Deep learning-based classification of peptide analytes from single-channel nanopore translocation events

Rapid and accurate detection of peptide biomarkers using nanopore biosensors is critical for disease diagnosis and other biomedical applications. Processing large, complex single-channel translocation data streams poses a significant challenge for peptide analyte classification. Here, we present a supervised deep learning data processing pipeline for peptide classification from translocation events. The first stage employs a convolutional and recurrent neural network, adapted from the Deep-Channel multi-channel classifier, to accurately classify raw current recordings into discrete conductance states, including partially blocked sub-conductance intermediates. The second stage, peptide classification, utilizes a novel branched input network with a temporal convolutional network for processing translocation event conductance state sequences and a dense network for incorporating computed event-level and global kinetic features. Using idealized simulated multi-state translocation data for seven peptides, we demonstrate high classification accuracy (0.9998 (±0.0006)) when global features are included alongside event-level features. For classifying mixture samples, where only event-level features are applicable, performance shows more modest accuracy (0.70 (±0.01)). Peptide mixture predictions showed reasonable accuracy (MAE 0.045-0.161), although misclassification resulted in false positives. Event stochasticity and the fact that some peptides possessed similar kinetic parameters posed challenging for event-level prediction. However, vote aggregation from translocation event streams achieves perfect 100% accuracy, when predicting pure peptide samples. This proof-of-concept study demonstrates a robust deep learning framework for nanopore peptide classification using simulated data, laying the groundwork for classifying peptides from complex mixtures using real experimental data with the anthrax toxin protective antigen nanopore.