Nanocaged enzymes with enhanced catalytic activity and increased stability against protease digestion

Abstract

Cells routinely compartmentalize enzymes for enhanced efficiency of their metabolic pathways. Here we report a general approach to construct DNA nanocaged enzymes for enhancing catalytic activity and stability. Nanocaged enzymes are realized by self-assembly into DNA nanocages with well-controlled stoichiometry and architecture that enabled a systematic study of the impact of both encapsulation and proximal polyanionic surfaces on a set of common metabolic enzymes. Activity assays at both bulk and single-molecule levels demonstrate increased substrate turnover numbers for DNA nanocage-encapsulated enzymes. Unexpectedly, we observe a significant inverse correlation between the size of a protein and its activity enhancement. This effect is consistent with a model wherein distal polyanionic surfaces of the nanocage enhance the stability of active enzyme conformations through the action of a strongly bound hydration layer. We further show that DNA nanocages protect encapsulated enzymes against proteases, demonstrating their practical utility in functional biomaterials and biotechnology.

DNA Antenna Tile‐Associated Deoxyribozyme Sensor with Improved Sensitivity

“…strategies for overcoming the diffusion limitations or the substrate channelling along the active sites of several enzymes. [14c,19] …”

How to Lengthen the Long-Term Stability of Enzyme Membranes: Trends and Strategies

“…Today, a molecule can be immobilized into a molecular size cavity [2,128]. There have been reports on stability enhancement achieved by controlling the environment at a molecular level [129,130].…”

Nanotechnology and the Unique Role of DNA

“…If a site-specific linkage is required and the POI contains accessible cysteine residues, the coupling order can be reversed, using amino-modified oligonucleotides [86]. The use of heterocrosslinkers is still one of the most largely employed methods for attachment of selected proteins to DNA nanostructures ( Figure 1.6(a)) [28].…”

“…Besides solving design challenges, scientists rapidly succeeded in demonstrating the use of those structures for realistic applications, from the development of addressable molecular pegboards for protein patterning [24][25][26][27] or encapsulation [28][29][30][31][32], to optoelectronic hybrid materials [33] and organic catalysts [34]. Another field in great expansion is coupled to the advancement of single-molecule technologies, enabling for example the precise localization and counting of molecules in spatially distributed samples or the disclosure of anomalous kinetic events occurring on a time scale normally not accessible by standard methods [8,[35][36][37][38][39][40].…”