Assuntos
Materiais Biocompatíveis/metabolismo , Oxigenases de Função Mista/metabolismo , Transportadores de Cassetes de Ligação de ATP/química , Sítios de Ligação , Materiais Biocompatíveis/química , Catálise , Domínio Catalítico , Cristalografia por Raios X , Proteínas de Escherichia coli/química , Oxigenases de Função Mista/química , Simulação de Acoplamento Molecular , Oxirredução , Especificidade por Substrato , Sulfetos/químicaRESUMO
Understanding the interaction of a protein with a relevant ligand is crucial for the design of an artificial metalloenzyme. Our own interest is focused on the synthesis of artificial monooxygenases. In an initial effort, we have used the periplasmic nickel-binding protein NikA from Escherichia coli and iron complexes in which N(2)Py(2) ligands (where Py is pyridine) have been varied in terms of charge, aromaticity, and size. Six "NikA/iron complex" hybrids have been characterized by X-ray crystallography, and their interactions and solution properties have been studied. The hybrids are stable as indicated by their K (d) values, which are all in the micromolar range. The X-ray structures show that the ligands interact with NikA through salt bridges with arginine residues and π-stacking with a tryptophan residue. We have further characterized these interactions using quantum mechanical calculations and determined that weak CH/π hydrogen bonds finely modulate the stability differences between hybrids. We emphasize the important role of the tryptophan residues. Thus, our study aims at the complete characterization of the factors that condition the interaction of an artificial ligand and a protein and their implications for catalysis. Besides its potential usefulness in the synthesis of artificial monooxygenases, our approach should be generally applicable in the field of artificial metalloenzymes.
Assuntos
Transportadores de Cassetes de Ligação de ATP/química , Proteínas de Escherichia coli/química , Escherichia coli/química , Compostos de Ferro/química , Metaloproteínas/química , Níquel/metabolismo , Transportadores de Cassetes de Ligação de ATP/metabolismo , Materiais Biomiméticos/química , Materiais Biomiméticos/metabolismo , Cristalografia por Raios X , Escherichia coli/metabolismo , Proteínas de Escherichia coli/metabolismo , Compostos de Ferro/metabolismo , Ligantes , Metaloproteínas/metabolismo , Modelos Moleculares , Piridinas/química , Piridinas/metabolismo , Água/químicaRESUMO
Catalytic antibodies with a metalloporphyrin cofactor or <
Assuntos
Anticorpos Catalíticos/metabolismo , Anticorpos Monoclonais/metabolismo , Peroxidases/metabolismo , Animais , Anticorpos Catalíticos/biossíntese , Anticorpos Monoclonais/biossíntese , Catálise , Oxirredução , Peroxidases/imunologiaRESUMO
Catalytic antibodies with a metalloporphyrin cofactor represent a new generation of biocatalysts tailored for selective oxidations. Thus monoclonal antibodies, 3A3, were raised against microperoxidase 8 (MP8), and the corresponding 3A3-MP8 complexes were shown previously to have a high peroxidase activity. This paper shows that those complexes also catalyzed efficiently the nitration of phenol into 2- and 4-nitrophenol by NO2- in the presence of H2O2. pH dependence studies suggested that no amino acid from the antibody protein participated in the heterolytic cleavage of the O-O bond of H2O2. The inhibition of the reaction by cyanide and radical scavengers suggested a MP8-mediated peroxidase-like mechanism, involving the reduction of high-valent iron-oxo species by NO2- and phenol producing, respectively, NO2* and phenoxy radicals, which then reacted to give nitrophenols. Finally, the antibody protein appears to have two major roles: (i) it protects MP8 toward oxidative degradations and (ii) it induces a regioselectivity of the reaction toward the formation of 2-nitrophenol.