McDonough Rowan, Williams Charlotte C, Hartley Carol J, French Nigel, Scott Colin, Lewis David A
Institute for Nanoscale Science and Technology, School of Chemical and Physical Sciences, Flinders University, Bedford Park, SA 5042, Australia.
CSIRO Manufacturing, Clayton, Melbourne, VIC 3168, Australia.
Langmuir. 2024 Apr 2;40(13):6685-6693. doi: 10.1021/acs.langmuir.3c02958. Epub 2024 Mar 25.
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.
了解界面酶动力学机制对于开发用于生产增值化学品的合成生物系统至关重要。在此，我们使用两种互补且相互支持的动力学方法，即酶过量和反应物（NAD）过量，研究了作用于连接在二氧化硅纳米颗粒（SiNPs）表面的NAD的β-烟酰胺腺嘌呤二核苷酸（NAD）依赖性酶催化的界面动力学。为这两种方法建立的动力学模型表征了几个关键反应步骤，包括可逆酶吸附、络合、解络合以及表面结合的酶/NAD复合物的催化。分析揭示了一种浓缩效应，导致颗粒表面酶和辅因子的局部浓度非常高，其中酶被表面结合的NAD饱和，促进了酶/NAD络合和催化的速率提高。与游离酶/NAD系统相比，这使得连接NAD系统内的酶效率很高，且随着酶浓度降低而增加。酶吸附到带有连接催化剂（如NAD）的固体底物上的作用，对于创建高效流动生物催化系统具有潜力。
