Amyloid fibrils form in supersaturated solutions via a nucleation and growth mechanism. Although the structural features of amyloid fibrils have become increasingly clearer, knowledge on the thermodynamics of fibrillation is limited. Furthermore, protein aggregation is not a target of calorimetry, one of the most powerful approaches used to study proteins. Here, with β2-microglobulin (β2m), a protein responsible for dialysis-related amyloidosis, we show direct heat measurements of the formation of amyloid fibrils using isothermal titration calorimetery (ITC). The spontaneous fibrillation after a lag phase was accompanied by exothermic heat. The thermodynamic parameters of fibrillation obtained under various protein concentrations and temperatures were consistent with the main-chain dominated structural model of fibrils, in which overall packing was less than that of the native structures. We also characterized the thermodynamics of amorphous aggregation enabling the comparison of protein folding, amyloid fibrillation and amorphous aggregation. These results indicate that ITC will become a promising approach for clarifying comprehensively the thermodynamics of protein folding and misfolding.
Tatsuya Ikenoue(a), Young-Ho Lee(a), József Kardos(b), Hisashi Yagi(a), Takahisa Ikegami(a), Hironobu Naiki(c)and Yuji Goto(a).
(a)Institute for Protein Research, Osaka University, Yamadaoka 3-2, Suita, Osaka 565-0871, Japan.
(b)Department of Biochemistry, Eötvös Loránd University, Pázmány sétány 1/C, Budapest, 1117, Hungary.
(c)Faculty of Medical Sciences, University of Fukui, Matsuoka, Fukui 910-1193, Japan.