Hetero-assembly of a dual β-amyloid variant peptide system

Abstract

Self-assembly of amyloid polypeptides (1) imparts biological effects depending on the size in over 20 amyloid diseases and (2) produces useful yet relatively untapped biomaterials. Unfortunately, our understanding of amyloid polypeptides, as related to biomedical implications and biomaterial applications, is limited by their self-assembling nature. In this study, we report the creation of a dual peptide system, where a pair of β-amyloid (Aβ) variants are not self-assembled but hetero-assembled in the presence of their assembly partners. We provide evidence that the resulting hetero-assemblies share molecular, structural and morphological similarities with typical amyloid self-assemblies formed by a single polypeptide (e.g., Aβ). We anticipate that our dual peptide system may readily be adapted for precise control of amyloid assembly, for the study of size-dependent neurotoxicity and precise fabrication of amyloid biomaterials.

Amyloid Aggregation of Bacillus circulans Xylanase under Native Conditions and its Modulation by β‐Amyloid‐Derived Peptide Fragments

“…After 14 days′ incubation, fibrillar aggregates were detected in all BCX and BCX‐KLVFWAK samples with the added peptides (Figure S4); this is consistent with the significant ThT fluorescence values at the end of the incubation. Similar to KLVFWAK, neither significant ThT fluorescence (Figure A) nor fibrillar aggregates were detected with free ELVFWAE alone during the incubation (Figure E), thus indicating little to no self‐assembly to form amyloid aggregates, as reported previously . The amyloid aggregation‐modulating effects of the peptide were dose‐dependent: for example, a lag phase prior to the onset of ThT fluorescence appeared longer with higher concentrations of free ELVFWAE present in BCX‐KLVFWAK solution (Figure S5).…”

“…Amyloidogenic sequences can often be engineered for reduced aggregation propensity by simple point mutations. For example, the fragment KLVFFAE, which is critical in the amyloid self‐assembly of Aβ, is highly self‐aggregating whereas its variants, KLVFWAK and ELVFWAE, are not, due to electrostatic self‐repulsion …”

“…During the 14‐day incubation, the CD signals of BCX‐KLVFWAK were gradually weakened, which is accounting for by the reduction in soluble protein fractions and/or the loss of secondary structure (Figure B), accompanied by the formation of fibrils in BCX‐KLVFWAK samples (Figure C). As reported previously, KLVFWAK itself was shown to display limited amyloid‐assembly under the incubation conditions—ThT fluorescence of KLVFWAK remained negligible during the 14‐day incubation (Figure A)—this is confirmed by the lack of amyloid fibrils (Figure D) due to electrostatic repulsion caused by the terminal lysine residues . The expression level of a BCX variant with an N‐terminal fusion of KLVFWAK was significantly low and, thus, the fusion was not studied further.…”

“…The expression level of a BCX variant with an N‐terminal fusion of KLVFWAK was significantly low and, thus, the fusion was not studied further. We also attempted C‐terminal fusion to BCX of a KLVFWAK variant, ELVFWAE, in which positive lysine residues were replaced by negative glutamate residues . The ELVFWAE fusion to BCX resulted in negligible expression as well, and, thus, was not studied further either.…”

“…C‐terminal fusion of KLVFWAK, however, accelerated the amyloid aggregation of BCX and lowered BCX′s kinetic stability, which could result from local rather than global structural alteration under native conditions. Interestingly, despite the limited self‐aggregation of KLVFWAK as described above (Figure A) and previously, a part of this fragment, in particular LVFWA, is still highly amyloidogenic, as judged by Tango score of ≈80 (Figure S8). Thus, one possible explanation for the observed reduction in lag phase for BCX amyloid aggregation by C‐terminal fusion of KLVFWAK (Figure A) might be the LVFWA sequence, which could contribute to assembly among BCX‐KLVFWAK.…”