Heme-Containing Oxygenases

A radical is any molecule that contains one or more unpaired electrons. Radicals are normally generated in many metabolic pathways. Some of these radicals can exist in a free form and subsequently interact with various tissue components resulting in dysfunction. The potential role of oxygen- or xenobiotic-derived free radicals in the pathology of several human diseases has stimulated extensive research linking the toxicity of numerous xenobiotics and disease processes to a free radical mechanism. However, because free radical-mediated changes are pervasive and often poorly understood, the question of whether such species are a major cause of tissue injury and human disease remains equivocal. This review discusses cellular sources of various radical species and their reactions with vital cellular constituents. Examples of purported free radical-mediated disorders are discussed in detail to provide insights into the controversy over whether free radicals are important mediators of tissue injury.

Free radicals as mediators of tissue injury and disease.

ion from camphor is affected by the choices made during setup.131 In this context, it should be noted that early QM/MM work on different P450cam intermediates and on different steps of the catalytic cycle employed separate setups that had been generated independently, which raises the question whether it is possible to handle the whole cycle with the same QM/MM setup. This is indeed the case: following a standard protocol that starts from the X-ray structure of the ferrous dioxygen complex and that involves careful assignment of protonation states and hydration, one can generate a common setup that gives QM/MM results for P450cam which are generally consistent with those obtained previously with individual setups.54 This confirms that setup issues can be resolved in a consistent manner if proper care is exercised. P450 Enzymes Modeled by QM/MM Calculations Chemical Reviews, 2010, Vol. 110, No. 2 955

P450 Enzymes: Their Structure, Reactivity, and Selectivity—Modeled by QM/MM Calculations

Structural factors that influence functional properties are examined in the case of four heme enzymes: cytochrome P-450, chloroperoxidase, horseradish peroxidase, and secondary amine mono-oxygenase. The identity of the axial ligand, the nature of the heme environment, and the steric accessibility of the heme iron and heme edge combine to play major roles in determining the reactivity of each enzyme. The importance of synthetic porphyrin models in understanding the properties of the protein-free metal center is emphasized. The conclusions described herein have been derived from studies at the interface between biological and inorganic chemistry.

https://science.sciencemag.org/content/sci/240/4851/433.full.pdf

Probing structure-function relations in heme-containing oxygenases and peroxidases

Antibiotic resistance can be achieved by horizontal acquisition of resistance genes (carried by plasmids or transposons), by recombination of foreign DNA into the chromosome, or by mutations in different chromosomal loci ([15][1]). In studies of molecular evolutionary biology, the term mutation rate

Mutation Frequencies and Antibiotic Resistance

The use of intramolecular isotope effects to distinguish between deprotonation and hydrogen atom abstraction mechanisms in cytochrome P-450- and peroxidase-catalyzed N-demethylation reactions.

(1988). The metabolic activation of nitroheterocyclic therapeutic agents. Drug Metabolism Reviews: Vol. 19, No. 1, pp. 33-62.

The metabolic activation of nitroheterocyclic therapeutic agents.

Human hepatic microsomal enzymes catalyzed the NADPH-dependent anaerobic reductive activation of [1-14C]metronidazole [1-(2-hydroxyethyl)-2-methyl-5-nitroimidazole] and [4,5-14C]ronidazole [(1-methyl-5-nitroimidazole-2-yl)methyl carbamate] to species that became covalently bound to proteins. Due to the low efficiency of the enzyme-catalyzed covalent binding of metronidazole, the stoichiometry of anaerobic reductive activation was studied with dithionite as the reductant. Two moles of dithionite was consumed per mole of [1-14C]metronidazole for maximal covalent binding to either DNA or immobilized sulfhydryl groups, demonstrating that four electrons are required for the reductive activation of metronidazole. These data implicate the involvement of a hydroxylamine in covalent binding. Maximal covalent binding of [4,5-14C]ronidazole to DNA also required four-electron reduction, consistent with previous studies of the covalent binding of this agent to immobilized sulfhydryl groups [Kedderis et al. (1988) Arch. Biochem. Biophys. 262, 40-48]. Studies of the covalent binding of variously radiolabeled ronidazole molecules to DNA suggested that the imidazole ring was intact while greater than 80% of the 2-carbamoyl group and the C4 proton were not present in the DNA adduct. Studies of both the chemical and human hepatic microsomal reduction of [4-3H]metronidazole demonstrated that covalent binding occurred with the stoichiometric loss of this label, implicating binding at the C4 position. These results suggest that the reductive activation of 5-nitroimidazoles generally proceeds via four-electron reduction to form hydroxylamines followed by nucleophilic attack at C4.

Gregory L. Kedderis

Papers

The use of intramolecular isotope effects to distinguish between deprotonation and hydrogen atom abstraction mechanisms in cytochrome P-450- and peroxidase-catalyzed N-demethylation reactions.

The metabolic activation of nitroheterocyclic therapeutic agents.

Covalent interaction of 5-nitroimidazoles with DNA and protein in vitro: mechanism of reductive activation.

Mechanisms of N-demethylation reactions catalyzed by cytochrome P-450 and peroxidases.

Mechanism of reductive activation of a 5-nitroimidazole by flavoproteins: model studies with dithionite.