Reduction of nitrite to nitric oxide catalyzed by xanthine oxidoreductase
Reduction of nitrite to nitric oxide catalyzed by xanthine oxidoreductase
复制标题
DOI：
10.1074/jbc.275.11.7757
复制
发表时间：
2000-03-17
影响因子：
4.8
通讯作者：
Harrison, R
中科院分区：
文献类型：
--
作者：
Godber, BLJ;Doel, JJ;Harrison, R
黄嘌呤氧化酶（XO）在NADH或黄嘌呤作为还原性底物存在的厌氧条件下催化亚硝酸盐还原为一氧化氮（NO）。臭氧化学发光直接证明了NO的产生，并显示出与NADH消耗的化学计量比约为2:1。当黄嘌呤作为还原性底物时，由于NO诱导XO转化为相对无活性的脱硫形式，导致酶失活，使NO生成动力学变得复杂。采用分光光度法测定了在厌氧条件下，由XO和黄嘌呤脱氢酶在亚硝酸盐存在下催化尿酸生成和NADH氧化的稳态动力学参数。在亚硝酸盐存在下，XO催化厌氧NO生成的最佳pH值为NADH 7.0，黄嘌呤小于等于6.0。异黄嘌呤与亚硝酸盐竞争性地抑制黄嘌呤氧化，表明了XO的钼位点参与亚硝酸盐还原。脱硫酶的nadh -亚硝酸盐还原酶活性相对较低，表明了对Mo=S的强烈偏好。结果表明，在黄嘌呤存在的情况下，XO的FAD位点不影响亚硝酸盐的还原，但当NADH作为还原底物时，它明显参与了亚硝酸盐的还原。NO的表观产量随着氧张力的增加而减少，这与NO与XO生成的超氧化物的反应一致，这表明XO衍生的NO在消化道中具有杀菌作用。
Xanthine oxidase (XO) was shown to catalyze the reduction of nitrite to nitric oxide (NO), under anaerobic conditions, in the presence of either NADH or xanthine as reducing substrate. NO production was directly demonstrated by ozone chemiluminescence and showed stoichiometry of approximately 2:1 versus NADH depletion. With xanthine as reducing substrate, the kinetics of NO production were complicated by enzyme inactivation, resulting from NO-induced conversion of XO to its relatively inactive desulfo-form. Steady-state kinetic parameters were determined spectrophotametrically for urate production and NADH oxidation catalyzed by XO and xanthine dehydrogenase in the presence of nitrite under anaerobic conditions. pH optima for anaerobic NO production catalyzed by XO in the presence of nitrite were 7.0 for NADH and less than or equal to 6.0 for xanthine. Involvement of the molybdenum site of XO in nitrite reduction was shown by the fact that alloxanthine inhibits xanthine oxidation competitively with nitrite. Strong preference for Mo=S over Mo=O was shown by the relatively very low NADH-nitrite reductase activity shown by desulfo-enzyme. The FAD site of XO was shown not to influence nitrite reduction in the presence of xanthine, although it was clearly involved when NADH was the reducing substrate, Apparent production of NO decreased with increasing oxygen tensions, consistent with reaction of NO with XO-generated superoxide, It is proposed that XO-derived NO fulfills a bactericidal role in the digestive tract.