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The amyloid pathology associated with long-term haemodialysis is due to the deposition of (2)-microglobulin, the non-polymorphic light chain of class I major histocompatibility complex, that accumulates at bone joints into amyloid fibrils. Several lines of evidence show the relevance of the tryptophan residue at position 60 for the fibrillogenic transition of the protein. A comparative N-15 NMR relaxation analysis is presented for wild-type human (2)-microglobulin and W60G (2)-microglobulin, i.e. the mutant with a glycyne replacing the natural tryptophan residue at position 60. The experimental data, collected at 11.4T and 310K, were analyzed by means of the reduced spectral density approach. Molecular dynamics (MD) simulations and corresponding thermodynamic integration, together with hydrodynamic calculations were performed to support data interpretation. The analysis results for the mutant protein are consistent with a reduced aggregation with respect to the wild-type counterpart, as a consequence of an increased conformational rigidity probed by either NMR relaxation and MD simulations. Although dynamics in solution is other than fibrillar competence, the assessed properties of the mutant protein can be related with its reduced ability of forming fibrils when seeded in 20% trifluoroethanol. Copyright (c) 2013 John Wiley & Sons, Ltd. MIURMinistero dell' Istruzione, dell' Universita e della Ricerca (MIUR) [RBRN07BMCT, 20083ERXWS]; EUEuropean Union (EU) [LSHM-CT-2005-037525] This work was supported by MIUR (grants FIRB No. RBRN07BMCT, PRIN No. 20083ERXWS), and by EU (EURAMY project LSHM-CT-2005-037525). Dr Rotter M. and D. Makek A. are acknowledged for the help with calculations and assistance. DG thanks Dr Alexandros S. Papadopoulos for the helpful discussion on statistical aspects of data analysis.

"Amyloidosis associated to hemodialysis is caused by persistently high beta(2)-microglobulin (beta(2)m) serum levels. beta(2)m is an intrinsically amyloidogenic protein whose capacity to assemble into amyloid fibrils in vitro and in vivo is concentration dependent; no beta(2)m genetic variant is known in the human population. We investigated the roles of two evolutionary conserved Trp residues in relation to beta(2)m structure, function and folding/misfolding by means of a combined biophysical and functional approach. We show that Trp60 plays a functional role in promoting the association of beta(2)m in class I major histocompatibility complex; it is exposed to the solvent at the apex of a protein loop in order to accomplish such function. The Trp60 -> Gly mutation has a threefold effect: it stabilizes beta(2)m, inhibits beta(2)m amyloidogenic propensity and weakens the interaction with the class I major histocompatibility complex heavy chain. On the contrary, Trp95 is buried in the beta(2)m core; the Trp95 -> Gly mutation destabilizes the protein, which is unfolded in solution, yielding nonfibrillar beta(2)m aggregates. Trp60 and Trp95 therefore play differential and complementary roles in beta(2)m, being relevant for function (Trp60) and for maintenance of a properly folded structure (Trp95) while affecting in distinct ways the intrinsic propensity of wild-type beta(2)m towards self-aggregation into amyloid fibrils. (c) 2008 Elsevier Ltd. All rights reserved."