Lucas M, Solano F
Departamento de Bioquímica y Biología Molecular, Hospital Universitario Virgen Macarena, Facultad de Medicina, Sevilla, Spain.
Anal Biochem. 1992 Nov 1;206(2):273-7. doi: 10.1016/0003-2697(92)90366-f.
The oxidation of free coelenterazine by superoxide anion was analyzed and compared to the oxidation by the semisynthetic photoprotein obelin, prepared by incorporation of synthetic coelenterazine into apoobelin. The oxidation of bound coelenterazine was triggered upon binding of calcium to the reconstituted photoprotein. The oxidation of free synthetic coelenterazine, in the absence of the apoprotein, was triggered by superoxide anion. The production of reactive oxygen metabolites by fMet-Leu-Phe- and 4b-phorbol 12b-myristate 13a-acetate-stimulated neutrophils was studied by means of the luminescence of synthetic coelenterazine. The features of this chemiluminescent probe were compared with those of luminol and are summarized as follows: (a) coelenterazine-dependent chemiluminescence was inhibited by superoxide dismutase; (b) coelenterazine was as sensitive as luminol in detecting the oxidative burst of neutrophils; (c) azide failed to inhibit coelenterazine chemiluminescence; (d) in contrast with luminol, which requires the catalytic removal of hydrogen peroxide, coelenterazine chemiluminescence did not depend on the activity of cell-derived myeloperoxidase. These results indicate the usefulness of coelenterazine as a very sensitive and specific chemiluminescence probe of superoxide anion.
分析了超氧阴离子对游离腔肠素的氧化作用，并将其与通过将合成腔肠素掺入脱辅基腔肠素制备的半合成光蛋白腔肠萤光素的氧化作用进行了比较。钙与重组光蛋白结合后引发结合态腔肠素的氧化。在没有脱辅基蛋白的情况下，超氧阴离子引发游离合成腔肠素的氧化。通过合成腔肠素的发光研究了甲酰甲硫氨酰-亮氨酰-苯丙氨酸和4β-佛波醇12β-肉豆蔻酸酯13α-乙酸酯刺激的中性粒细胞产生活性氧代谢产物的情况。将这种化学发光探针的特性与鲁米诺的特性进行了比较，总结如下：(a)超氧化物歧化酶抑制腔肠素依赖性化学发光；(b)在检测中性粒细胞的氧化爆发方面，腔肠素与鲁米诺一样敏感；(c)叠氮化物不能抑制腔肠素化学发光；(d)与需要催化去除过氧化氢的鲁米诺不同，腔肠素化学发光不依赖于细胞来源的髓过氧化物酶的活性。这些结果表明腔肠素作为超氧阴离子非常敏感和特异的化学发光探针是有用的。
