Characterization and modulation of the unimolecular conformation of integrins with nanopore sensors

Integrins are a large family of heterodimeric mammalian transmembrane receptors that connect extracellular matrix (ECM) and the cytoskeleton. Integrin-mediated adhesion governs the cellular adhesion, mobility, and cell mechanobiology. Investigations on different integrins in cellular and the conformation change regulated by divalent metal ions and oligopeptides have been documented, they mainly adopted the cellular assays or surface manipulation to achieve the information of ensemble integrins with proteins and small molecules. This work presents the nanopore approach for the characterization of unimolecular integrin via resistive pulse sensing. The orientation selection in distinct nanopore in size and pH condition discloses the tiny disparity of the conformation and the local surface charge distribution. The divalent metal ions could efficiently modulate the conformation alteration of integrin, in which Ca2+ and Mn2+ could activate integrin alpha L beta 2 to take larger physical dimension than Mg2+, and Ca2+ is superior to Mn2+ in the activation when integrin alpha L beta 2 is treated in the mixed metal ions. The oligopeptide cRGD (cyclic Arg-Gly-Asp) and its derivatives cRGDfk and cRGDyk show specific binding with integrin alpha V beta 3 and the complex of alpha V beta 3-cRGD exhibits largest geometry upon Ca2+-mediation with characteristic nanopore translocation feature. This work provides a platform for unimolecular integrin sensing and the modulation of the local conformation transition regulated by divalent metal ions and oligopeptides, which will be valuable for the representation of integrin-related cellular behavior and bio-function.