Monitoring the interaction between ?2-microglobulin and the molecular chaperone ?B-crystallin by NMR and mass spectrometry: ?B-crystallin dissociates ?2-microglobulin oligomers.

ABSTRACT: The interaction at neutral pH between wild-type and a variant form (R3A) of the amyloid fibril-forming protein ?2-microglobulin (?2m) and the molecular chaperone ?B-crystallin was investigated by thioflavin T fluorescence, NMR spectroscopy, and mass spectrometry. Fibril formation of R3A?2m was potently prevented by ?B-crystallin. ?B-crystallin also prevented the unfolding and nonfibrillar aggregation of R3A?2m. From analysis of the NMR spectra collected at various R3A?2m to ?B-crystallin molar subunit ratios, it is concluded that the structured ?-sheet core and the apical loops of R3A?2m interact in a nonspecific manner with the ?B-crystallin. Complementary information was derived from NMR diffusion coefficient measurements of wild-type ?2m at a 100-fold concentration excess with respect to ?B-crystallin. Mass spectrometry acquired in the native state showed that the onset of wild-type ?2m oligomerization was effectively reduced by ?B-crystallin. Furthermore, and most importantly, ?B-crystallin reversibly dissociated ?2m oligomers formed spontaneously in aged samples. These results, coupled with our previous studies, highlight the potent effectiveness of ?B-crystallin in preventing ?2m aggregation at the various stages of its aggregation pathway. Our findings are highly relevant to the emerging view that molecular chaperone action is intimately involved in the prevention of in vivo amyloid fibril formation.

SUBMITTER: Esposito G 

PROVIDER: S-EPMC3682583 | biostudies-literature | 2013 Jun

REPOSITORIES: biostudies-literature