Artificial metalloenzymes

An artificial metalloenzyme (26) was designed by Breslow et al. 24). It was the first example of a complete artificial enzyme, having a substrate binding cyclodextrin cavity and a Ni2+ ion-chelated nucleophilic group for catalysis. Metalloenzyme (26) behaves a real catalyst, exhibiting turnover, and enhances the rate of hydrolysis of p-nitrophenyl acetate more than 103 fold. The catalytic group of 26 is a -Ni2+ complex which itself is active toward the substrate 1, but not toward such a substrate having no metal ion affinity at a low catalyst concentration. It is appearent that the metal ion in 26 activates the oximate anion by chelation, but not the substrate directly as believed in carboxypeptidase. 

Cieus M, Ward TR. Designed evolution of artificial metalloenzymes protein catalysts made to order. Org Biomol Chem 2007 5 1835-1844. 

Figure .17 Artificial metalloenzymes (a) Strategyto incorporate a catalytically active metal fragment within a host protein.

Chin, J., Developing artificial hydrolytic metalloenzymes by a unified mechanistic approach , Acc. Chem. Res., 24, 145-152 (1991). 

In natural processes, metal ions are often in high oxidation states (2 or 3), whereas in chemical systems the metals are in low oxidation states (0 or 1). This fact inverts the role of the metal center, such that it acts as a one-electron sink in a natural system, but as a nucleophile in an artificial ones (see other chapters of this book and the review by Aresta et al. [109]). Nevertheless, important biochemical processes such as the reversible enzymatic hydration of C02, or the formation of metal carbamates, may serve as natural models for many synthetic purposes. Starting from the properties of carbonic anhydrase (a zinc metalloenzyme that performs the activation of C02), Schenk et al. proposed a review [110] of perspectives to build biomimetic chemical catalysts by means of high-level DFT or ah initio calculations for both the gas phase and in the condensed state. The fixation of C02 by Zn(II) complexes to undergo the hydration of C02 (Figure 4.17) the use of Cr, Co, or Zn complexes as catalysts for the coordination-insertion reaction of C02 with epoxides and the theoretical aspects of carbamate synthesis, especially for the formation of Mg2+ and Li+ carbamates, are discussed in the review of Schenk... 

FIGURE 28 Artificial metalloenzymes (A) strategy for incorporating a catalytically active metal fragment into a host protein (Wilson and Whitesides (97)) (B) hydrogenation of alkenes via biotin-(strep)avidin methodology (Wilson and Whitesides (97) and Skander et al. (9S)). (For a color version of this figure, the reader is referred to the Web version of this chapter.)... 

The preparation and characterization of novel man-ganese(III) complexes of various porphyrin and porphyrin-likes macrocycles have continued to attract strong attention especially because of their importance in catalytical oxidation processes through the formation of a Mn(V)0 intermediate (see Section 6) and as model for metalloenzymes. In this line, an artificial enzyme formed through a directed assembly of a molecular square that encapsulated a Mn porphyrin has been prepared and investigated as a catalyst. In contrast to symmetrical binuclear bis(phenoxo) bridged macrocyclic Mn(III)Mn(III) complexes, unsymmetrical ones are rare. A new series of these kinds of carboxylate-free complexes has been described and their redox properties investigated. ... 

Letondor C, Ward TR. Artificial metalloenzymes for enantiose-lective catalysis recent advances. ChemBioChem. 2006 7 1845-1852. 

Abe S, Ueno T, Reddy PAN, Okazaki S, Hikage T, Suzuki A, Yamane T, Nakajima H, Watanabe Y. Design and structure analysis of artificial metalloproteins selective coordination of His64 to copper complexes with square-planar structure in the apo-myoglobin scaffold. Inorg. Chem. 2007 46 5137-5139. Ohashi M, Koshiyama T, Ueno T, Yanase M, Fuji H, Watanabe Y. Preparation of artificial metalloenzymes by insertion of chromium Schiff base complexes into apomyoglobin mutants. Angew. Chem Int. Ed. 2003 42 1005-1008. 

