TY - JOUR
T1 - Decreased protein nitration in macrophages that overexpress indoleamine 2, 3-dioxygenase
AU - Keskin, Derin B.
AU - Marshall, Brendan
AU - Munn, David
AU - Mellor, Andrew L.
AU - Gearhart, Debra A.
PY - 2007
Y1 - 2007
N2 - The activity of indoleamine 2, 3-dioxygenase (IDO; E.C. 1.13.11.42) catalyzes the oxidative cleavage of tryptophan to form kynurenine. IDO activity consumes superoxide anions; therefore, we postulated that over-expression of IDO might mitigate superoxide-anion dependent, oxidative modification of cellular proteins in vitro. We prepared and characterized RAW 264.7 macrophages that were stably transfected with either an IDO expression vector or the control (empty) vector. We detected IDO mRNA, protein, and enzyme activity in the IDO-transfected macrophages, but not in the macrophages transfected with the empty vector. To generate superoxide anions in situ, we treated the IDO- and control-transfected cultures with xanthine or hypoxanthine, and then used ELISA methods to quantitate the relative levels of oxidatively modified proteins in total cell lysates. The levels of protein carbonyls were similar in IDO-transfected and vector-transfected macrophages; however, protein nitration was significantly less in IDO-transfected cells compared to control transfectants. In addition, steady-state levels of superoxide anions were significantly lower in the IDO-transfected cultures compared with control transfectants. Our results are consistent with the concept that, besides degrading tryptophan, IDO activity may protect cells from oxidative damage.
AB - The activity of indoleamine 2, 3-dioxygenase (IDO; E.C. 1.13.11.42) catalyzes the oxidative cleavage of tryptophan to form kynurenine. IDO activity consumes superoxide anions; therefore, we postulated that over-expression of IDO might mitigate superoxide-anion dependent, oxidative modification of cellular proteins in vitro. We prepared and characterized RAW 264.7 macrophages that were stably transfected with either an IDO expression vector or the control (empty) vector. We detected IDO mRNA, protein, and enzyme activity in the IDO-transfected macrophages, but not in the macrophages transfected with the empty vector. To generate superoxide anions in situ, we treated the IDO- and control-transfected cultures with xanthine or hypoxanthine, and then used ELISA methods to quantitate the relative levels of oxidatively modified proteins in total cell lysates. The levels of protein carbonyls were similar in IDO-transfected and vector-transfected macrophages; however, protein nitration was significantly less in IDO-transfected cells compared to control transfectants. In addition, steady-state levels of superoxide anions were significantly lower in the IDO-transfected cultures compared with control transfectants. Our results are consistent with the concept that, besides degrading tryptophan, IDO activity may protect cells from oxidative damage.
KW - Indoleamine 2, 3-dioxygenase
KW - Oxidative stress
KW - Protein oxidation
KW - RAW 264.7 macrophages
KW - Superoxide anion
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U2 - 10.2478/s11658-006-0048-9
DO - 10.2478/s11658-006-0048-9
M3 - Article
C2 - 17103092
AN - SCOPUS:34249003035
SN - 1425-8153
VL - 12
SP - 82
EP - 102
JO - Cellular and Molecular Biology Letters
JF - Cellular and Molecular Biology Letters
IS - 1
ER -