Oxygenation reactions catalyzed by the F557V mutant of soybean Lipoxygenase-1: Evidence for two orientations of substrate binding.

Oxygenation reactions catalyzed by the F557V mutant of soybean Lipoxygenase-1: Evidence for two orientations of substrate binding. Arch Biochem Biophys. 2019 Aug 29;:108082 Authors: Hershelman D, Kahler KM, Price MJ, Lu I, Fu Y, Plumeri PA, Karaisz F, Bassett NF, Findeis PM, Clapp CH Abstract Plant lipoxygenases oxygenate linoleic acid to produce 13(S)-hydroperoxy-9Z,11E-octadecadienoic acid (13(S)-HPOD) or 9-hydroperoxy-10Z,12E-octadecadienoic acid (9(S)-HPOD). The manner in which these enzymes bind substrates and the mechanisms by which they control regiospecificity are uncertain. Hornung et al. (Proc. Natl. Acad. Sci. USA96 (1999) 4192-4197) have identified an important residue, corresponding to phe-557 in soybean lipoxygenase-1 (SBLO-1). These authors proposed that large residues in this position favored binding of linoleate with the carboxylate group near the surface of the enzyme (tail-first binding), resulting in formation of 13(S)-HPOD. They also proposed that smaller residues in this position facilitate binding of linoleate in a head-first manner with its carboxylate group interacting with a conserved arginine residue (arg-707 in SBLO-1), which leads to 9(S)-HPOD. In the present work, we have tested these proposals on SBLO-1. The F557V mutant produced 33% 9-HPOD (S:R = 87:13) from linoleic acid at pH 7.5, compared with 8% for the wild-type enzyme and 12% with the F557V, R707L double mutant. Experiments with 11(S)-deuteri...
Source: Archives of Biochemistry and Biophysics - Category: Biochemistry Authors: Tags: Arch Biochem Biophys Source Type: research