Evaluating cadmium-free quantum dots along with mixed nanoparticle clusters as scaffolds for multienzymatic glycolytic channeling

Abstract Allowing coupled enzymes to crosslink with nanoparticles (NPs) into nanoclusters has been shown to facilitate them engaging in the most efficient form of multienzymatic catalysis, namely that of intermediary channeling. Utilizing a previously validated nanoparticle-scaffolded seven enzyme cascade from glycolysis that processes glucose into 3-phosphoglycerate, we begin by confirming that non-cadmium containing ZnSe/ZnS core/shell quantum dots (QDs) made from non-toxic and earth abundant materials can replace Cd-containing QDs as a scaffolding material in the multienzyme clusters while still providing access to improved channeling activity. We then investigate the role of enzyme assembly order within mixed NP systems that consist of both spherical QDs and rectangular 2-dimensional nanoplatelets (NPLs). Along with physicochemical confirmation of enzyme assembly to the QDs and enzyme-induced cluster formation, the rate of overall catalytic flux for each of the systems was monitored under different assembly conditions. The results reveal that adjusting relative NP concentration normalized to surface area, enzyme assembly order, and choice of initial material in any mixed NP clustered configuration are critical to attaining further improvements in catalytic flux via channeling. The potential ramifications of these observations in the context of assembling designer biosynthetic cascades that use bulk feedstock materials derived from agriculture to create new and useful products are then discussed. Graphical Abstract Schematic of a self-assembled mixed QD-NPL-enzyme system engaged in 7-enzyme sequential substrate channeling.