13 publications

13 publications

Artificial Metalloenzyme for Enantioselective Sulfoxidation Based on Vanadyl-Loaded Streptavidin

Ward, T.R.

J. Am. Chem. Soc. 2008, 130, 8085-8088, 10.1021/ja8017219

Nature’s catalysts are specifically evolved to carry out efficient and selective reactions. Recent developments in biotechnology have allowed the rapid optimization of existing enzymes for enantioselective processes. However, the ex nihilo creation of catalytic activity from a noncatalytic protein scaffold remains very challenging. Herein, we describe the creation of an artificial enzyme upon incorporation of a vanadyl ion into the biotin-binding pocket of streptavidin, a protein devoid of catalytic activity. The resulting artificial metalloenzyme catalyzes the enantioselective oxidation of prochiral sulfides with good enantioselectivities both for dialkyl and alkyl-aryl substrates (up to 93% enantiomeric excess). Electron paragmagnetic resonance spectroscopy, chemical modification, and mutagenesis studies suggest that the vanadyl ion is located within the biotin-binding pocket and interacts only via second coordination sphere contacts with streptavidin.


Metal: V
Ligand type: Water
Host protein: Streptavidin (Sav)
Anchoring strategy: Supramolecular
Optimization: Genetic
Reaction: Sulfoxidation
Max TON: 27
ee: 93
PDB: ---
Notes: ---

Artificial Metalloenzymes as Selective Catalysts in Aqueous Media

Review

Ward, T.R.

Coord. Chem. Rev. 2008, 252, 751-766, 10.1016/j.ccr.2007.09.016

The fusion of homogeneous and enzymatic catalysis has recently drawn attention due to reported novel activities and high selectivities. The incorporation of metal-catalysts into proteins combines the advantages of both catalytic strategies. Herein we summarize recent approaches of artificial metalloenzymes applied to catalysis. The discussion includes different strategies of anchoring and screening for improved selectivity.


Notes: ---

Artificial Metalloenzymes for Asymmetric Allylic Alkylation on the Basis of the Biotin–Avidin Technology

Ward, T.R.

Angew. Chem. Int. Ed. 2008, 47, 701-705, 10.1002/anie.200703159

Palladium in the active site: The incorporation of a biotinylated palladium diphosphine within streptavidin yielded an artificial metalloenzyme for the title reaction (see scheme). Chemogenetic optimization of the catalyst by the introduction of a spacer (red star) between biotin (green triangle) and palladium and saturation mutagenesis at position S112X afforded both R‐ and S‐selective artificial asymmetric allylic alkylases.


Metal: Pd
Ligand type: Phosphine
Host protein: Streptavidin (Sav)
Anchoring strategy: Supramolecular
Optimization: Chemical & genetic
Reaction: Allylic alkylation
Max TON: 10
ee: 93
PDB: ---
Notes: ---

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

Review

Ward, T.R.

Chimia 2008, 62, 956-961, 10.2533/chimia.2008.956

Artificial metalloenzymes, based on the incorporation of a biotinylated catalytically active organometallic moiety within streptavidin, offer an attractive alternative to homogeneous, heterogeneous and enzymatic catalysis. In this account, we outline our recent results and implications in the developments of such artificial metalloenzymes for various asymmetric transformations, including hydrogenation, transfer hydrogenation, allylic alkylation and sulfoxidation.


Notes: ---

Biosynthesis of a Site-Specific DNA Cleaving Protein

Schultz, P.G.

J. Am. Chem. Soc. 2008, 130, 13194-13195, 10.1021/ja804653f

An E. coli catabolite activator protein (CAP) has been converted into a sequence-specific DNA cleaving protein by genetically introducing (2,2′-bipyridin-5-yl)alanine (Bpy-Ala) into the protein. The mutant CAP (CAP-K26Bpy-Ala) showed comparable binding affinity to CAP-WT for the consensus operator sequence. In the presence of Cu(II) and 3-mercaptopropionic acid, CAP-K26Bpy-Ala cleaves double-stranded DNA with high sequence specificity. This method should provide a useful tool for mapping the molecular details of protein−nucleic acid interactions.


