19 publications

19 publications

A De Novo Designed Metalloenzyme for the Hydration of CO2

Pecoraro, V. L.

Angew. Chem., Int. Ed., 2014, 10.1002/anie.201404925

Protein design will ultimately allow for the creation of artificial enzymes with novel functions and unprecedented stability. To test our current mastery of nature’s approach to catalysis, a ZnII metalloenzyme was prepared using de novo design. α3DH3 folds into a stable single‐stranded three‐helix bundle and binds ZnII with high affinity using His3O coordination. The resulting metalloenzyme catalyzes the hydration of CO2 better than any small molecule model of carbonic anhydrase and with an efficiency within 1400‐fold of the fastest carbonic anhydrase isoform, CAII, and 11‐fold of CAIII.


Metal: Zn
Ligand type: Amino acid
Host protein: α3D peptide
Anchoring strategy: Dative
Optimization: Chemical & genetic
Max TON: ---
ee: ---
PDB: ---
Notes: kcat/KM ≈ 3.8*104 M-1*s-1

A Designed Supramolecular Protein Assembly with In Vivo Enzymatic Activity

Tezcan, F. A.

Science, 2014, 10.1126/science.1259680

The generation of new enzymatic activities has mainly relied on repurposing the interiors of preexisting protein folds because of the challenge in designing functional, three-dimensional protein structures from first principles. Here we report an artificial metallo-β-lactamase, constructed via the self-assembly of a structurally and functionally unrelated, monomeric redox protein into a tetrameric assembly that possesses catalytic zinc sites in its interfaces. The designed metallo-β-lactamase is functional in the Escherichia coli periplasm and enables the bacteria to survive treatment with ampicillin. In vivo screening of libraries has yielded a variant that displays a catalytic proficiency [(kcat/Km)/kuncat] for ampicillin hydrolysis of 2.3 × 106 and features the emergence of a highly mobile loop near the active site, a key component of natural β-lactamases to enable substrate interactions.


Metal: Zn
Ligand type: Amino acid
Host protein: Cytochrome cb562
Anchoring strategy: Dative
Optimization: Genetic
Max TON: ---
ee: ---
PDB: 4U9E
Notes: ---

A Highly Specific Metal-Activated Catalytic Antibody

Janda, K. D.; Lerner, R. A.

J. Am. Chem. Soc., 1993, 10.1021/ja00064a068

n/a


Metal: Zn
Ligand type: Undefined
Host protein: IgG 84A3
Anchoring strategy: Undefined
Optimization: ---
Max TON: ---
ee: ---
PDB: ---
Notes: Substrate specificty

A Metal Ion Regulated Artificial Metalloenzyme

Roelfes, G.

Dalton Trans., 2017, 10.1039/C7DT00533D

Regulation of enzyme activity is essential in living cells. The rapidly increasing number of designer enzymes with new-to-nature activities makes it necessary to develop novel strategies for controlling their catalytic activity. Here we present the development of a metal ion regulated artificial metalloenzyme created by combining two anchoring strategies, covalent and supramolecular, for introducing a regulatory and a catalytic site, respectively. This artificial metalloenzyme is activated in the presence of Fe2+ ions, but only marginally in the presence of Zn2+.


Metal: Fe
Ligand type: Bypyridine
Host protein: LmrR
Anchoring strategy: Covalent
Optimization: Genetic
Max TON: 14
ee: 75
PDB: ---
Notes: ---

Metal: Zn
Ligand type: Bypyridine
Host protein: LmrR
Anchoring strategy: Covalent
Optimization: Genetic
Max TON: 6
ee: 80
PDB: ---
Notes: ---

Catalysis by a De Novo Zinc-Mediated Protein Interface: Implications for Natural Enzyme Evolution and Rational Enzyme Engineering

Kuhlman, B.

Biochemistry, 2012, 10.1021/bi201881p


Metal: Zn
Ligand type: Amino acid
Anchoring strategy: Dative
Optimization: Chemical & genetic
Max TON: >50
ee: ---
PDB: 3V1C
Notes: ---

Computational Redesign of a Mononuclear Zinc Metalloenzyme for Organophosphate Hydrolysis

Baker, D.

Nat. Chem. Biol., 2012, 10.1038/NChemBio.777


Metal: Zn
Ligand type: Amino acid
Anchoring strategy: Dative
Optimization: Genetic
Max TON: >140
ee: ---
PDB: 3T1G
Notes: kcat/KM ≈ 104 M-1*s-1

Design and Evolution of New Catalytic Activity with an Existing Protein Scaffold

Kim, H. S.

Science, 2006, 10.1126/science.1118953


Metal: Zn
Ligand type: Amino acid
Host protein: Glyoxalase II (Human)
Anchoring strategy: Dative
Optimization: Genetic
Max TON: ---
ee: ---
PDB: 2F50
Notes: kcat/KM ≈ 184 M-1*s-1

Engineered Metal Regulation of Trypsin Specificity

Craik, C. S.

Biochemistry, 1995, 10.1021/bi00007a010


Metal: Zn
Ligand type: Amino acid
Host protein: Trypsin
Anchoring strategy: Dative
Optimization: Genetic
Max TON: ---
ee: ---
PDB: ---
Notes: Substrate specificty

Metal: Ni
Ligand type: Amino acid
Host protein: Trypsin
Anchoring strategy: Dative
Optimization: Genetic
Max TON: ---
ee: ---
PDB: ---
Notes: Substrate specificty

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.

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.

Importance of Scaffold Flexibility/Rigidity in the Design and Directed Evolution of Artificial Metallo-β-Lactamases

Song, W. J.; Tezcan, F. A.

