Post-translational regulation of human indoleamine 2,3-dioxygenase activity by nitric oxide.
The heme protein indoleamine 2,3-dioxygenase (IDO) is induced by the proinflammatory cytokine interferon-gamma (IFNgamma) and plays an important role in the immune response by catalyzing the oxidative degradation of L-tryptophan (Trp) that contributes to immune suppression and tolerance. Here we examined the mechanism by which nitric oxide (NO) inhibits human IDO activity. Exposure of IFNgamma-stimulated human monocyte-derived macrophages (MDM) to NO donors had no material impact on IDO mRNA or protein expression, yet exposure of MDM or transfected COS-7 cells expressing active human IDO to NO donors resulted in reversible inhibition of IDO activity. NO also inhibited the activity of purified recombinant human IDO (rhIDO) in a reversible manner and this correlated with NO binding to the heme of rhIDO. Optical absorption and resonance Raman spectroscopy identified NO-inactivated rhIDO as a ferrous iron (Fe(II))-NO-Trp adduct. Stopped-flow kinetic studies revealed that NO reacted most rapidly with Fe(II) rhIDO in the presence of Trp. These findings demonstrate that NO inhibits rhIDO activity reversibly by binding to the active site heme to trap the enzyme as an inactive nitrosyl-Fe(II) enzyme adduct with Trp bound and O2 displaced. Reversible inhibition by NO may represent an important mechanism in controlling the immune regulatory actions of IDO.
Thomas
Shane R
SR
Centre for Vascular Research, Faculty of Medicine, University of New South Wales, Sydney 2052, Australia. shane.thomas@unsw.edu.au
Terentis
Andrew C
AC
Cai
Hong
H
Takikawa
Osamu
O
Levina
Aviva
A
Lay
Peter A
PA
Freewan
Mohammed
M
Stocker
Roland
R
D001665:Binding Sites; D002478:Cells, Cultured; D004789:Enzyme Activation; D006801:Humans; D050503:Indoleamine-Pyrrole 2,3,-Dioxygenase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D007700:Kinetics; D008264:Macrophages / Q000378:metabolism; D009569:Nitric Oxide / Q000494:pharmacology*; D020030:Nitric Oxide Donors / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism; D013057:Spectrum Analysis; D014364:Tryptophan / Q000378:metabolism
D050503:Indoleamine-Pyrrole 2,3,-Dioxygenase; D020030:Nitric Oxide Donors; D009569:Nitric Oxide; D014364:Tryptophan; D010100:Oxygen