An Emerging Nanozyme Class for à la carte Enzymatic‐Like Activities based on Protein‐Metal Nanocluster Hybrids

Abstract

In this study, the goal is to fabricate robust and highly efficient peroxidase‐like nanozymes that can ultimately be assembled into films for their easy reuse in catalytic cycles. Nanozymes are designed by mimicking the strategy adopted by metalloproteins to accommodate metal cofactors within their protein structure. The engineered consensus tetratricopeptide repeat (CTPR) protein module is selected as the scaffold to guide the growth and the stabilization of a library of in situ synthesized metal nanoclusters. A deep investigation of the interplay between the composition and function of the nanozymes reveals the impact of the protein templates and nanocluster composition on the peroxidase‐like activity of the hybrids. Moreover, among a total of 24 hybrids, a top‐performing nanozyme results from the growth of Au/Pt bimetallic nanoclusters on a CTPR protein with engineered histidine coordination sites. These nanozymes exhibit improved thermostability and resistance to hydrogen peroxide compared to natural peroxidases like horseradish peroxidase. Finally, it shows the easy fabrication of nanozyme composite films guided throughout the intrinsic self‐assembling properties of the CTPR scaffold. These heterogeneous solid materials are reused in several reaction cycles without significant loss of the catalytic performance, proving these protein‐templated nanozymes as an advantageous alternative to natural enzymes.

In vitro Production of Hemin‐Based Artificial Metalloenzymes

“…(Figure S2). The assembly procedure is performed at pH 10, a condition under which the protein is known to be in a partially extended conformation [24] to reduce steric impediments and foster the incorporation of the metalorganic molecule within the cleft. After the assembly, the native configuration of the protein was restored at pH 7, as evidenced by its circular dichroism spectrum (Figure S3).…”

“…Specifically, the designed protein acted as metal‐free biocatalysts for (3+2) cycloadditions [23] . Our group also demonstrated the capability of CTPR proteins to act as scaffolds for coordinating metal nanoclusters, achieving tunable, highly catalytic, stable, and reusable nanozymes, proving them as an advantageous alternative to natural enzymes [24] . This study presents a new approach: the development of an engineered CTPR3 variant with the capacity to coordinate a hemin porphyrin, simulating the structural conformation of HRP.…”

The roles of templates consisting of amino acids in the synthesis and application of gold nanoclusters

“…Using engineered CTPR protein with histidine coordination sites as a ligand to form Au/Pt bimetallic NCs, the highest activity was obtained. 117 BSA–AuNCs were used for the detection of dopamine (DA) and uranyl with decrease of POD-like activity of BSA–AuNCs (Fig. 7A), respectively.…”

Enzyme-mimic catalytic activities and biomedical applications of noble metal nanoclusters

“…, Au NCs or Pt NCs protected by CTPR). 41 These Au/Pt NCs can be reused in multiple reaction cycles without significant loss of catalytic activity.…”