Real-time monitoring of voltage shift based on enzymatically released pyrophosphate using phenylboronic acid-immobilized gate field-effect transistor

Pyrophosphate (PPi) is ubiquitous in living cells and is often produced by enzymatic reactions, e.g., DNA synthesis by DNA polymerase. We have developed a novel detection system for the voltage shift associated with the change in PPi concentration resulting from an enzymatic reaction using a phenylboronic acid (PBA)-coated gate field-effect transistor (FET), since PBA coating is effective for detecting ion accumulation associated with PPi production from enzymatic reactions. To detect enzymatic reactions more efficiently, we employed the enzyme-electrode conjugation method using specific peptide sequences, which are spontaneously tethered to a gold substrate. The combination of the enzyme-electrode conjugation method with the charge detection using the PBA-coated FET enables the effective detection of enzymatic reactions.