Bäckvall, J.E.; Diéguez, M.; Pàmies, O.

Adv. Synth. Catal. 2015, 357, 1567-1586, 10.1002/adsc.201500290

Artificial metalloenzymes combine the excellent selective recognition/binding properties of enzymes with transition metal catalysts, and therefore many asymmetric transformations can benefit from these entities. The search for new successful strategies in the construction of metal‐enzyme hybrid catalysts has therefore become a very active area of research. This review discusses all the developed strategies and the latest advances in the synthesis and application in asymmetric catalysis of artificial metalloenzymes with future directions for their design, synthesis and application (Sections 2–4). Finally, advice is presented (to the non‐specialist) on how to prepare and use artificial metalloenzymes (Section 5).

Craik, C.S.

Biochemistry 1995, 34, 2172-2180, 10.1021/bi00007a010

Histidine substrate specificity has been engineered into trypsin by creating metal binding sites for Ni2+ and Zn2+ ions. The sites bridge the substrate and enzyme on the leaving-group side of the scissile bond. Application of simple steric and geometric criteria to a crystallographically derived enzyme- substrate model suggested that histidine specificity at the P2' position might be acheived by a tridentate site involving amino acid residues 143 and 151 of trypsin. Trypsin N143H/E151H hydrolyzes a P2'- His-containing peptide (AGPYAHSS) exclusively in the presence of nickel or zinc with a high level of catalytic efficiency. Since cleavage following the tyrosine residue is normally highly disfavored by trypsin, this result demonstrates that a metal cofactor can be used to modulate specificity in a designed fashion. The same geometric criteria applied in the primary SI binding pocket suggested that the single-site mutation D189H might effect metal-dependent His specificity in trypsin. However, kinetic and crystallographic analysis of this variant showed that the design was unsuccessful because His 189 rotates away from substrate causing a large perturbation in adjacent surface loops. This observation suggests that the reason specificity modification at the trypsin S1 site requires extensive mutagenesis is because the pocket cannot deform locally to accommodate alternate PI side chains. By taking advantage of the extended subsites, an alternate substrate specificity has been engineered into trypsin.

Coleman, J.E.

Nature 1967, 214, 193-194, 10.1038/214193a0

ACETAZOLAMIDE (2-acetylamino-1,3,4-thiadiazole-5-sulphonamide, ‘Diamox’) is the most potent known inhibitor of the zinc enzyme carbonic anhydrase. This communication reports the direct demonstration that binding of acetazolamide to human carbonic anhydrase requires the presence of a metal ion at the active site and that binding depends on the species of divalent metal ion present. Zinc (II) and cobalt (II) ions are the only ions which induce the formation of very stable acetazolamide carbonic anhydrase complexes and are also the ions which most effectively catalyse the hydration of carbon dioxide and the hydrolysis of p-nitrophenyl acetate. Metal-binding monodentate ions, CN−, HS−, OCN−, and N3−, known as effective carbonic anhydrase inhibitors, compete for the acetazolamide binding site of the zinc enzyme.