Metal: Cu
Ligand type: Bipyridine
Anchoring strategy: ---
Optimization: Chemical & genetic
Max TON: ---
ee: ---
PDB: ---
Notes: Catabolite activator protein from E. coli

Metal: Fe
Ligand type: Bipyridine
Anchoring strategy: ---
Optimization: Chemical & genetic
Max TON: ---
ee: ---
PDB: ---
Notes: Catabolite activator protein from E. coli

Chemogenetic Protein Engineering: An Efficient Tool for the Optimization of Artificial Metalloenzymes

Review

Ward, T.R.

Chem. Commun. 2008, 4239, 10.1039/b806652c

Artificial metalloenzymes, based on the incorporation of a catalytically active organometallic moiety within a host protein, lie at the interface between organometallic and enzymatic catalysis. In terms of activity, reaction repertoire, substrate range and operating conditions, they take advantage of the versatility of the organometallic chemistry. In contrast, the enantioselectivity is determined by the biomolecular scaffold, which provides a well defined second coordination sphere to the organometallic moiety, reminiscent of enzymes. The attractive feature of such systems is their optimization potential, which combines chemical and genetic methods (i.e. chemogenetic) to screen diversity space. This feature article describes the implementation of such an optimization protocol for artificial transfer hydrogenases, for which we have the most detailed understanding.


Notes: ---

Control of the Coordination Structure of Organometallic Palladium Complexes in an Apo-Ferritin Cage

Ueno, T.; Watanabe, Y.

J. Am. Chem. Soc. 2008, 130, 10512-10514, 10.1021/ja802463a

We report the preparation of organometallic Pd(allyl) dinuclear complexes in protein cages of apo-Fr by reactions with [Pd(allyl)Cl]2 (allyl = η3-C3H5). One of the dinuclear complexes is converted to a trinuclear complex by replacing a Pd-coordinated His residue to an Ala residue. These results suggest that multinuclear metal complexes with various coordination structures could be prepared by the deletion or introduction of His, Cys, and Glu at appropriate positions on protein surface.


Metal: Pd
Ligand type: Allyl
Host protein: Ferritin
Anchoring strategy: Dative
Optimization: ---
Reaction: Suzuki coupling
Max TON: ---
ee: ---
PDB: 2ZG7
Notes: ---

Functionalized Antibodies as Biosensing Materials and Catalysts

Review

Harada, A.

Chem. Lett. 2008, 37, 1184-1189, 10.1246/cl.2008.1184

Monoclonal antibodies have been prepared against water-soluble porphyrins, viologen derivatives, and transition-metal complexes, respectively. These monoclonal antibodies were utilized to devise biosensing and catalytic systems.


Notes: ---

Hemozymes Peroxidase Activity Of Artificial Hemoproteins Constructed From the Streptomyces Lividans Xylanase A and Iron(III)-Carboxy-Substituted Porphyrins

Mahy, J.-P.

Bioconjug. Chem. 2008, 19, 899-910, 10.1021/bc700435a

To develop artificial hemoproteins that could lead to new selective oxidation biocatalysts, a strategy based on the insertion of various iron-porphyrin cofactors into Xylanase A (Xln10A) was chosen. This protein has a globally positive charge and a wide enough active site to accommodate metalloporphyrins that possess negatively charged substituents such as microperoxidase 8 (MP8), iron(III)-tetra-α4-ortho-carboxyphenylporphyrin (Fe(ToCPP)), and iron(III)-tetra-para-carboxyphenylporphyrin (Fe(TpCPP)). Coordination chemistry of the iron atom and molecular modeling studies showed that only Fe(TpCPP) was able to insert deeply into Xln10A, with a KD value of about 0.5 µM. Accordingly, Fe(TpCPP)-Xln10A bound only one imidazole molecule, whereas Fe(TpCPP) free in solution was able to bind two, and the UV–visible spectrum of the Fe(TpCPP)-Xln10A-imidazole complex suggested the binding of an amino acid of the protein on the iron atom, trans to the imidazole. Fe(TpCPP)-Xln10A was found to have peroxidase activity, as it was able to catalyze the oxidation of typical peroxidase cosubstrates such as guaiacol and o-dianisidine by H2O2. With these two cosubstrates, the KM value measured with the Fe(TpCPP)-Xln10A complex was higher than those values observed with free Fe(TpCPP), probably because of the steric hindrance and the increased hydrophobicity caused by the protein around the iron atom of the porphyrin. The peroxidase activity was inhibited by imidazole, and a study of the pH dependence of the oxidation of o-dianisidine suggested that an amino acid with a pKA of around 7.5 was participating in the catalysis. Finally, a very interesting protective effect against oxidative degradation of the porphyrin was provided by the protein.