J. Am. Chem. Soc., 2017, 10.1021/jacs.7b08981


Metal: Zn
Ligand type: Amino acid
Host protein: Zn8:AB54
Anchoring strategy: Dative
Optimization: Genetic
Reaction: Hydrolysis
Max TON: ---
ee: ---
PDB: 5XZI
Notes: Supramolecular protein scaffold constructed from cytochrome cb562 building blocks, Ampicillin hydrolysis: kcat/KM = 130 min-1 * M-1

Metal: Zn
Ligand type: Amino acid
Host protein: Zn8:AB54 (mutant C96T)
Anchoring strategy: Dative
Optimization: Genetic
Reaction: Hydrolysis
Max TON: ---
ee: ---
PDB: 5XZJ
Notes: Supramolecular protein scaffold constructed from cytochrome cb562 building blocks, Ampicillin hydrolysis: kcat/KM = 210 min-1 * M-1

Influence of Active Site Location on Catalytic Activity in De Novo-Designed Zinc Metalloenzymes

Pecoraro, V. L.

J. Am. Chem. Soc., 2013, 10.1021/ja401537t


Metal: Hg; Zn
Ligand type: Amino acid
Host protein: TRI peptide
Anchoring strategy: Dative
Optimization: Chemical & genetic
Max TON: ---
ee: ---
PDB: 3PBJ
Notes: Influence of position of Zn and Hg ion on catalytic activity of the ArM tested. PDB ID 3PBJ = Structure of an analogue.

Metal Incorporated Horseradish Peroxidase (HRP) Catalyzed Oxidation of Resveratrol: Selective Dimerization or Decomposition

Pan, Y.

RSC Adv., 2013, 10.1039/c3ra43784a


Metal: Ca; Co; Mn; Ni; Zn
Ligand type: Undefined
Anchoring strategy: Undefined
Optimization: Chemical
Reaction: Oxidation
Max TON: ---
ee: ---
PDB: ---
Notes: Oxidation of resveratrol. Dimerisation product obtained.

Multifunctional Nanoenzymes from Carbonic Anhydrase Skeleton

Yilmaz, F.

Process Biochem., 2018, 10.1016/j.procbio.2018.06.005


Metal: Zn
Ligand type: Amino acid
Host protein: Carbonic anhydrase (CA)
Anchoring strategy: Metal substitution
Optimization: Chemical
Reaction: Hydrolysis
Max TON: ---
ee: ---
PDB: ---
Notes: Cross-linked carbonic anhydrase nano-enzyme particles (93 nm in diameter). Hydrolysis of 4-nitrophenyl acetate.

Metal: Rh
Ligand type: Amino acid
Host protein: Carbonic anhydrase (CA)
Anchoring strategy: Metal substitution
Optimization: Chemical
Reaction: Hydration
Max TON: ---
ee: ---
PDB: ---
Notes: Cross-linked carbonic anhydrase nano-enzyme particles (93 nm in diameter). Hydration of styrene.

Metal: Mn
Ligand type: Amino acid
Host protein: Carbonic anhydrase (CA)
Anchoring strategy: Metal substitution
Optimization: Chemical
Reaction: Oxidation
Max TON: ---
ee: ---
PDB: ---
Notes: Cross-linked carbonic anhydrase nano-enzyme particles (93 nm in diameter). Oxidation of styrene.

Neocarzinostatin-Based Hybrid Biocatalysts with a RNase like Activity

Mahy, J.-P.; Ricoux, R.

Bioorg. Med. Chem., 2014, 10.1016/j.bmc.2014.05.063


Metal: Zn
Ligand type: Poly-pyridine
Anchoring strategy: Supramolecular
Optimization: ---
Max TON: ---
ee: ---
PDB: ---
Notes: kcat/KM = 13.6 M-1 * s-1

Photoinduced Electron Transfer within Supramolecular Hemoprotein Co-Assemblies and Heterodimers Containing Fe and Zn Porphyrins

Oohora, K.

J. Inorg. Biochem., 2019, 10.1016/j.jinorgbio.2019.01.001


Metal: Fe; Zn
Ligand type: Protoporphyrin IX
Host protein: Cytochrome b562
Anchoring strategy: Cystein-maleimide; Supramolecular
Optimization: Chemical & genetic
Reaction: Electron transfer
Max TON: ---
ee: ---
PDB: ---
Notes: ---

Selection and Evolution of Enzymes from a Partially Randomized Non-Catalytic Scaffold

Seelig, B.; Szostak, J. W.

Nature, 2007, 10.1038/nature06032


Metal: Zn
Ligand type: Amino acid
Anchoring strategy: Dative
Optimization: Genetic
Reaction: RNA ligation
Max TON: >7
ee: ---
PDB: ---
Notes: ---

Sequence-Specific Peptide Cleavage Catalyzed by an Antibody

Lerner, R. A.

Science, 1989, 10.1126/science.2922606


Metal: Zn
Ligand type: Tetramine
Host protein: Antibody 28F11
Anchoring strategy: Supramolecular
Optimization: Chemical
Max TON: 400
ee: ---
PDB: ---
Notes: ---

Structure and Dynamics of a Primordial Catalytic fold Generated by In Vitro Evolution

Seelig, B.

Nat. Chem. Biol., 2012, 10.1038/nchembio.1138


Metal: Zn
Ligand type: Amino acid
Anchoring strategy: Dative
Optimization: Genetic
Reaction: RNA ligation
Max TON: ---
ee: ---
PDB: 2LZE
Notes: ---