MAK33 antibody light chain amyloid fibrils are similar to oligomeric precursors

Abstract

Little structural information is available so far on amyloid fibrils consisting of immunoglobulin light chains. It is not understood which features of the primary sequence of the protein result in fibril formation. We report here MAS solid-state NMR studies to identify the structured core of κ-type variable domain light chain fibrils. The core contains residues of the CDR2 and the β-strands D, E, F and G of the native immunoglobulin fold. The assigned core region of the fibril is distinct in comparison to the core identified in a previous solid-state NMR study on AL-09 by Piehl at. al, suggesting that VL fibrils can adopt different topologies. In addition, we investigated a soluble oligomeric intermediate state, previously termed the alternatively folded state (AFS), using NMR and FTIR spectroscopy. The NMR oligomer spectra display a high degree of similarity when compared to the fibril spectra, indicating a high structural similarity of the two aggregation states. Based on comparison to the native state NMR chemical shifts, we suggest that fibril formation via domain-swapping seems unlikely. Moreover, we used our results to test the quality of different amyloid prediction algorithms.

A Substantial Structural Conversion of the Native Monomer Leads to in‐Register Parallel Amyloid Fibril Formation in Light‐Chain Amyloidosis

“…Both proteins have been comprehensively characterized by in vitro fibrillization, and their monomer structures have been determined by X‐ray crystallography and solution‐state NMR . Two recent studies determined secondary‐structure elements of the V L s from AL‐09 and MAK33 light‐chain fibrils, both belonging to the κ‐family . In‐register parallel arrangements of β‐sheets have been suggested for short light‐chain fragments in the context of cryo‐EM studies .…”

“…The flexibility introduced by the mutation might help to form a compatible arrangement between the β‐sheets and the disulfide bonds, and ease the α‐helix‐to‐β‐strand transition of the CDR1 . One should note that for the mutant forms of the V L domains of MAK33 (S20N) and AL‐09 (seven mutations), S20N and six out of seven mutations in AL‐09 are located outside the assigned β‐strand regions, (which are restricted to the very N‐ and C‐terminal regions in AL‐09, and to residues 60–87 in MAK33). Both fibrils were fibrillized under highly acidic conditions (pH 2).…”

Insights into protein misfolding and aggregation enabled by solid-state NMR spectroscopy

“…Others have advocated domain swapping as a potential contributor to this type of aggregation process, noting a correlation between domain swapping propensity and aggregation propensity. To the best of our knowledge, truly domain-swapped conformations have not yet been demonstrated within amyloid fibrils, but may be part of non-amyloid aggregates or pre-amyloid oligomers [32–35]. …”

“…The study of the variable domain of AL-09 included a comparison to patient-derived material, based not on the seeding of labeled protein, but rather the direct acquisition of patient material’s natural abundance 13 C spectrum. The fibrillar and oligomeric states of the variable domain of MAK33 were very similar to each other in terms of their ssNMR spectra, whilst the fibrils were also reported not to reflect a domain swapped state [35]. Interestingly, the regions that form the amyloid core seem to differ between the two light chain variants, arguing that further ssNMR studies will be valuable to probe what (if any) commonalities are present between different amyloidogenic light chains.…”

Protein resonance assignment by BSH‐CP‐based 3D solid‐state NMR experiments: A practical guide

“…BSH‐CP‐based experiments are suited for a wide range of samples and have already been successfully applied to different kinds of proteins such as membrane proteins, amyloid fibrils, and microcrystalline protein samples, as well as protein filaments and supramolecular assemblies …”