title: Structure, stability, and aggregation of β-2 microglobulin mutants: insights from a Fourier transform infrared study in solution and in the crystalline state.
abstract: β-2 microglobulin (β2m) is an amyloidogenic protein involved in dialysis-related amyloidosis. We report here the study of the structural properties of the protein in solution and in the form of single crystals by Fourier transform infrared (FTIR) spectroscopy and microspectroscopy. The investigation has been extended to four β2m mutants previously characterized by x-ray crystallography: Asp(53)Pro, Asp(59)Pro, Trp(60)Gly, and Trp(60)Val. These variants displayed very similar three-dimensional structures but different thermal stability and aggregation propensity, investigated here by FTIR spectroscopy. For each variant, appreciable spectral differences were found between the protein in solution and in single crystals, consisting in a downshift of the main β-sheet band and in better resolved turn and loop bands, indicative of reduced protein secondary structure dynamics in the crystalline state. Notably, the well-resolved spectra of the β2m crystalline variants enabled us to identify structural differences induced by the single amino acid mutations. Such differences encompass turn and loop structures that might affect the stability and aggregation propensity of the investigated β2m variants. This study highlights the potential of FTIR microspectroscopy to acquire useful structural information on protein crystals, complementary to the crystallographic analyses.
authors.affiliation: Department of Biotechnology and Biosciences, University of Milano-Bicocca, Milan, Italy.
authors.forename: Diletta
authors.initials: D
authors.lastname: Ami
authors.forename: Stefano
authors.initials: S
authors.lastname: Ricagno
authors.forename: Martino
authors.initials: M
authors.lastname: Bolognesi
authors.forename: Vittorio
authors.initials: V
authors.lastname: Bellotti
authors.forename: Silvia Maria
authors.initials: SM
authors.lastname: Doglia
authors.forename: Antonino
authors.initials: A
authors.lastname: Natalello
mesh_terms: D018360:Crystallography, X-Ray; D007700:Kinetics; D008958:Models, Molecular; D050505:Mutant Proteins / Q000737:chemistry* / Q000235:genetics; D009154:Mutation; D055503:Protein Multimerization; D055550:Protein Stability; D020836:Protein Structure, Quaternary; D012996:Solutions; D017550:Spectroscopy, Fourier Transform Infrared; D013696:Temperature; D001613:beta 2-Microglobulin / Q000737:chemistry* / Q000235:genetics
chemical_list: D050505:Mutant Proteins; D012996:Solutions; D001613:beta 2-Microglobulin