Unraveling antibody-induced mechanical stability of antigen: Insights from single-molecule studies.

Abstract

Antigen-antibody interaction, as a prominent ligand-receptor reaction, plays a crucial role in immunological responses. Notably, antigens can contain multiple ligand binding sites that define their intermolecular interactions more intricately and thereby make them context-dependent. Here, we have investigated the binding-induced effect of the largest antibody isotype, IgM, on protein L mechanical stability using single-molecule magnetic tweezers. Our results showed that IgM elevates the protein L mechanical stability by increasing its unfolding time. Interestingly, we were able to resolve distinct IgM-bound states of protein L by characterizing their unfolding dwell time: while the IgM-unbound state has the lowest dwell time, it increases with the IgM concentration via binding to either one or both of its binding sites, reconciling the IgM-induced protein L mechanical stability. To delve into the plausible mechanism of such intricate phenomena, we performed steered a molecular dynamic simulation of protein L and determined its unfolding rupture force at those multiple IgM-bound states, their corresponding molecular insights, and interaction gymnastics through binding interfaces. Additionally, we unraveled the mechanical response of these binding interfaces to be different; and during dimer IgM complex formation, these binding interfaces synergistically increase the mechanical stability of the complex. This provides the underlying principles of IgM-induced protein L stability under mechanical constraints. Overall, this study provides an in-depth understanding of a generic mechanism of antibody-induced mechanical stability of antigenic substrate under physiological sheer stress.