Circular Permutated PQQ‐Glucose Dehydrogenase as an Ultrasensitive Electrochemical Biosensor

Abstract

Protein biosensors play an increasingly important role as reporters for research and clinical applications. Here we present an approach for the construction of fully integrated but modular electrochemical biosensors based on the principal component of glucose monitors PQQ‐glucose dehydrogenase (PQQ‐GDH). We designed allosterically regulated circular permutated variants of PQQ‐GDH that show large (>10‐fold) changes in enzymatic activity following intramolecular scaffolding of the newly generated N‐ and C termini by ligand binding domain/ligand complexes. The developed biosensors demonstrated sub‐nanomolar affinities for small molecules and proteins in colorimetric and electrochemical assays. For instance, the concentration of Cyclosporine A could be measured in 1 μL of undiluted blood with the same accuracy as the leading diagnostic technique that uses 50 times more sample. We further used this biosensor to construct highly porous gold bioelectrodes capable of robustly detecting concentrations of Cyclosporine A as low as 20 pM and retained functionality in samples containing at least 60 % human serum.

Creation of a Self-Powered, Real-Time Sensor for Therapeutics in Blood: from Protein Engineering to Electronic Integration

“…We hypothesized that the 4-HT speci c conformational changes in the LBD would modulate the redox activity of GDH, leading to changes in the electrical signals. In previous studies, PQQ-GDH was allosterically engineered to sense α-amylase 14 and cyclosporine A 15 by inserting corresponding recognition domains with calmodulin as the conformational switch. However, invasively disrupting or splitting the structure of PQQ-GDH heavily impeded enzyme turnover for glucose oxidation and slowed signal generation (~ 30 min).…”

Machine learning-assisted flexible wearable device for tyrosine detection

“…[19][20][21] These biosensors include electrochemical biosensors, uorescence-based biosensors, colorimetric biosensors, localized surface plasmon resonance (LSPR) and surface-enhanced Raman scattering (SERS) biosensors. [22][23][24][25] Among them, electrochemical and SERS biosensors are the most popular. For example, a label-free electrochemical immunosensor was prepared using a N-GQD/ GNR composite-modied electrode to detect carbohydrate antigen 15-3 (CA15-3) biomarkers.…”