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Use of inhibitors in elucidating the mechanism of an enzyme

In this review paper theoretical insights obtained through studies of non-heme iron-containing enzymes are discussed.In particular, research on oxygen-utilizing enzymes with a 2-His/1-Asp or 3-His structural motif is the focus of this work with recent examples from studies of cysteine dioxygenase (CDO) and taurine/α-ketoglutarate dioxygenase (Tau D) enzymes.Thus, theoretical modeling can be extremely useful in the characterization of short-lived intermediates which either cannot be detected experimentally or have a lifetime that is too short to enable spectroscopic characterization.

CDO uses molecular oxygen and transfers both oxygen atoms to cysteine to form cysteine sulfinic acid products.

By contrast, in Tau D the first oxygen atom of O is donated to α-ketoglutarate to give succinate, carbon dioxide and an oxo–iron active species.

The latter is the oxidant that is able to abstract a hydrogen atom from a substrate and rebounds the hydroxyl group to form hydroxylated products.

The theoretical (density functional theory (DFT)) studies show that the first step in this dioxygen activation process is the rate-determining step and that the reactions leading to products are highly exothermic as would be expected from an efficient catalytic cycle of an enzyme.

Furthermore, theory predicts Tau D to be a much more aggressive and efficient catalyst then for instance monoxygenase enzymes such as the cytochromes P450 (P450s).

Non-heme enzymes generally react with substrates via single-state reactivity (SSR) patterns on a dominant quintet spin-state surface.

Theory has found no evidence of possible alternative oxidants in the reaction mechanisms.

The theoretical studies give fundamental insight into processes that proceed too fast to be experimentally studied and give answers to questions regarding the efficiency and nature of the oxidizing species of this fascinating class of enzymes.

-alkoxyresorufin dealkylation assays since the 1990s, thousands of inhibitors of cytochrome P450 family 1 enzymes (P450s 1A1, 1A2, and 1B1) have been identified and studied.

Generally, planar polycyclic molecules such as polycyclic aromatic hydrocarbons, stilbenoids, and flavonoids are considered to potentially be effective inhibitors of these enzymes, however, the details of the structure-activity relationships and selectivity of these inhibitors are still ambiguous.

Comments Use of inhibitors in elucidating the mechanism of an enzyme