De Novo Sequencing of Antibody Light Chain Proteoforms from Patients with Multiple Myeloma

In multiple myeloma diseases, monoclonal immunoglobulin light chains (LCs) are abundantly produced, with, as a consequence in some cases, the formation of deposits affecting various organs, such as the kidney, while in other cases remaining soluble up to concentrations of several g·L–1 in plasma. Th...

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Bibliographic Details
Published in:Analytical chemistry (Washington) 2021-08, Vol.93 (30), p.10627-10634
Main Authors: Dupré, Mathieu, Duchateau, Magalie, Sternke-Hoffmann, Rebecca, Boquoi, Amelie, Malosse, Christian, Fenk, Roland, Haas, Rainer, Buell, Alexander K, Rey, Martial, Chamot-Rooke, Julia
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Amino acids
Analytical chemistry
Antibodies
Chains
Chemical Sciences
Chemistry
Humans
Immunoglobulin Light Chains
Immunology
Life Sciences
Light chains
Multiple Myeloma
Organs
Patients
Peptides
Proteomics
Sequence Analysis, Protein
Trypsin
Workflow
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Summary:In multiple myeloma diseases, monoclonal immunoglobulin light chains (LCs) are abundantly produced, with, as a consequence in some cases, the formation of deposits affecting various organs, such as the kidney, while in other cases remaining soluble up to concentrations of several g·L–1 in plasma. The exact factors crucial for the solubility of LCs are poorly understood, but it can be hypothesized that their amino acid sequence plays an important role. Determining the precise sequences of patient-derived LCs is therefore highly desirable. We establish here a novel de novo sequencing workflow for patient-derived LCs, based on the combination of bottom-up and top-down proteomics without database search. PEAKS is used for the de novo sequencing of peptides that are further assembled into full length LC sequences using ALPS. Top-down proteomics provides the molecular masses of proteoforms and allows the exact determination of the amino acid sequence including all posttranslational modifications. This pipeline is then used for the complete de novo sequencing of LCs extracted from the urine of 10 patients with multiple myeloma. We show that for the bottom-up part, digestions with trypsin and Nepenthes digestive fluid are sufficient to produce overlapping peptides able to generate the best sequence candidates. Top-down proteomics is absolutely required to achieve 100% final sequence coverage and characterize clinical samples containing several LCs. Our work highlights an unexpected range of modifications.
ISSN:0003-2700
1520-6882
DOI:10.1021/acs.analchem.1c01955