Abstract
Two isoforms of prostaglandin H synthase have been described: isoform-1 (PGHS-1), which is ascribed a role in basal or housekeeping prostaglandin synthesis; and isoform-2 (PGHS-2), which has been found to be strongly inducible in many tissues and has been associated with inflammatory processes. Recent observations have indicated that cyclooxygenase catalysis by the two isoforms can be differentially regulated when both are present simultaneously (Reddy, S. T., and Herschman, H. R. (1994) J. Biol. Chem. 269, 15473-15480). The requirement of the cyclooxygenase for hydroperoxide initiator has been proposed as an important limit on cellular prostaglandin synthesis (Marshall, P. J., Kulmacz, R. J., and Lands, W. E. M. (1987) J. Biol. Chem. 262, 3510-3517). To compare the levels of hydroperoxide required for cyclooxygenase initiation in the two PGHS isoforms, we have examined the ability of a hydroperoxide scavenger, glutathione peroxidase, to suppress the cyclooxygenase activity of purified preparations of human PGHS-2, ovine PGHS-2, and ovine PGHS-1. Half-maximal prostaglandin synthetic activity was found to require a much lower hydroperoxide level with human PGHS-2 (2.3 nM) and ovine PGHS-2 (2.2 nM) than with ovine PGHS-1 (21 nM). Similar results were obtained when cyclooxygenase activity was monitored by chromatographic analyses of radiolabeled arachidonate metabolites or with oxygen electrode measurements. Mixing four parts of ovine PGHS-1 with one part of human PGHS-2 did not markedly change the sensitivity of the overall cyclooxygenase activity to inhibition by glutathione peroxidase, indicating that the PGHS-1 activity was not easily initiated by PGHS-2 activity in the same vessel. Effective catalysis by PGHS-2 can thus proceed at hydroperoxide levels too low to sustain appreciable catalysis by PGHS-1. This difference in catalytic characteristics provides a biochemical mechanism for differential control of prostaglandin synthesis by the two PGHS isoforms, even when both are present in the same intracellular compartment.
Highlights
Prostaglandin H synthase (PGHS)1 catalyzes a key irreversible step in the biosynthesis of prostaglandins, the oxygenation and rearrangement of arachidonic acid to form prostaglandin G2 (Samuelsson et al, 1978)
It is thought that prostaglandin synthesis; and isoform-2 (PGHS-2), which is strongly induced by various mitogens, has a role in inflammation, whereas prostaglandin H synthase have been described: isoform-1 (PGHS-1), whose cellular levels vary over a smaller range, is a housekeeping enzyme (Mitchell et al, 1993; Masferrer et al, 1994)
Immunoblotting with antibody directed at the hPGHS-2 C-terminal peptide insert stained the same 74-kDa band, confirming that the major protein component in the preparation was the inducible human PGHS isoform
Summary
Prostaglandin H synthase (PGHS)1 catalyzes a key irreversible step in the biosynthesis of prostaglandins, the oxygenation and rearrangement of arachidonic acid to form prostaglandin G2 (Samuelsson et al, 1978). Theoretical predictions of the sensitivities of the cyclooxygenase activities of oPGHS-1, hPGHS-2, and mixtures of the two enzymes to inhibition by GSP were generated with an expanded version of an earlier kinetic model (Kulmacz and Lands, 1983).
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