Real-time structural signatures of protein allostery by electrically- and mechanically-coupled sensing

Accurate detection and identification of conformational evolutions are essential to describe the micro-structural mechanisms of protein allostery. Here we establish an approach by integrating nanopore sensing technique with atomic force microscopy, based on molecular dynamics simulations. A theoretical method is further developed to real-time estimate the low-resolution structures of those evolved conformations. Tests for the forced allostery of αXβ2 integrin, a typical protein with multiple functional states, demonstrate that the spheroidal shape identifiers of those intermediate conformers present sufficient sensitivity to resolve two allosteric patterns of αXβ2 from bent-down to stand-up states under the nanopore confinement. The strong steric confinement from the nanopore restricts the geometric displacements of key domains in conformational extension dynamics, thereby increasing the intramolecular interaction energy barrier to be overcome for standing. Prolongation of the neck-shoulder region of αXβ2 and opening at the thigh-calf joint form a competitive compensation for the stretched height, resulting in two tendentious allosteric patterns under various confined diameters and steered speeds. Thus, this study proposes a new method to real-time visualize the conformational dynamics during protein allostery, favoring to depict structural characteristics for the conformations of those transient states and evaluate the impacts of the nanopore confinement on protein allostery. [Display omitted] •A MD approach of combining nanopore and AFM to probe protein conformational dynamics.•Shape signatures of evolutive conformations resolved by SMD-spheroid approximation.•Nanopore confinement enhanced the energy barrier of structural extension of αXβ2.•Intramolecular domains were competitive in compensating for the stretched height.•Two allosteric patterns under varied confinements and steered speeds were formed.