Kinetic Model for the Heterogeneous Biocatalytic Reactions Using Tethered Cofactors.

Understanding the mechanism of interfacial enzyme kinetics is critical to the development of synthetic biological systems for the production of value-added chemicals. Here, the interfacial kinetics of the catalysis of β-nicotinamide adenine dinucleotide (NAD)-dependent enzymes acting on NAD tethered to the surface of silica nanoparticles (SiNPs) has been investigated using two complementary and supporting kinetic approaches: enzyme excess and reactant (NAD) excess. Kinetic models developed for these two approaches characterize several critical reaction steps including reversible enzyme adsorption, complexation, decomplexation, and catalysis of the surface-bound enzyme/NAD complex. The analysis reveals a concentrating effect resulting in a very high local concentration of enzyme and cofactor on the particle surface, in which the enzyme is saturated by surface-bound NAD, facilitating a rate enhancement of enzyme/NAD complexation and catalysis. This resulted in high enzyme efficiency within the tethered NAD system compared to that of the free enzyme/NAD system, which increases with decreasing enzyme concentration. The role of enzyme adsorption onto solid substrates with a tethered catalyst (such as NAD) has potential for creating highly efficient flow biocatalytic systems.