Destruction of Amyloid Fibrils of a β2-Microglobulin Fragment by Laser Beam Irradiation

Daisaku Ozawa,Hisashi Yagi,Tadato Ban,Atsushi Kameda,Toru Kawakami,Hironobu Naiki,Yuji Goto

doi:10.1074/jbc.m805118200

Daisaku Ozawa, Hisashi Yagi + Show 5 more

Open Access

https://doi.org/10.1074/jbc.m805118200

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Abstract

To understand the mechanism by which amyloid fibrils form, we have been making real-time observations of the growth of individual fibrils, using total internal fluorescence microscopy combined with an amyloid-specific fluorescence dye, thioflavin T (ThT). At neutral pH, irradiation at 442 nm with a laser beam to excite ThT inhibited the fibril growth of beta(2)-microglobulin (beta2-m), a major component of amyloid fibrils deposited in patients with dialysis-related amyloidosis. Examination with a 22-residue K3 fragment of beta2-m showed that the inhibition of fibril growth and moreover the destruction of preformed fibrils were coupled with the excitation of ThT. Several pieces of evidence suggest that the excited ThT transfers energy to ground state molecular oxygen, producing active oxygen, which causes various types of chemical modifications. The results imply a novel strategy for preventing the deposition of amyloid fibrils and for destroying preformed amyloid deposits.

Highlights

Amyloid fibrils are associated with the pathogenesis of more than 20 serious diseases, including Alzheimer, Parkinson, and Huntington diseases, and dialysis-related amyloidosis [1, 2]
Our results suggest that it may be possible to selectively breakdown amyloid fibrils coupled with amyloid-specific dyes like thioflavin T (ThT), suggesting a new strategy for destroying amyloid fibrils leading to the prevention and treatment of amyloidosis
TIRFM images indicated the presence of short fibrils in each sample, confirming that amyloid-specific fluorescence from ThT enables to the visualization of both ␤2-m and K3 fibrils

Summary

Introduction

Amyloid fibrils are associated with the pathogenesis of more than 20 serious diseases, including Alzheimer, Parkinson, and Huntington diseases, and dialysis-related amyloidosis [1, 2]. Further understanding of the structure, mechanism of formation, and roles in amyloidosis is one of the most important issues of protein science today. Fluorescence microscopy (TIRFM) is combined with amyloidspecific thioflavin T (ThT) fluorescence (4 – 8) This technique provides important information about the morphology, growth rate, and extension direction of fibrils in real time at the single fibril level. We have applied the technique to the fibrils of ␤2-microglobulin (␤2-m) responsible for dialysis-related amyloidosis and amyloid ␤ associated with Alzheimer disease (4 – 8). ␤2-m is a major component of amyloid fibrils deposited in dialysis-related amyloidosis, a common and serious complication in patients receiving hemodialysis for more than 10 years (9 –11). An increase in the concentration of ␤2-m is the most important risk factor for fibrillation, how ␤2-m forms amyloid fibrils under physiological conditions is unknown

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