Inhibition of indoleamine 2,3 dioxygenase activity by H2O2.
Indoleamine 2,3-dioxygenase is the first and rate limiting enzyme of the kynurenine pathway of tryptophan metabolism, has potent effects on cell proliferation and mediates antimicrobial, antitumorogenic, and immunosuppressive effects. As a potent cytotoxic effector, the mechanisms of indoleamine 2,3-dioxygenase inhibition deserve greater attention. The work presented here represents the first systematic study exploring the mechanisms by which low levels of hydrogen peroxide (10-100 microM) inhibit indoleamine 2,3-dioxygenase in vitro. Following brief peroxide exposure both enzyme inhibition and structural changes were observed. Loss of catalysis was accompanied by oxidation of several cysteine residues to sulfinic and sulfonic acids, observed by electrospray and MALDI mass spectrometry. Enzyme activity could in part be preserved in the presence of sulfhydryl containing compounds, particularly DTT and methionine. However, these structural alterations did not prevent substrate (l-tryptophan) binding. Some enzyme activity could be recovered in the presence of thioredoxin, indicating that the inhibitory effect of H(2)O(2) is at least partially reversible in vitro. We present evidence that cysteine oxidation represents one mechanism of indoleamine 2,3-dioxygenase inhibition.
Poljak
Anne
A
Bioanalytical Mass Spectrometry Facility, University of New South Wales, Sydney, Australia. a.poljak@unsw.edu.au
Grant
Ross
R
Austin
Chris J D
CJ
Jamie
Joanne F
JF
Willows
Robert D
RD
Takikawa
Osamu
O
Littlejohn
Tamantha K
TK
Truscott
Roger J W
RJ
Walker
Mark J
MJ
Sachdev
Perminder
P
Smythe
George A
GA
D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D007108:Immune Tolerance / Q000502:physiology; D050503:Indoleamine-Pyrrole 2,3,-Dioxygenase / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D009369:Neoplasms / Q000201:enzymology / Q000276:immunology; D010084:Oxidation-Reduction / Q000187:drug effects; D011485:Protein Binding / Q000187:drug effects; D017434:Protein Structure, Tertiary / Q000187:drug effects; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013379:Substrate Specificity / Q000187:drug effects; D013438:Sulfhydryl Compounds / Q000737:chemistry; D014364:Tryptophan / Q000737:chemistry / Q000276:immunology / Q000378:metabolism
D004791:Enzyme Inhibitors; D050503:Indoleamine-Pyrrole 2,3,-Dioxygenase; D013438:Sulfhydryl Compounds; D014364:Tryptophan; D006861:Hydrogen Peroxide