Manganese-Substituted Carbonic Anhydrase as a New Peroxidase
Chem. - Eur. J. 2006, 12, 1587-1596, 10.1002/chem.200501413
Carbonic anhydrase is a zinc metalloenzyme that catalyzes the hydration of carbon dioxide to bicarbonate. Replacing the active‐site zinc with manganese yielded manganese‐substituted carbonic anhydrase (CA[Mn]), which shows peroxidase activity with a bicarbonate‐dependent mechanism. In the presence of bicarbonate and hydrogen peroxide, (CA[Mn]) catalyzed the efficient oxidation of o‐dianisidine with kcat/KM=1.4×106 m−1 s−1, which is comparable to that for horseradish peroxidase, kcat/KM=57×106 m−1 s−1. CA[Mn] also catalyzed the moderately enantioselective epoxidation of olefins to epoxides (E=5 for p‐chlorostyrene) in the presence of an amino‐alcohol buffer, such as N,N‐bis(2‐hydroxyethyl)‐2‐aminoethanesulfonic acid (BES). This enantioselectivity is similar to that for natural heme‐based peroxidases, but has the advantage that CA[Mn] avoids the formation of aldehyde side products. CA[Mn] degrades during the epoxidation limiting the yield of the epoxidations to <12 %. Replacement of active‐site residues Asn62, His64, Asn67, Gln92, or Thr200 with alanine by site‐directed mutagenesis decreased the enantioselectivity demonstrating that the active site controls the enantioselectivity of the epoxidation.
Max TON: 22ee: 67PDB: ---Notes: ---
Max TON: 9.5ee: 55PDB: 4CACNotes: PDB ID 4CAC = Structure of Zn containing hCAII
Stereoselective Hydrogenation of Olefins Using Rhodium-Substituted Carbonic Anhydrase—A New Reductase
Chem. - Eur. J. 2009, 15, 1370-1376, 10.1002/chem.200801673
One useful synthetic reaction missing from nature's toolbox is the direct hydrogenation of substrates using hydrogen. Instead nature uses cofactors like NADH to reduce organic substrates, which adds complexity and cost to these reductions. To create an enzyme that can directly reduce organic substrates with hydrogen, researchers have combined metal hydrogenation catalysts with proteins. One approach is an indirect link where a ligand is linked to a protein and the metal binds to the ligand. Another approach is direct linking of the metal to protein, but nonspecific binding of the metal limits this approach. Herein, we report a direct hydrogenation of olefins catalyzed by rhodium(I) bound to carbonic anhydrase (CA‐[Rh]). We minimized nonspecific binding of rhodium by replacing histidine residues on the protein surface using site‐directed mutagenesis or by chemically modifying the histidine residues. Hydrogenation catalyzed by CA‐[Rh] is slightly slower than for uncomplexed rhodium(I), but the protein environment induces stereoselectivity favoring cis‐ over trans‐stilbene by about 20:1. This enzyme is the first cofactor‐independent reductase that reduces organic molecules using hydrogen. This catalyst is a good starting point to create variants with tailored reactivity and selectivity. This strategy to insert transition metals in the active site of metalloenzymes opens opportunities to a wider range of enzyme‐catalyzed reactions.
Metal: RhLigand type: CODOptimization: GeneticReaction: HydrogenationMax TON: 15.8ee: ---PDB: ---Notes: ---
Metal: RhLigand type: CODOptimization: GeneticReaction: HydrogenationMax TON: 80.5ee: ---PDB: 4CACNotes: PDB ID 4CAC = Structure of Zn containing hCAII
Transforming Carbonic Anhydrase into Epoxide Synthase by Metal Exchange
ChemBioChem 2006, 7, 1013-1016, 10.1002/cbic.200600127
Enantioselective epoxidation of styrene was observed in the presence of manganese‐containing carbonic anhydrase as catalyst. The probable oxygen‐transfer reagent is peroxymonocarbonate, which has a structural similarity with the hydrogenocarbonate substrate of the natural reaction. Styrene was chosen as the enzyme possesses a small hydrophobic cavity close to the active site.
Max TON: 4.1ee: 52PDB: ---Notes: ---
Max TON: 10.3ee: 40PDB: ---Notes: ---