Real-time monitoring of mass-transport-related enzymatic reaction kinetics in a nanochannel-array reactor.
To understand the fundamentals of enzymatic reactions confined in micro-/nanosystems, the construction of a small enzyme reactor coupled with an integrated real-time detection system for monitoring the kinetic information is a significant challenge. Nano-enzyme array reactors were fabricated by covalently linking enzymes to the inner channels of a porous anodic alumina (PAA) membrane. The mechanical stability of this nanodevice enables us to integrate an electrochemical detector for the real-time monitoring of the formation of the enzyme reaction product by sputtering a thin Pt film on one side of the PAA membrane. Because the enzymatic reaction is confined in a limited nanospace, the mass transport of the substrate would influence the reaction kinetics considerably. Therefore, the oxidation of glucose by dissolved oxygen catalyzed by immobilized glucose oxidase was used as a model to investigate the mass-transport-related enzymatic reaction kinetics in confined nanospaces. The activity and stability of the enzyme immobilized in the nanochannels was enhanced. In this nano-enzyme reactor, the enzymatic reaction was controlled by mass transport if the flux was low. With an increase in the flux (e.g., >50 microL min(-1)), the enzymatic reaction kinetics became the rate-determining step. This change resulted in the decrease in the conversion efficiency of the nano-enzyme reactor and the apparent Michaelis-Menten constant with an increase in substrate flux. This nanodevice integrated with an electrochemical detector could help to understand the fundamentals of enzymatic reactions confined in nanospaces and provide a platform for the design of highly efficient enzyme reactors. In addition, we believe that such nanodevices will find widespread applications in biosensing, drug screening, and biochemical synthesis.
Li
Su-Juan
SJ
Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093, China.
Wang
Chen
C
Wu
Zeng-Qiang
ZQ
Xu
Jing-Juan
JJ
Xia
Xing-Hua
XH
Chen
Hong-Yuan
HY
D001692:Biological Transport; D002384:Catalysis; D004800:Enzymes, Immobilized / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics
D004800:Enzymes, Immobilized