An Artificial Hemoprotein with Inducible Peroxidase‐ and Monooxygenase‐Like Activities

Abstract

An ovel inducible artificial metalloenzyme obtained by covalent attachment of am anganese(III)-tetraphenylporphyrin (MnTPP) to the artificial bidomain repeat protein, (A3A3')Y26C,i sr eported. The protein is part of the aRep family. The biohybrid was fully characterized by MALDI-ToF mass spectrometry,c irculard ichroism and UV/Vis spectroscopies. The peroxidase and monooxygenase activities were evaluated on the originala nd modified scaffolds including those that have a) an additional imidazole, b) a specific aRep bA3-2 that is known to induce the opening of the (A3A3')i nterdomain region and c) ad erivative of the aRep bA3-2 inducer extended with aH is 6-Tag (His 6-bA3-2). Catalytic profiles are highly dependento nt he presence of co-catalysts with the best activity obtained with His 6-bA3-2. The entire mechanism wasr ationalized by an integrative molecular modeling study that includes protein-ligand docking and large-scale molecular dynamics. This constitutes the first example of an entirely artificial metalloenzyme with inducible peroxidase and monooxygenase activities, reminiscent of allostericr egulation of natural enzymatic pathways.

Allosteric cooperation in a de novo-designed two-domain protein

“…Therefore, the addition of a domain to an existing protein expands, alters, or modulates its functionality ( 5 ). Protein engineers have been inspired by this modularity to generate artificial multidomain proteins with improved properties or to create nanostuctured and sensor devices ( 6 – 10 ). Toward this end, different methodologies have been developed to fuse the different domains by: 1) introducing designed or naturally occurring peptide linkers ( 11 ); 2) superimposing and fusing one or two turns of terminal alpha helices of connecting helical proteins ( 12 , 13 ); and 3) computationally designing new structural elements to interface the different domains in a fragment based approach ( 14 ).…”

Integrated experimental/computational approaches to characterize the systems formed by vanadium with proteins and enzymes

“…8 In a similar way, the interaction of metallic species with macromolecular hosts has a major application in biotechnology from the development of metallopeptides to the de novo design and redesign of enzymes. 9,10 In the latter cases, the catalytic activity and selectivity of the process can be handled by introducing appropriate interactions at the cofactor-biomolecule interface, e.g. favouring those that stabilize the cofactor in a specific region or with a determined orientation for the reaction to take place.…”

“…Recent applications are the binding elucidation of Pt-and Ru-based metallodrugs in proteins as well as the Cu-and Mn-based cofactors in artificial metalloenzymes. 9,[174][175][176] In the field of metallodrug design, the binding sites of antiarthritic Auranofin and other cytotoxic gold compounds under experimentation share the same structural motif and the [Au I (PEt 3 )] + moiety is released in aqueous solution. [177][178][179] HSA is the protein candidate for the transport of Au(I) species in humans, even though no X-ray determinations are reported for the adducts between the Au I (PEt 3 ) + moiety and HSA.…”

Photocatalytic Hydrogen Production and Carbon Dioxide Reduction Catalyzed by an Artificial Cobalt Hemoprotein

“…We have thus previously reported a proof of concept by creating artificial Diels–Alderases by covalent attachment of phenanthroline– or terpyridine–Cu(II) complexes in a specific αRep single-chain asymmetric bidomain variant named (A3A3′)Y26C [ 20 ]. This protein has been shown to adopt a closed bivalve shell-like conformation and molecular modeling identified that this cavity is large enough for the anchoring of even larger metal complexes such as meso -tetraphenylporphyrin [ 21 ]. Thus, from an enzyme designer’s point of view, the artificial αRep proteins bring together the properties required for the development of artificial metalloenzymes dedicated to H 2 production.…”