Alteration of human leukotriene A(4) hydrolase activity after site-directed mutagenesis: serine-415 is a regulatory residue

Leukotriene A(4) hydrolase (LTA-H) is a bifunctional protein that has amino peptidase activity, but also contains an epoxide hydrolase activity that co nverts leukotriene (LT)A(4) to LTB4. The lipid metabolic activity of this e nzyme plays a central role in the control of polymorphonuclear leukocyte fu nction and in the development of inflammation. LTA-H is widely spread in ma ny mammalian tissues, although it appears to be inactive in many cases. Reg ulation of this enzyme's activity by phosyhorylation of a serine at residue 415 has recently been described. Since the activation of LTA-H in the pres ence of activated PMNL would likely lead to a substantial increase in the p roduction of inflammatory lipids, regulation of LTA-H presents a novel pote ntial target for anti-inflammatory therapy. We have now made a series of si te-directed mutants at this site to test the importance of this residue to the activity of LTA-H. Replacement of the critical serine with threonine or glutamine has little effect on either the epoxide hydrolase or aminopeptid ase activities. However, replacing serine with a negatively charged amino a cid (either aspartate or glutamate), intended to mimic phosphorylation at t hat site, causes significant reduction in epoxide hydrolase activity (50-70 %). These mutations have little effect on the aminopeptidase activity of th e LTA-H, suggesting that the mutation models the regulatory event and is no t simply due to improper folding of the protein. (C) 1994 Elsevier Science B.V. All rights reserved.