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Metal ions of metalloenzymes participate in catalysis either by acting as Lewis acid catalysts or by changing their oxidation states. The most important metal ion that participates as a Lewis acid catalyst in the actions of metalloenzymes is the Zn(II) ion, and virtually all types of organic reactions are catalyzed by Zn(II)metalloenzymes.

Effective artificial enzymes have been designed by adopting macromolecular systems for catalyst-substrate complexes. Artificial active sites comprising two or more organic functional groups were built on macromolecular backbones, leading to several types of organic artificial proteases. The activity of metal centers for peptide or DNA hydrolysis was greatly enhanced by attachment to polystyrene, leading to artificial metallopeptidases with substrate selectivity as well as artificial metallonucleases with high catalytic activity for double stranded DNA. A small artificial protease selective for a macromolecular target protein was synthesized. Target-specific artificial proteases can be used as protein-cleaving catalytic drugs.

Poly(chloromethylstyrene-co-divinylbenzene) (PCD) with 2% cross-linkage is developed as a backbone of immobile artificial enzymes. As the first artificial enzyme built on PCD, an artificial metalloproteinase is prepared by attaching the Cu(II) complex of cyclen as the catalytic center and guanidinium ion as the binding unit. The PCD derivatives prepared were characterized by scanning electron microscopy, IR spectroscopy, elemental analysis, titration of chloromethylphenyl moiety with triethylamine, complexation of p-nitrobenzoate ion to the guanidinium moiety, quantification of the cyclen moieties retaining high affinity for the Cu(II) ion, and determination of log K f for the Cu(II) binding sites. The proteinase activity was measured with γ-globulin (Gbn) by following cleavage of the two chains of Gbn by electrophoresis. The catalytic activity of the Cu(II) complex of cyclen toward Gbn was enhanced by more than 104 times upon attachment to PCD. Gbn complexed to the PCD derivative containing both Cu(II)-cyclen and guanidinium moieties was cleaved by hydrolysis into many pieces with a half-life as short as 10−30 min at pH 4.5−7 and 4 °C. Kinetic data revealed that guanidinium ions attached to PCD acted as effective binding sites for the protein, contributing considerably to the overall catalytic power of the immobile artificial metalloproteinase.

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A peptide-cleaving catalyst selective for peptide deformylase (PDF) was obtained from a library containing about 15 000 catalyst candidates. The catalyst cleaved the polypeptide backbone of PDF at Gln(152)-Arg(153). Docking simulations suggested multiple modes of interactions in the complex formed between the catalyst and PDF.

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Catalytic drugs based on target-selective artificial proteases have been proposed as a new paradigm in drug design. Peptide-cleavage agents selective for pathogenic proteins of Alzheimer's disease, type 2 diabetes mellitus or Parkinson's disease have been prepared using the Co(III) aqua complex (Co(III)cyclen) of 1,4,7,10-tetraazacyclododecane as the catalytic center. In the present study, the Co(III) aqua complex (Co(III)oxacyclen) of 1-oxa-4,7,10-triazacyclododecane was examined in search of an improved catalytic center for peptide-cleavage agents. An X-ray crystallographic study of [Co(oxacyclen)(CO(3))](ClO(4)), titration of Co(III)oxacyclen, and kinetic studies on the cleavage of albumin, gamma-globulin, lysozyme, and myoglobin by Co(III)oxacyclen were carried out. Considerably higher proteolytic activity was observed for Co(III)oxacyclen in comparison with Co(III)cyclen, indicating that better target-selective artificial metalloproteases would be obtained using Co(III)oxacyclen as the catalytic center. The improved proteolytic activity was attributed to either steric effects or the increased Lewis acidity of the Co(III) center. The kinetic data also predicted that side effects due to the cleavage of nontarget proteins by a catalytic drug based on Co(III)oxacyclen would be insignificant.