Prostaglandin G2 levels during reaction of prostaglandin H synthase with arachidonic acid

doi:10.1016/0090-6980(87)90246-2

Prostaglandins

Volume 34, Issue 2, August 1987, Pages 225-240

Prostaglandins

https://doi.org/10.1016/0090-6980(87)90246-2 Get rights and content

Abstract

Prostaglandin H synthase catalyzes the formation of prostaglandin (PG) G₂ from arachidonic acid (cyclooxygenase activity), and also the reduction of PGG₂ to PGH₂ (peroxidase activity). The ability of the pure synthase to accumulate the hydroperoxide, PGG₂, under conditions allowing the concurrent function of both catalytic activities was investigated. The peroxidase velocity was continuously determined from the absorbance increases at 611 nm that accompanied oxidation of a peroxidase cosubstrate, N,N,N′N′-tetramethyl-phenylenediamine, and PGG₂ concentrations were calculated from the peroxidase velocities and the peroxidase V_max and K_m values. Cyclooxygenase velocities were than calculated from the changes in PGG₂. Parallel reactions monitored by the use of radiolabelled arachidonate or with a polarographic oxygen electrode were used to confirm the calculated PGG₂ levels and the cyclooxygenase velocities. The concentration of PGG₂ was found to follow a transient course as the reaction of the synthase progressed, rapidly rising to a maximum of 0.7μM in the first 10 s, and then declining slowly, reaching 0.1 μM after 60 s. The maximal level of PGG₂ achieved during the reaction was constant at about 0.7 μM with higher amounts of added cyclooxygenase capacity (0.3–0.6 μM PGG₂/s) but was only about 0.4 μM when the added cyclooxygenase capacity was 0.1 μM PGG₂/s. The peroxidase was found to lose only 30% of its activity after 90 s, a point where the cyclooxygenase was almost completely inactive. These results support the concept of a burst of catalytic action from the cyclooxygenase and a reactive, more sustained, catalytic action from the peroxidase during the reaction of the synthase with arachidonic acid.

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Prostaglandin G2 levels during reaction of prostaglandin H synthase with arachidonic acid

Abstract

J. Biol. Chem.

J. Biol. Chem.

Arch. Biochem. Biophys.

Biochim. Biophys. Acta

J. Biol. Chem.

Prostaglandins

Arch. Biochem. Biophys.

Prostaglandins

J. Biol. Chem.

Prostaglandins

J. Biol. Chem.

J. Biol. Chem.

Prostaglandin G₂ levels during reaction of prostaglandin H synthase with arachidonic acid