Herein, immunoglobulin light chains (LCs) native state was studied in the context of the pathology known as light chain amyloidosis (AL). This pathology is characterized by LCs overexpression, which leads to toxicity and aggregation into amyloid fibrils in target organs, with heart being the most affected one. Due to genetic rearrangement and somatic hypermutation, virtually, each AL patient presents a different amyloidogenic LC (Merlini, 2017). Because of such complexity, the fine molecular determinants of LC aggregation propensity and proteotoxicity are, to date, unclear; significantly, their decoding requires investigating large sets of cases. This project is aimed to unravel the molecular determinants linked with LCs toxicity. First, we screened several independent biophysical and structural properties of the LCs native state. In particular, we considered hydrophobicity, fold stability, flexibility and 3D structure. Our experimental approach considered two LCs sets called ‘H’ and ‘M’. The H set is composed of eight LCs from AL patients while the M set by LCs from multiple myeloma (MM) patients. M LCs were chosen as control since they are overexpressed as the toxic H LCs but they do not lead to toxicity or aggregation. To date, the molecular bases leading to LC proteotoxicity remain to be elucidated. Our data show that low fold stability and high protein flexibility correlate with amyloidogenic LCs, while hydrophobicity, structural rearrangements and nature of the LC dimeric association interface (as observed in seven crystal structures here presented) do not appear to play a significant role in protein aggregation. Additionally, it has been demonstrated that the LCs toxicity in vivo is linked to copper (Cu2+) (Diomede et al., 2017a) by increasing the radical oxygen species (ROS) production. We aimed our studied to clarify Cu2+ LCs interaction. Moreover, we wanted to assess whether Cu2+ is able to alter the biophysical properties of the native state to more aggregation prone states. Our findings reveal that H LCs interacts with Cu2+ with a higher affinity than M LCs and that His residues may be involved in Cu2+ binding. Indeed the affinity decreases in presence of protonated His residues. Moreover, data suggest that the interaction with Cu2+ increases the molecular flexibility and decreases the fold stability. These observations suggests that protein aggregation cannot be evaluated through one single parameters but by the co-action of several biophysical traits. Moreover, our results suggest that the presence of Cu2+ can alter the native LCs properties leading to a higher toxicity in vivo.

EXPLORING THE MOLECULAR AND BIOPHYSICAL MECHANISMS OF PROTEOTOXIC IMMUNOGLOBULIN LIGHT CHAINS IN AL AMYLOIDOSIS / L. Oberti ; scientific tutor: S. Ricagno. - : . DIPARTIMENTO DI BIOSCIENZE, 2019 Mar 15. ((31. ciclo, Anno Accademico 2018. [10.13130/oberti-luca_phd2019-03-15].

EXPLORING THE MOLECULAR AND BIOPHYSICAL MECHANISMS OF PROTEOTOXIC IMMUNOGLOBULIN LIGHT CHAINS IN AL AMYLOIDOSIS

L. Oberti
2019-03-15

Abstract

Herein, immunoglobulin light chains (LCs) native state was studied in the context of the pathology known as light chain amyloidosis (AL). This pathology is characterized by LCs overexpression, which leads to toxicity and aggregation into amyloid fibrils in target organs, with heart being the most affected one. Due to genetic rearrangement and somatic hypermutation, virtually, each AL patient presents a different amyloidogenic LC (Merlini, 2017). Because of such complexity, the fine molecular determinants of LC aggregation propensity and proteotoxicity are, to date, unclear; significantly, their decoding requires investigating large sets of cases. This project is aimed to unravel the molecular determinants linked with LCs toxicity. First, we screened several independent biophysical and structural properties of the LCs native state. In particular, we considered hydrophobicity, fold stability, flexibility and 3D structure. Our experimental approach considered two LCs sets called ‘H’ and ‘M’. The H set is composed of eight LCs from AL patients while the M set by LCs from multiple myeloma (MM) patients. M LCs were chosen as control since they are overexpressed as the toxic H LCs but they do not lead to toxicity or aggregation. To date, the molecular bases leading to LC proteotoxicity remain to be elucidated. Our data show that low fold stability and high protein flexibility correlate with amyloidogenic LCs, while hydrophobicity, structural rearrangements and nature of the LC dimeric association interface (as observed in seven crystal structures here presented) do not appear to play a significant role in protein aggregation. Additionally, it has been demonstrated that the LCs toxicity in vivo is linked to copper (Cu2+) (Diomede et al., 2017a) by increasing the radical oxygen species (ROS) production. We aimed our studied to clarify Cu2+ LCs interaction. Moreover, we wanted to assess whether Cu2+ is able to alter the biophysical properties of the native state to more aggregation prone states. Our findings reveal that H LCs interacts with Cu2+ with a higher affinity than M LCs and that His residues may be involved in Cu2+ binding. Indeed the affinity decreases in presence of protonated His residues. Moreover, data suggest that the interaction with Cu2+ increases the molecular flexibility and decreases the fold stability. These observations suggests that protein aggregation cannot be evaluated through one single parameters but by the co-action of several biophysical traits. Moreover, our results suggest that the presence of Cu2+ can alter the native LCs properties leading to a higher toxicity in vivo.
RICAGNO, STEFANO
light chain amyloidosis; protein aggregation; amyloids; biophysics
Settore BIO/10 - Biochimica
EXPLORING THE MOLECULAR AND BIOPHYSICAL MECHANISMS OF PROTEOTOXIC IMMUNOGLOBULIN LIGHT CHAINS IN AL AMYLOIDOSIS / L. Oberti ; scientific tutor: S. Ricagno. - : . DIPARTIMENTO DI BIOSCIENZE, 2019 Mar 15. ((31. ciclo, Anno Accademico 2018. [10.13130/oberti-luca_phd2019-03-15].
Doctoral Thesis
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Descrizione: PhD thesis Luca Oberti
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/2434/630665
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