A partially structured species of beta 2-microglobulin is significantly populated under physiological conditions and involved in fibrillogenesis.
The folding of beta(2)-microglobulin (beta(2)-m), the protein forming amyloid deposits in dialysis-related amyloidosis, involves formation of a partially folded conformation named I(2), which slowly converts into the native fold, N. Here we show that the partially folded species I(2) can be separated from N by capillary electrophoresis. Data obtained with this technique and analysis of kinetic data obtained with intrinsic fluorescence indicate that the I(2) conformation is populated to approximately 14 +/- 8% at equilibrium under conditions of pH and temperature close to physiological. In the presence of fibrils extracted from patients, the I(2) conformer has a 5-fold higher propensity to aggregate than N, as indicated by the thioflavine T test and light scattering measurements. A mechanism of aggregation of beta(2)-m in vivo involving the association of the preformed fibrils with the fraction of I(2) existing at equilibrium is proposed from these results. The possibility of isolating and quantifying a partially folded conformer of beta(2)-m involved in the amyloidogenesis process provides new opportunities to monitor hemodialytic procedures aimed at the reduction of such species from the pool of circulating beta(2)-m but also to design new pharmaceutical approaches that consider such species as a putative molecular target.
Chiti
F
F
Dipartimento di Scienze Biochimiche, Università di Firenze, Viale Morgagni 50, 50134 Florence, Italy.
De Lorenzi
E
E
Grossi
S
S
Mangione
P
P
Giorgetti
S
S
Caccialanza
G
G
Dobson
C M
CM
Merlini
G
G
Ramponi
G
G
Bellotti
V
V
D052160:Benzothiazoles; D002942:Circular Dichroism; D004396:Coloring Agents / Q000494:pharmacology; D003224:Congo Red / Q000494:pharmacology; D019075:Electrophoresis, Capillary; D005456:Fluorescent Dyes / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008027:Light; D008854:Microscopy, Electron; D008954:Models, Biological; D008956:Models, Chemical; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D012542:Scattering, Radiation; D013696:Temperature; D013844:Thiazoles / Q000494:pharmacology; D013997:Time Factors; D014466:Ultraviolet Rays; D001613:beta 2-Microglobulin / Q000737:chemistry* / Q000378:metabolism*
D052160:Benzothiazoles; D004396:Coloring Agents; D005456:Fluorescent Dyes; D013844:Thiazoles; D001613:beta 2-Microglobulin; C009462:thioflavin T; D003224:Congo Red