A conformational fingerprint for amyloidogenic light chains
Curation statements for this article:-
Curated by eLife
eLife Assessment
This important study identifies the "H-state" as a potential conformational marker distinguishing amyloidogenic from non-amyloidogenic light chains, addressing a critical problem in protein misfolding and amyloidosis. By combining advanced techniques such as small-angle X-ray scattering, molecular dynamics simulations, and H-D exchange mass spectrometry, the authors provide convincing evidence for their novel findings. However, incomplete experimental descriptions, limitations in SAXS data interpretation, and the way HDX MS data is presented affect the strength and generalizability of the conclusions. Strengthening these aspects would enhance the impact of this work for researchers in amyloidosis and protein misfolding.
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Abstract
Immunoglobulin light chain amyloidosis (AL) shares with multiple myeloma (MM) the overproduction of one clonal light chain (LC), but whereas in MM patients LC molecules remain soluble in circulation, AL LCs misfold into toxic soluble species and amyloid fibrils that accumulate in internal organs, leading to completely different clinical manifestations. The large sequence variability of LCs has hampered our understanding of the mechanism leading to LC aggregation. Nevertheless, some biochemical properties associated with AL-LC are emerging. The stability of the dimeric LCs seems to play a role, but conformational dynamics and susceptibility to proteolysis have been identified as biophysical parameters that, under native conditions, can better distinguish AL-LCs from LCs found in MM. In this study, our goal was to delineate a conformational fingerprint that could discriminate AL from MM LCs. By subjecting four AL and two MM LCs to in vitro analysis under native conditions using small-angle X-ray scattering (SAXS), we observed that the AL LCs exhibited a slightly larger radius of gyration and greater deviation from the experimentally determined structure, indicating enhanced conformational dynamics. Integrating SAXS with molecular dynamics (MD) simulations to generate a conformational ensemble revealed that LCs can adopt multiple states, with VL and CL domains either bent or straight. AL-LCs favored a distinct state in which both domains were in a straight conformation, maximizing solvent accessibility at their relative interfaces. This unique conformation was experimentally validated by hydrogen-deuterium exchange mass spectrometry (HDX-MS). Such findings reconcile a wealth of experimental observations and provide a precise structural target for drug design investigations.
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eLife Assessment
This important study identifies the "H-state" as a potential conformational marker distinguishing amyloidogenic from non-amyloidogenic light chains, addressing a critical problem in protein misfolding and amyloidosis. By combining advanced techniques such as small-angle X-ray scattering, molecular dynamics simulations, and H-D exchange mass spectrometry, the authors provide convincing evidence for their novel findings. However, incomplete experimental descriptions, limitations in SAXS data interpretation, and the way HDX MS data is presented affect the strength and generalizability of the conclusions. Strengthening these aspects would enhance the impact of this work for researchers in amyloidosis and protein misfolding.
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Reviewer #1 (Public review):
The study investigates light chains (LCs) using three distinct approaches, with a focus on identifying a conformational fingerprint to differentiate amyloidogenic light chains from multiple myeloma light chains. The study's major contribution is the identification of a low-populated "H state," which the authors propose as a unique marker for AL-LCs. While this finding is promising, the review highlights several strengths and weaknesses. Strengths include the valuable contribution of identifying the H state and the use of multiple approaches, which provide a comprehensive understanding of LC structural dynamics. However, the study suffers from weaknesses, particularly in the interpretation of SAXS data, lack of clarity in presentation, and methodological inconsistencies. Critical concerns include high error …
Reviewer #1 (Public review):
The study investigates light chains (LCs) using three distinct approaches, with a focus on identifying a conformational fingerprint to differentiate amyloidogenic light chains from multiple myeloma light chains. The study's major contribution is the identification of a low-populated "H state," which the authors propose as a unique marker for AL-LCs. While this finding is promising, the review highlights several strengths and weaknesses. Strengths include the valuable contribution of identifying the H state and the use of multiple approaches, which provide a comprehensive understanding of LC structural dynamics. However, the study suffers from weaknesses, particularly in the interpretation of SAXS data, lack of clarity in presentation, and methodological inconsistencies. Critical concerns include high error margins between SAXS profiles and MD fits, unclear validation of oligomeric species in SAXS measurements, and insufficient quantitative cross-validation between experimental (HDX) and computational data (MD). This reviewer calls for major revisions including clearer definitions, improved methodology, and additional validation, to strengthen the conclusions.
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Reviewer #2 (Public review):
Summary:
This well-written manuscript addresses an important but recalcitrant problem - the molecular mechanism of protein misfolding in Ig light chain (LC) amyloidosis (AL), a major life-threatening form of systemic human amyloidosis. The authors use expertly recorded and analyzed small-angle X-ray scattering (SAXS) data as a restraint for molecular dynamics simulations (called M&M) and to explore six patient-based LC proteins. The authors report that a highly populated "H-state" determined computationally, wherein the two domains in an LC molecule acquire a straight rather than bent conformation, is what distinguishes AL from non-AL LCs. They then use H-D exchange mass spectrometry to verify this conclusion. If confirmed, this is a novel and interesting finding with potentially important translational …
Reviewer #2 (Public review):
Summary:
This well-written manuscript addresses an important but recalcitrant problem - the molecular mechanism of protein misfolding in Ig light chain (LC) amyloidosis (AL), a major life-threatening form of systemic human amyloidosis. The authors use expertly recorded and analyzed small-angle X-ray scattering (SAXS) data as a restraint for molecular dynamics simulations (called M&M) and to explore six patient-based LC proteins. The authors report that a highly populated "H-state" determined computationally, wherein the two domains in an LC molecule acquire a straight rather than bent conformation, is what distinguishes AL from non-AL LCs. They then use H-D exchange mass spectrometry to verify this conclusion. If confirmed, this is a novel and interesting finding with potentially important translational implications.
Strengths:
Expertly recorded and analyzed SAXS data combined with clever M&M simulations lead to a novel and interesting conclusion.
Regardless of whether or not the CL-CL domain interface is destabilized in AL LCs explored in this (Figure 6) and other studies, stabilization of this interface is an excellent idea that may help protect at least a subset of AL LCs from misfolding in amyloid. This idea increases the potential impact of this interesting study.
Weaknesses:
The HDX analysis could be strengthened.
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Reviewer #3 (Public review):
Summary:
This study identifies confirmational fingerprints of amylodogenic light chains, that set them apart from the non-amylodogenic ones.
Strengths:
The research employs a comprehensive combination of structural and dynamic analysis techniques, providing evidence that conformational dynamics at VL-CL interface and structural expansion are distinguished features of amylodogenic LCs.
Weaknesses:
The sample size is limited, which may affect the generalizability of the findings. Additionally, the study could benefit from deeper analysis of specific mutations driving this unique conformation to further strengthen therapeutic relevance.
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