Metal: Fe
Ligand type: Porphyrin
Host protein: Xylanase A (XynA)
Anchoring strategy: Supramolecular
Optimization: ---
Max TON: ---
ee: ---
PDB: ---
Notes: kcat/KM = 1083 M-1 * s-1

Molecular Design of Heteroprotein Assemblies Providing a Bionanocup as a Chemical Reactor

Ueno, T.; Watanabe, Y.

Small 2008, 4, 50-54, 10.1002/smll.200700855

A bionanocup chemical reactor is constructed from a heteroprotein assembly from bacteriophage T4. The preparation of a stable iron(III) porphyrin–bionanocup composite is described. The hydrophobic cup provides a space suitable for the fixation of low‐water‐solubility iron(III) porphyrins. The application of the iron(III) porphyrin–bionanocup composites for the catalysis of sulfoxidation of thioanisoles is demonstrated (see figure).


Metal: Fe
Host protein: (gp27-gp5)3
Anchoring strategy: Cystein-maleimide
Optimization: ---
Max TON: ---
ee: ---
PDB: ---
Notes: ---

Protein Scaffold of a Designed Metalloenzyme Enhances the Chemoselectivity in Sulfoxidation of Thioanisole

Lu, Y.

Chem. Commun. 2008, 1665, 10.1039/b718915j

We demonstrate that incorporation of MnSalen into a protein scaffold enhances the chemoselectivity in sulfoxidation of thioanisole and find that both the polarity and hydrogen bonding of the protein scaffold play an important role in tuning the chemoselectivity.


Metal: Mn
Ligand type: Salen
Host protein: Myoglobin (Mb)
Anchoring strategy: Supramolecular
Optimization: Chemical & genetic
Reaction: Sulfoxidation
Max TON: 5.2
ee: 60
PDB: ---
Notes: Sperm whale myoglobin

Synthesis of a New Estradiol–Iron Metalloporphyrin Conjugate Used to Build up a New Hybrid Biocatalyst for Selective Oxidations by the ‘Trojan Horse’ Strategy

Mahy, J.-P.

Tetrahedron Lett. 2008, 49, 1865-1869, 10.1016/j.tetlet.2008.01.022

The synthesis of a new cationic iron metalloporphyrin–estradiol conjugate is reported. After a study of its association with the anti-estradiol antibody 7A3 by UV–visible spectroscopy, the influence of the antibody on the sulfoxidation of thioanisole by H2O2 catalyzed by the iron–metalloporphyrin has been investigated.


Metal: Fe
Ligand type: Porphyrin
Host protein: Antibody 7A3
Anchoring strategy: Supramolecular
Optimization: ---
Reaction: Sulfoxidation
Max TON: 12
ee: 8
PDB: ---
Notes: ---

X-Ray Structure and Designed Evolution of an Artificial Transfer Hydrogenase

Ward, T.R.

Angew. Chem. Int. Ed. 2008, 47, 1400-1404, 10.1002/anie.200704865

A structure is worth a thousand words: Guided by the X‐ray structure of an S‐selective artificial transfer hydrogenase, designed evolution was used to optimize the selectivity of hybrid catalysts. Fine‐tuning of the second coordination sphere of the ruthenium center (see picture, orange sphere) by introduction of two point mutations allowed the identification of selective artificial transfer hydrogenases for the reduction of dialkyl ketones.


Metal: Ru
Ligand type: Amino-sulfonamide; Benzene
Host protein: Streptavidin (Sav)
Anchoring strategy: Supramolecular
Optimization: Chemical & genetic
Max TON: 100
ee: 92
PDB: 2QCB
Notes: ---

Metal: Ru
Ligand type: Amino-sulfonamide; P-cymene
Host protein: Streptavidin (Sav)
Anchoring strategy: Supramolecular
Optimization: Chemical & genetic
Max TON: 97
ee: 96
PDB: 2QCB
Notes: ---