TY - JOUR
T1 - Revealing the Membrane-Bound Catalytic Oxidation of NADH by the Drug Target Type-II NADH Dehydrogenase
AU - Godoy-Hernandez, Albert
AU - Tate, Daniel J.
AU - McMillan, Duncan G.G.
PY - 2019
Y1 - 2019
N2 - 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.
AB - 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.
UR - http://www.scopus.com/inward/record.url?scp=85073248507&partnerID=8YFLogxK
U2 - 10.1021/acs.biochem.9b00752
DO - 10.1021/acs.biochem.9b00752
M3 - Article
C2 - 31592658
SN - 0006-2960
VL - 58
SP - 4272
EP - 4275
JO - Biochemistry
JF - Biochemistry
IS - 42
ER -