| Recent developments in applications of electronic coupling of redox enzymes to electrodes, an area named bioelectrocatalysis, have led to the conception of innovative biosensors and systems for chiral synthesis. In this work, we investigated both electron-transfer mediators and direct electron exchange for the electronic coupling of electrode and enzyme active sites.; In developing a rapid, accurate, multi-enzyme creatine-creatinine sensor for measurements in clinical samples, the indirect coupling of sarcosine dehydrogenase to an electrode was demonstrated with a variety of electron transfer mediators. The best mediator for this sensor system in terms of enzyme kinetics and current output was 2,6-dichlorophenol indophenol (2,6-DCPIP). The creatine sensor exhibited an apparent Km of 361 ± 82 μM, while the creatinine sensor displayed an apparent Km of 348 ± 90 μM; thus, both sensors displayed a linear response over the 40–150 μM range in normal humans. Experimental and theoretical estimates of mass transport and kinetic parameters revealed that the current response of the overall system probably was limited by diffusion of 2,6-DCPIP mediator in the immobilized enzyme layer.; Direct or unmediated electron transfer from enzymes was effectively demonstrated for D-fructose sensor based on hydrophobic immobilization of fructose dehydrogenase (FDH) in a self-assembled monolayer on gold. Cyclic voltammograms confirmed direct electron transfer between FDH and a gold foil electrode, but the catalytic anodic peaks gradually disappeared over extensive potential sweeping. Constant applied potential experiments further confirmed direct electron transfer, and the sensor exhibited rapid response times of approximately 1.91 seconds. However, the current was saturated at relatively low concentrations, possibly owing to inefficiency of electron transfer between the enzyme redox center and the gold electrode.; Successful mediated coupling of L(+)-lactate dehydrogenase (LDH), purified from A. fulgidus, to gold electrodes was achieved to investigate the kinetic properties of this enzyme for potential applications in bioelectrosynthesis. LDH from A. fulgidus showed high substrate selectivity, because it yielded appreciable activity only toward L-(+)-lactic acid and 2-hydroxy butyric acid. This demonstration of mediated electronic coupling illustrates the potential of this system for bioelectrosynthesis applications. |
