Signal-on CoA-dependent electrochemical biosensor for highly sensitive and label-free detection of Citrate synthase activity.

We report here a label-free and sensitive electrochemical method for probing Citrate synthase (CS) activity based on detailed investigations into the nucleic acid-mimicking coordination polymer (CP) formed from the coenzyme A (CoA)-Ag(I) repeat units. Our biosensing approach provides an especial and significant detection mechanism: CS can catalyze the essential condensation reaction between acetyl-coenzyme A (Ac-CoA) and oxaloacetate (OAA) to form citrate and CoA; then, in the presence of Ag(I), CoA-Ag(I) CP can be in situ formed because of the strong complexation ability of thiol groups of CoA toward Ag(I). The generated CoA-Ag(I) CP attaches to graphene-modified glassy carbon electrode surface by multiple adenine bases deriving from CoA and acting as the side groups along the polymeric backbone, which displays efficient HO-electrocatalyzing activity. More importantly, by using the formed polymer as signal output, the process is implemented to quantitatively analyze the activity of CS. Under the optimal conditions, CS with a detection limit as low as 0.00165U/µL could be sensitively probed with a wide linear range from 0.0033 to 0.264U/µL. Furthermore, with the character of label-free detection, high sensitivity and excellent selectivity, this strategy offers a convenient and specific method for CS activity detection and relevant inhibitors screening, which holds a promising potential in the practical application of CS-based biochemical research, disease diagnosis and drug discovery.