Investigation of sulfur containing amino acids at the lipoxygenase active site using a platinum complex.
Inactivation of native soybean lipoxygenase-1 was observed upon preincubation with (NEt4)[PtCl3(P(Bun)3)]. Removal of the platinum complex(es) from the inactivated enzyme by treatment with sodium diethyldithiocarbamate (Naddtc) which reverses methionine but not cysteine binding, restores most of the activity. Linoleic acid, an enzyme substrate, protects it from inactivation. The quenching of the fluorescence of the putative active site tryptophans which accompanies inactivation disappears after Naddtc reactivation. The (NEt4)[PtCl3(P(Bun)3)]-inactivated enzyme iron(II) cannot be oxidized at variance with that of the native or Naddtc reactivated enzyme, as checked by EPR spectroscopy. These results show that at least one methionine is close to the iron binding site in soybean lipoxygenase-1.
Michaud-Soret
I
I
Laboratoire de Chimie et Biochimie Pharmacologiques et Toxicologiques, Université René Descartes, Paris, France.
Chottard
J C
JC
D001665:Binding Sites; D003545:Cysteine; D004050:Ditiocarb / Q000494:pharmacology*; D004578:Electron Spin Resonance Spectroscopy; D007700:Kinetics; D008084:Lipoxygenase / Q000378:metabolism*; D016859:Lipoxygenase Inhibitors / Q000494:pharmacology*; D008715:Methionine; D009944:Organoplatinum Compounds / Q000138:chemical synthesis / Q000494:pharmacology*; D011487:Protein Conformation; D012639:Seeds / Q000201:enzymology; D013025:Glycine max / Q000201:enzymology; D013050:Spectrometry, Fluorescence; D014364:Tryptophan
D016859:Lipoxygenase Inhibitors; D009944:Organoplatinum Compounds; D014364:Tryptophan; D004050:Ditiocarb; D008715:Methionine; D008084:Lipoxygenase; D003545:Cysteine