Native globular ferritin nanopore sensor.

High-resolution nanopore analysis technology relies on the design of novel transmembrane protein platforms. Traditional barrel-shaped protein channels are preferred for constructing nanopore sensors, which may miss protein candidates in non-barrel structures. Here, we demonstrate the globular ferritin displays excellent membrane-insertion capacity and stable transmembrane ionic current owing to its hydrophobic four-fold channels and hydrophilic three-fold channels. The ionic current rectification and voltage-gating characteristics are discovered in single-ferritin ionic current measurement. Notably, the ferritin is used as a nanopore sensor, by which we achieve the high resolution discrimination of L-cysteine, L-homocysteine, and cysteine-containing dipeptides with the assistance of equivalent Cu(2+). The mechanistic studies by multiple controlled experiments and quantum mechanics/all-atom/coarse-grained multiscale MD simulations reveal that analytes are synergistically captured by His114, Cys126, and Glu130 within C3 channel, causing the current blockage signals. The promising ferritin nanopore sensor provides a guide to discovering new protein nanopores without shape restrictions.