Mimochrome, a metalloporphyrin‐based catalytic Swiss knife†

Leone, Linda; Chino, Marco; Nastri, Flavia; Maglio, Ornella; Pavone, Vincenzo; Lombardi, Angela

doi:10.1002/bab.1985

Cited by 29 publications

(36 citation statements)

References 307 publications

(345 reference statements)

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“…Although OPs are meant to eradicate specific pests, their high application has led to their accumulation in the environment, contaminating water resources, soils, and foods (vegetables, fruits) and affecting the health of non-target organisms, including biota and human beings [4,5]. A few simple catalysts are emerging for environmental remediation [6][7][8][9]. However, their application is still far to come, being urged to identify contaminated areas and feedstocks.…”

Section: Introductionmentioning

confidence: 99%

A 3D printable adapter for solid-state fluorescence measurements: the case of an immobilized enzymatic bioreceptor for organophosphate pesticides detection

et al. 2022

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The widespread use of pesticides in the last decades and their accumulation into the environment gave rise to major environmental and human health concerns. To address this topic, the scientific community pointed out the need to develop methodologies to detect and measure the presence of pesticides in different matrices. Biosensors have been recently explored as fast, easy, and sensitive methods for direct organophosphate pesticides monitoring. Thus, the present work aimed at designing and testing a 3D printed adapter useful on different equipment, and a membrane support to immobilize the esterase-2 from Alicyclobacillus acidocaldarius (EST2) bioreceptor. The latter is labelled with the IAEDANS, a bright fluorescent probe. EST2 was selected since it shows a high specificity toward paraoxon. Our results showed good stability and replicability, with an increasing linear fluorescent intensity recorded from 15 to 150 pmol of labelled EST2. Linearity of data was also observed when using the immobilized labelled EST2 to detect increasing amounts of paraoxon, with a limit of detection (LOD) of 0.09 pmol. This LOD value reveals the high sensitivity of our membrane support when mounted on the 3D adapter, comparable to modern methods using robotic workstations. Notably, the use of an independent support significantly simplified the manipulation of the membrane during experimental procedures and enabled it to match the specificities of different systems. In sum, this work emphasizes the advantages of using 3D printed accessories adapted to respond to the newest research needs. Graphical abstract

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Section: Introductionmentioning

confidence: 99%

A 3D printable adapter for solid-state fluorescence measurements: the case of an immobilized enzymatic bioreceptor for organophosphate pesticides detection

et al. 2022

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“…Indeed, other artificial heme-enzyme systems have been reported to have increased activity in the presence of TFE. 46 , 47 However, we do not suggest that co-solvent variation is a generalizable tool to affect the flexibility/rigidity of all enzymes but illustrates the principle that tuning flexibility can be used to tune activity.…”

Section: Discussionmentioning

confidence: 60%

Rigidifying a De Novo Enzyme Increases Activity and Induces a Negative Activation Heat Capacity

et al. 2021

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Conformational sampling profoundly impacts the overall activity and temperature dependence of enzymes. Peroxidases have emerged as versatile platforms for high-value biocatalysis owing to their broad palette of potential biotransformations. Here, we explore the role of conformational sampling in mediating activity in the de novo peroxidase C45. We demonstrate that 2,2,2-triflouoroethanol (TFE) affects the equilibrium of enzyme conformational states, tending toward a more globally rigid structure. This is correlated with increases in both stability and activity. Notably, these effects are concomitant with the emergence of curvature in the temperature-activity profile, trading off activity gains at ambient temperature with losses at high temperatures. We apply macromolecular rate theory (MMRT) to understand enzyme temperature dependence data. These data point to an increase in protein rigidity associated with a difference in the distribution of protein dynamics between the ground and transition states. We compare the thermodynamics of the de novo enzyme activity to those of a natural peroxidase, horseradish peroxidase. We find that the native enzyme resembles the rigidified de novo enzyme in terms of the thermodynamics of enzyme catalysis and the putative distribution of protein dynamics between the ground and transition states. The addition of TFE apparently causes C45 to behave more like the natural enzyme. Our data suggest robust, generic strategies for improving biocatalytic activity by manipulating protein rigidity; for functional de novo protein catalysts in particular, this can provide more enzyme-like catalysts without further rational engineering, computational redesign, or directed evolution.

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“…Thus, the same products can be obtained by incubation of one apoprotein with different small molecule complexes, [5] & [6] respectively. The differences between Hybrids 1 & 3 and Hybrids 2 & 4 must result from different topologies around the metal‐protein interactions, possibly different chirality at the metal centre [70, 71] . The higher melting point together with the observation that more folded proteins tend to produce ion series of lower charge state distribution in the mass spectrometer indicates that the protein scaffold has adopted a fold with more extensive intramolecular bonding in Hybrids 1 & 3 than in Hybrids 2 & 4 [72] …”

Section: Resultsmentioning

confidence: 99%

Controlled Ligand Exchange Between Ruthenium Organometallic Cofactor Precursors and a Naïve Protein Scaffold Generates Artificial Metalloenzymes Catalysing Transfer Hydrogenation

et al. 2021

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Many natural metalloenzymes assemble from proteins and biosynthesised complexes, generating potent catalysts by changing metal coordination. Here we adopt the same strategy to generate artificial metalloenzymes (ArMs) using ligand exchange to unmask catalytic activity. By systematically testing RuII(η6‐arene)(bipyridine) complexes designed to facilitate the displacement of functionalised bipyridines, we develop a fast and robust procedure for generating new enzymes via ligand exchange in a protein that has not evolved to bind such a complex. The resulting metal cofactors form peptidic coordination bonds but also retain a non‐biological ligand. Tandem mass spectrometry and 19F NMR spectroscopy were used to characterise the organometallic cofactors and identify the protein‐derived ligands. By introduction of ruthenium cofactors into a 4‐helical bundle, transfer hydrogenation catalysts were generated that displayed a 35‐fold rate increase when compared to the respective small molecule reaction in solution.

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Mimochrome, a metalloporphyrin‐based catalytic Swiss knife†

Cited by 29 publications

References 307 publications

A 3D printable adapter for solid-state fluorescence measurements: the case of an immobilized enzymatic bioreceptor for organophosphate pesticides detection

A 3D printable adapter for solid-state fluorescence measurements: the case of an immobilized enzymatic bioreceptor for organophosphate pesticides detection

Rigidifying a De Novo Enzyme Increases Activity and Induces a Negative Activation Heat Capacity

Controlled Ligand Exchange Between Ruthenium Organometallic Cofactor Precursors and a Naïve Protein Scaffold Generates Artificial Metalloenzymes Catalysing Transfer Hydrogenation

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