Abstract
While designing the anti-inflammatory agents targeting cyclooxygenase-2 (COX-2), we first identified a water loop around the heme playing critical role in the enzyme catalysis. The results of molecular dynamic studies supported by the strong hydrogen-bonding equilibria of the participating atoms, radical stabilization energies, the pKa of the H-donor/acceptor sites and the cyclooxygenase activity of pertinent muCOX-2 ravelled the working of the water–peptide channel for coordinating the flow of H·/electron between the heme and Y385. Based on the working of H·/electron transfer channel between the 12.5 Å distant radical generation and the radical disposal sites, a series of molecules was designed and synthesized. Among this category of compounds, an appreciably potent anti-inflammatory agent exhibiting IC50 0.06 μM against COX-2 and reversing the formalin induced analgesia and carageenan induced inflammation in mice by 90% was identified. Further it was revealed that, justifying its bidentate design, the compound targets water loop (heme bound site) and the arachidonic acid binding pockets of COX-2.
Highlights
While designing the anti-inflammatory agents targeting cyclooxygenase-2 (COX-2), we first identified a water loop around the heme playing critical role in the enzyme catalysis
The limitation of the static crystal structure, not truly representing the reactive coordinates of the enzyme, incited us to investigate the dynamics of COX-2—arachidonic acid (AA)—heme complex if somehow the conformational changes bring ferryl heme and Y385 close to each other
The H transfer processes are difficult to monitor in proteins but the variety of computer aided simulations tagged with the experimental evidences provided enough proof in favour of proton coupled electron transfer in cyclooxygenase
Summary
While designing the anti-inflammatory agents targeting cyclooxygenase-2 (COX-2), we first identified a water loop around the heme playing critical role in the enzyme catalysis. Based on the working of H·/electron transfer channel between the 12.5 Å distant radical generation and the radical disposal sites, a series of molecules was designed and synthesized. Among this category of compounds, an appreciably potent anti-inflammatory agent exhibiting IC50 0.06 μM against COX-2 and reversing the formalin induced analgesia and carageenan induced inflammation in mice by 90% was identified. Bearing in mind the radical mechanism of AA m etabolism[23], here, we first explored the channel for the radical transfer from its generation site heme to the disposal site Y385 (Fig. 1C) and tried to disturb this channel (in addition to the arachidonic acid binding site) for developing the anti-inflammatory agents
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