Functionalization of amyloid fibrils via the Bri2 BRICHOS domain

Rakesh Kumar,Anna Katharina Schellhaus,Henrik Biverstål,Axel Abelein,Médoune Sarr,Jan Johansson,Nico P Dantuma

doi:10.1038/s41598-020-78732-1

Rakesh Kumar, Anna Katharina Schellhaus + Show 5 more

Open Access

https://doi.org/10.1038/s41598-020-78732-1

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Abstract

Amyloid fibrils are mechanically robust and partly resistant to proteolytic degradation, making them potential candidates for scaffold materials in cell culture, tissue engineering, drug delivery and other applications. Such applications of amyloids would benefit from the possibility to functionalize the fibrils, for example by adding growth factors or cell attachment sites. The BRICHOS domain is found in a family of human proteins that harbor particularly amyloid-prone regions and can reduce aggregation as well as toxicity of several different amyloidogenic peptides. Recombinant human (rh) BRICHOS domains have been shown to bind to the surface of amyloid-β (Aβ) fibrils by immune electron microscopy. Here we produce fusion proteins between mCherry and rh Bri2 BRICHOS and show that they can bind to different amyloid fibrils with retained fluorescence of mCherry in vitro as well as in cultured cells. This suggests a “generic” ability of the BRICHOS domain to bind fibrillar surfaces that can be used to synthesize amyloid decorated with different protein functionalities.

Highlights

Amyloid fibrils are mechanically robust and partly resistant to proteolytic degradation, making them potential candidates for scaffold materials in cell culture, tissue engineering, drug delivery and other applications
His6-NT*-Bri[2] BRICHOS-mCherry protein was produced in E. coli, purified by immobilized metal affinity chromatography (IMAC), and cleaved with thrombin to release recombinant human Bri[2] BRICHOS-mCherry, which was isolated by a second IMAC step
The abilities of rh Bri[2] BRICHOS to inhibit amyloid fibril formation of Aβ42 is retained in the rh Bri[2] BRICHOS-mCherry fusion protein as shown by kinetics measured by thioflavin T (ThT) fluorescence (Fig. 1C), indicating that the BRICHOS function is not perturbed by linking it to mCherry

Summary

Introduction

Amyloid fibrils are mechanically robust and partly resistant to proteolytic degradation, making them potential candidates for scaffold materials in cell culture, tissue engineering, drug delivery and other applications. We demonstrate here that mCherry-linked Bri[2] BRICHOS can bind to the fibril surface of six different protein nanofibers, which include four typical human disease-related amyloid fibrils as well as a de novo-designed β-structure protein. The abilities of rh Bri[2] BRICHOS to inhibit amyloid fibril formation of Aβ42 is retained in the rh Bri[2] BRICHOS-mCherry fusion protein as shown by kinetics measured by thioflavin T (ThT) fluorescence (Fig. 1C), indicating that the BRICHOS function is not perturbed by linking it to mCherry.

Results

Conclusion