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

In multiple myeloma diseases, monoclonal immunoglobulin light chains (LC) are abundantly produced, with the consequence in some cases to form deposits affecting various organs, such as kidney, while in other cases to remain soluble up to concentrations of several g.L^-1 in plasma. The exact factors crucial for the solubility of light chains are poorly understood, but it can be hypothesized that their amino acid sequence plays an important role. Determining the precise sequences of patient-derived light chains 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. 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 fluid extract are sufficient to produce overlapping peptides able to generate the best sequence candidates. For the sequencing of intact LC proteoforms, combining activation methods is key to achieve single amino acid resolution.