normal alpha-synuclein oligomers the resultant structures were extremely stable and resistant to GdnHCl and SDS. In contrast to normal beta-sheet-containing oligomers, the tTG/alpha-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 alpha-synuclein variants. We propose that tTG cross-linking imposes structural constraints on alpha-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.
Segers-Nolten
Ine M J
IM
Biophysical Engineering Group, MESA+ Institute for Nanotechnology and Institute for Biomedical Technology, University of Twente, 7500 AE Enschede, The Netherlands.
Wilhelmus
Micha M M
MM
Veldhuis
Gertjan
G
van Rooijen
Bart D
BD
Drukarch
Benjamin
B
Subramaniam
Vinod
V
D002942:Circular Dichroism; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007700:Kinetics; D018625:Microscopy, Atomic Force; D009154:Mutation; D010300:Parkinson Disease / Q000378:metabolism; D011485:Protein Binding; D020349:Surface Plasmon Resonance; D011503:Transglutaminases / Q000378:metabolism*; D051844:alpha-Synuclein / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D051844:alpha-Synuclein; D011503:Transglutaminases