posted on 2019-10-10, 14:47authored byAlbert Godoy-Hernandez, Daniel J. Tate, Duncan G. G. McMillan
Type-II NADH:quinone
oxidoreductases (NDH-2s) are an important
element of microbial pathogen electron transport chains and an attractive
drug target. Despite being widely studied, its mechanism and catalysis
are still poorly understood in a hydrophobic membrane environment.
A recent report for the Escherichia coli NDH-2 showed
NADH oxidation in a solution-based assay but apparently showed the
reverse reaction in electrochemical studies, calling into question
the validity of the electrochemical approach. Here we report electrochemical
catalysis in the well-studied NDH-2 from Caldalkalibacillus
thermarum (CthNDH-2). In agreement with
previous reports, we demonstrated CthNDH-2 NADH oxidation
in a solution assay and electrochemical assays revealed a system artifact
in the absence of quinone that was absent in a membrane system. However,
in the presence of either immobilized quinone or mobile quinone in
a membrane, NADH oxidation was observed as in solution-phase assays.
This conclusively establishes surface-based electrochemistry as a
viable approach for interrogating electron transfer chain drug targets.