6 publications

6 publications

A Clamp-Like Biohybrid Catalyst for DNA Oxidation

Nolte, R. J. M.

Nat. Chem., 2013, 10.1038/NCHEM.1752

In processive catalysis, a catalyst binds to a substrate and remains bound as it performs several consecutive reactions, as exemplified by DNA polymerases. Processivity is essential in nature and is often mediated by a clamp-like structure that physically tethers the catalyst to its (polymeric) template. In the case of the bacteriophage T4 replisome, a dedicated clamp protein acts as a processivity mediator by encircling DNA and subsequently recruiting its polymerase. Here we use this DNA-binding protein to construct a biohybrid catalyst. Conjugation of the clamp protein to a chemical catalyst with sequence-specific oxidation behaviour formed a catalytic clamp that can be loaded onto a DNA plasmid. The catalytic activity of the biohybrid catalyst was visualized using a procedure based on an atomic force microscopy method that detects and spatially locates oxidized sites in DNA. Varying the experimental conditions enabled switching between processive and distributive catalysis and influencing the sliding direction of this rotaxane-like catalyst.


Metal: Mn
Ligand type: Porphyrin
Host protein: gp45
Anchoring strategy: Covalent
Optimization: ---
Max TON: ---
ee: ---
PDB: 1CZD
Notes: ---

Alteration of the Oxygen-Dependent Reactivity of De Novo Due Ferri Proteins

DeGrado, W. F.

Nat. Chem., 2012, 10.1038/NCHEM.1454

De novo proteins provide a unique opportunity to investigate the structure–function relationships of metalloproteins in a minimal, well-defined and controlled scaffold. Here, we describe the rational programming of function in a de novo designed di-iron carboxylate protein from the Due Ferri family. Originally created to catalyse the O2-dependent, two-electron oxidation of hydroquinones, the protein was reprogrammed to catalyse the selective N-hydroxylation of arylamines by remodelling the substrate access cavity and introducing a critical third His ligand to the metal-binding cavity. Additional second- and third-shell modifications were required to stabilize the His ligand in the core of the protein. These structural changes resulted in at least a 106-fold increase in the relative rate between the arylamine N-hydroxylation and hydroquinone oxidation reactions. This result highlights the potential for using de novo proteins as scaffolds for future investigations of the geometric and electronic factors that influence the catalytic tuning of di-iron active sites.


Metal: Fe
Ligand type: Amino acid
Host protein: Due Ferro 1
Anchoring strategy: Dative
Optimization: Genetic
Reaction: N-Hydroxylation
Max TON: ---
ee: ---
PDB: 2LFD
Notes: ---

Evolving Artificial Metalloenzymes via Random Mutagenesis

Lewis, J. C.

Nat. Chem., 2018, 10.1038/nchem.2927


Metal: Rh
Ligand type: OAc
Anchoring strategy: Covalent
Optimization: Chemical & genetic
Reaction: Cyclopropanation
Max TON: 66
ee: 94
PDB: 5T88
Notes: Mutagenesis of the ArM by error-prone PCR

Metal: Rh
Ligand type: OAc
Anchoring strategy: Covalent
Optimization: Chemical & genetic
Reaction: N-H Insertion
Max TON: 73
ee: 40
PDB: 5T88
Notes: Mutagenesis of the ArM by error-prone PCR

Metal: Rh
Ligand type: OAc
Anchoring strategy: Covalent
Optimization: Chemical & genetic
Reaction: S-H Insertion
Max TON: 64
ee: 32
PDB: 5T88
Notes: Mutagenesis of the ArM by error-prone PCR

Metal: Rh
Ligand type: OAc
Anchoring strategy: Covalent
Optimization: Chemical & genetic
Reaction: Si-H Insertion
Max TON: 35
ee: 64
PDB: 5T88
Notes: Mutagenesis of the ArM by error-prone PCR

Hydrolytic Catalysis and Structural Stabilization in a Designed Metalloprotein

Pecoraro, V. L.

Nat. Chem., 2011, 10.1038/NCHEM.1201


Metal: Hg; Zn
Ligand type: Amino acid
Host protein: TRI peptide
Anchoring strategy: Dative
Optimization: Chemical & genetic
Max TON: >10
ee: ---
PDB: 3PBJ
Notes: Zn ion for catalytic activity, Hg ion for structural stability of the ArM. PDB ID 3PBJ = Structure of an analogue.

Metal: Hg; Zn
Ligand type: Amino acid
Host protein: TRI peptide
Anchoring strategy: Dative
Optimization: Chemical & genetic
Max TON: ---
ee: ---
PDB: 3PBJ
Notes: Zn ion for catalytic activity, Hg ion for structural stability of the ArM, kcat/KM ≈ 1.8*105 M-1*s-1. PDB ID 3PBJ = Structure of an analogue.

Reconstitution of [Fe]-Hydrogenase Using Model Complexes

Hu, X.; Shima, S.

Nat. Chem., 2015, 10.1038/Nchem.2382


Metal: Fe
Ligand type: Amino acid
Anchoring strategy: Covalent
Optimization: Chemical
Max TON: ---
ee: ---
PDB: ---
Notes: DFT calculations of the reaction mechanism.

Synthetic Cascades are Enabled by Combining Biocatalysts with Artificial Metalloenzymes

Turner, N. J.; Ward, T. R.

Nat. Chem., 2013, 10.1038/NCHEM.1498


Metal: Ir
Ligand type: Amino-sulfonamide; Cp*
Host protein: Streptavidin (Sav)
Anchoring strategy: Supramolecular
Optimization: Genetic
Max TON: 100
ee: > 99
PDB: ---
Notes: Cascade