A Residue-specific Shift in Stability and Amyloidogenicity of Antibody Variable Domains

Abstract

Background: Amyloid fibrils of variable (V L ) domains are the main cause of death in light chain amyloidosis. Results: Conserved V L residue 2 is crucial for structural integrity and amyloidogenicity of V L domains. Conclusion: Our data reveal novel insights into the architecture of variable domains in general and the prerequisite for fibrillation. Significance: This is important for understanding the principles of antibody structure and amyloidogenicity.

A single residue switch reveals principles of antibody domain integrity

“…However, as we showed for a murine V L , the N-and C-terminal residues, i.e. Ile 2 and Arg 108 play a major role for the integrity of the domain (5,6). Together with other recent data (16), this revives the question of domain boundaries, especially in the context of the increased use of antibody fragments as diagnostics and therapeutics.…”

“…The structural elements determining antibody domain stability are not sufficiently understood. For the V L domain, we showed that the nature of the terminal residues is important for its integrity (5,6). To test the impact of C-terminal residues on an antibody domain that is followed by a linker and another Ig domain, we chose the MAK33 IgG1 C H 2 domain.…”

“…So far, mainly the hydrophobic core of the ␤-barrel and the disulfide bridge have been considered in this context (3,4). Recently, we found for the variable light chain (V L ) domain that residues at the very termini (Ile 2 and Arg 108 ) are important for the integrity of the domain (5,6). Furthermore, the constant light chain (C L ) domain was destabilized when the N-terminal Arg 108 was deleted (6).…”

ThT 101: a primer on the use of thioflavin T to investigate amyloid formation

“…Fibril formation is accompanied by changes in secondary structure. The native state of amyloidogenic precursor proteins can be highly diverse, ranging from purely a-helical (myoglobin [114]), mixtures of a-helices and b-sheets (insulin [115] and lysozyme [116]), a/b-mixed structures (HypF N-terminal domain [117]), b-barrels (Ig light chain [118]) or natively unfolded (a-synuclein [119] and glucagon [120]). Upon fibril formation, some proteins undergo first a decrease in ordered secondary structure and a concomitant increase in random coil structures, followed by a gain in b-sheet-rich features, which make up the backbone of amyloid fibrils.…”

“…However, there is not always a major change in secondary structure, since some proteins adopt the amyloid fibril structure directly from their b-sheet-rich native state (i.e. Ig light chains [118]) or via a b-sheet intermediate state [121]. Importantly, aggregated proteins often increase the noise of the signal due to light scattering propensity of amorphous and amyloid structures, which leads in some cases to unexpected far-UV spectra [122].…”