Collot J, Gradinaru J, Humbert N, Skander M, Zocchi A, Ward TR. Artificial metalloenzymes for enantioselective catalysis based on biotin-avidin. J. Am. Chem. Soc. 2003 125 9030-9031. Letondor C, Humbert N, Ward TR. Artificial metalloenzymes based on biotin-avidin technology for the enantioselective reduction of ketones by transfer hydrogenation. I roc. Natl. Acad. Sci. U.S.A. 2005 102 4683-4687. 

Thomas CM, Letondor C, Humbert N, Ward TR. Aqueous oxidation of alcohols catalyzed by artificial metalloenzymes based on the biotin-avidin technology. J. Organomet. Chem. 2005 690 4488 491. 

Material scientists have exploited a range of ferritin superfamily proteins as supramolecular templates to encapsulate nanoparticles and/or as well-defined building blocks for fabrication of higher order assembly. For example, the organometallic Rh(nbd) (nbd = norbomadiene) can be immobilised at specific sites within the apoferritin molecule where it can catalyse the polymerisation of phenylacetylene within the protein shell (Figure 19.10). This is but one example of the quest to develop highly effective artificial metalloenzymes by rational design of metal coordination sites within the ferritin molecule. 

Furthermore, subsequent bimolecular reaction with the second molecule of diiron(II) complex is suppressed in dendrimers, thus allowing the direct observation and spectroscopic characterization of the Fe(II)Fe(III) intermediate (presumably, a super-oxo complex).96 Oxygen-activating metallodendrimers open a new way toward designing artificial analogues of metalloenzymes. 

The examples in this section have been chosen to provide an in-depth presentation showing how RSSF currently has been applied to the study of biological systems. These applications include the study of isotope effects on enzyme-catalyzed reactions, the investigation of substrate-metal ion interactions in metalloenzymes, the search for and identification of covalent intermediates in enzyme-catalyzed processes, the analysis of the effects of site-directed mutations on enzyme catalytic mechanism, and the exploitation of natural and artificial chromophores as probes of allosteric processes. 

Information on catalytic roles of metal ions acting as Lewis acid catalysts in metallopeptidases provides insights into designing artificial metallopeptidases. Here, the mechanistic studies on carboxypeptidase A is described as an example. Since it is very difficult to determine the catalytic roles of such metal ions directly by using the metalloenzymes, several fines of circumstantial evidence have been collected to obtain clues for the catalytic roles. 

An effective multinuclear artificial metalloenzyme would be obtained if an artificial active site comprising of two or more proximal metal centers is designed. A trinuclear artificial metallopeptidase was prepared by using BH... 

Multinuclear metallocatalyst BJ manifested both catalytic activity and substrate selectivity in the hydrolysis of small peptides. The metal centers of the artificial active site of BJ were utilized both in substrate recognition and in catalytic conversion. The structure of the active site obtained by using the bowlshaped molecule, however, is unknown. In addition, it is not possible to synthesize a variety of artificial multinuclear metalloenzymes by the method of transferring catalytic elements confined in a prebuilt cage to a synthetic polymer. 

Fig. 2 Artificial metalloenzymes and asymmetric reactions catalyzed by them. Metal cofactors are introduced by a covalent modification of biotin, b double anchoring to myoglobin, and c non-covalent insertion to serum albumin...

Artificial Metalloenzymes for Enantioselective Catalysis Based on the Biotin-Avidin Technology... 

Abstract Artificial metalloenzymes can be created by incorporating an active metal catalyst precursor in a macromolecular host. When considering such artificial metalloenzymes, the first point to address is how to localize the active metal moiety within the protein scaffold. Although a covalent anchoring strategy may seem most attractive at first, supramolecular anchoring strategy has proven most successful thus far. 

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