Modification of β2-microglobulin with D-glucose or 3-deoxyglucosone inhibits Aβ2M amyloid fibril extensionin vitro

Abstract

β₂-microglobulin (β2M) is a major constituent of amyloid fibrils (fAβ2M) deposited in patients with Aβ2M amyloidosis. Recently, advanced glycation end products (AGE) of β2M and fAβ2M have been suggested to play an important role in the pathogenesis of APβM amyloidosis. We first characterized the states of AGE modification of fAβ2M. Western blot analysis with a monoclonal anti-AGE antibody showed that purified fAβ2M was naturally modified with AGE. Immunohistochemical studies of amyloid-deposired tissue have revealed a patchy distribution of the AGE-modified area in the amyloid deposits. Then we modified β2-m either with D-glucose or with 3-deoxyglucosone (3-DG) and investigated the effect of these modification on fAβ2M extension in vitro, using the recently established first-order kinetic model of fAβ2M extension in vitro. Western blot analysis and enzyme linked immunosorbent assay with a monoclonal anti-AGE antibody showed that these sugar-modfied β2M contained AGE. During the incubation of fAβ2M with native β2-m at 37°C the fluorescence of thioflavin T increased without a lag phase and proceeded to equilibrium. On the contrary, only a slight increase in fluorescence was observed during the incubation of fAβ2M with sugar-modified β2M. Moreover, sugar-modified β2M exhibited a dose-dependent inhibitory effect on the extension reaction of fAβ2Mwith native β2M. These results may suggest that in some in vivo situations, the modification of β2-m with AGE could play an inhibitory role for the formation of fAβ2M.