Abstract
Immunoglobulin light chain-derived (AL) amyloidosis is a debilitating disease without known cure. Almost nothing is known about the structural factors driving the amyloidogenesis of the light chains. This study aimed to identify the fibrillogenic hotspots of the model protein 6aJL2 and in pursuing this goal, two complementary approaches were applied. One of them was based on several web-based computational tools optimized to predict fibrillogenic/aggregation-prone sequences based on different structural and biophysical properties of the polypeptide chain. Then, the predictions were confirmed with an ad-hoc synthetic peptide library. In the second approach, 6aJL2 protein was proteolyzed with trypsin, and the products incubated in aggregation-promoting conditions. Then, the aggregation-prone fragments were identified by combining standard proteomic methods, and the results validated with a set of synthetic peptides with the sequence of the tryptic fragments. Both strategies coincided to identify a fibrillogenic hotspot located at the CDR1 and β-strand C of the protein, which was confirmed by scanning proline mutagenesis analysis. However, only the proteolysis-based strategy revealed additional fibrillogenic hotspots in two other regions of the protein. It was shown that a fibrillogenic hotspot associated to the CDR1 is also encoded by several κ and λ germline variable domain gene segments. Some parts of this study have been included in the chapter “The Structural Determinants of the Immunoglobulin Light Chain Amyloid Aggregation”, published in Physical Biology of Proteins and Peptides, Springer 2015 (ISBN 978-3-319-21687-4).
Highlights
Www.nature.com/scientificreports characteristics of amyloid[11]
These findings suggest that as yet unidentified amyloidogenic sequences encoded in the germline IGLV6-57 could be part of the structural factors that make λ6 light chains highly prone to aggregate as amyloid fibrils[17,24,25]
We show that a fibrillogenic hotspot centre within the Complementary Determining Region 1 (CDR1) is encoded by other germline gene segments belonging to different κ and λ variable domain (VL) subgroup, indicating the general applicability of our findings for AL
Summary
Www.nature.com/scientificreports characteristics of amyloid[11]. it has been shown that the aggregation of the full-length amyloid precursor can be modulated by targeting their fibrillogenic region(s) with site-directed mutagenesis or by the binding with specific ligands[12,13]. We have shown that 6aJL2 can form amyloid-like fibrils in physiological relevant conditions of pH, temperature, and ionic strength[24] These findings suggest that as yet unidentified amyloidogenic sequences encoded in the germline IGLV6-57 could be part of the structural factors that make λ6 light chains highly prone to aggregate as amyloid fibrils[17,24,25]. The aggregation-prone fragments were identified by combining standard proteomic methods, and the results validated by analysing the aggregation behaviour of a set of synthetic peptides with the sequence of the tryptic fragments Both strategies coincided to identify a fibrillogenic hotspot located at the CDR1 and β-strand C of the protein, which was confirmed by scanning proline mutagenesis analysis. We show that a fibrillogenic hotspot centre within the CDR1 is encoded by other germline gene segments belonging to different κ and λ VL subgroup, indicating the general applicability of our findings for AL
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