Prostaglandin H synthase-catalyzed oxidation of all-trans- and 13-cis-retinoic acid to carbon-centered and peroxyl radical intermediates.
Prostaglandin H synthase-catalyzed oxidation of all-trans- and 13-cis-retinoic acid to carbon-centered and peroxyl radical intermediates.
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DOI：
10.1021/tx960027p
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发表时间：
1996
影响因子：
4.1
通讯作者：
M. A. Freyaldenhoven;R. Lloyd;V. Samokyszyn
中科院分区：
文献类型：
--
作者：
M. A. Freyaldenhoven;R. Lloyd;V. Samokyszyn
由于全反式维甲酸（RA）在治疗各种皮肤病中的重要性以及前列腺素H合酶（PGHS）在组织中的广泛分布，我们进一步研究了PGHS对RA及其异构体13-顺式维甲酸（（13 Z）-RA）的过氧化氢依赖性共氧化的机制。使用电子自旋共振（ESR）自旋捕获技术和5-苯基-4-戊烯-1-基过氧化氢（PPHP）或13-氢过氧基-9-顺式-11-反式十八碳二烯酸（13-OOH-18：2）作为过氧化氢底物，表明RA和（13 Z）-RA的过氧化氢依赖性PGHS催化氧化可形成自由基加合物。利用α-苯基-N-叔丁基硝酮（PBN）的自旋捕获，检测到（13 Z）-RA-PBN和RA-PBN加合物，其光谱特征分别为aH = 4.16/aN = 15.69和aH = 3.01/aN =15.92。在厌氧条件下使用自旋捕获器2-甲基-2-亚硝基丙烷（NtB）进行相同的实验，其产生氮氧加合物，其光谱特征在于具有（13 Z）-RA加合物的aH = 3.49/aN = 15.84和RA加合物的aH = 3.49/aN = 15.88的双重峰的三重峰。这些结果表明二次碳中心自由基的形成。我们还用（+）-苯并[a]芘7（S），8（S）-二氢二醇（（+）-BP-7，8-二醇）作为过氧自由基探针。结果表明形成了（+）-BP-7，8-二醇衍生的四醇，其中反式-反式四醇代表了（13 Z）-RA或RA发生PPHP依赖性、PGHS催化氧化的系统中的主要氧化产物。这些结果与这些体系中过氧自由基的形成是一致的。在所有实验中，与全反式异构体相比，（13 Z）-RA异构体似乎是酶的更好底物。总的来说，这些结果提供了进一步的证据，支持先前提出的类维生素A氧化的PGHS涉及C4碳中心的自由基，随后与分子氧反应，产生类维生素A衍生的过氧自由基的中间机制。
Due to the importance of all-trans-retinoic acid (RA) in the treatment of various dermatological conditions and the wide distribution of prostaglandin H synthase (PGHS) in tissues, we have further examined the mechanisms involved in the hydroperoxide-dependent cooxidation of RA and its isomer, 13-cis-retinoic acid ((13Z)-RA), by PGHS. Hydroperoxide-dependent, PGHS-catalyzed oxidation of RA and (13Z)-RA was shown to form free radical adducts, using electron spin resonance (ESR) spin trapping techniques and 5-phenyl-4-penten-1-yl hydroperoxide (PPHP) or 13-hydroperoxy-9-cis-11-trans-octadecadienoic acid (13-OOH-18:2) as hydroperoxide substrates. Utilization of the spin trap alpha-phenyl-N-tert-butylnitrone (PBN) resulted in the detection of (13Z)-RA-PBN and RA-PBN adducts whose spectra were characterized by hyperfine coupling constants of aH = 4.16/aN = 15.69 and aH = 3.01/aN =15.92, respectively. Identical experiments under anaerobic conditions were carried out using the spin trap 2-methyl-2-nitrosopropane (NtB) which yielded nitroxide adducts whose spectra were characterized by a triplet of doublets with values of aH = 3.49/aN = 15.84 for the (13Z)-RA adduct and aH = 3.49/aN = 15.88 for the RA adduct. These results are indicative of secondary carbon-centered radical formation. We also used (+)-benzo[a]pyrene 7(S),8(S)-dihydrodiol ((+)-BP-7,8-diol) as a peroxyl radical probe. The results demonstrated the formation of (+)-BP-7,8-diol-derived tetrols, with the trans-anti tetrol representing the major oxidation product in systems undergoing PPHP-dependent, PGHS-catalyzed oxidation of (13Z)-RA or RA. These results are consistent with the formation of peroxyl radicals in these systems. In all experiments, the (13Z)-RA isomer appeared to be a better substrate for the enzyme compared to the all-trans isomer. Collectively these results provide further evidence to support the previously proposed mechanism for retinoid oxidation by PGHS involving the intermediacy of C4 carbon-centered radicals which subsequently react with dioxygen, yielding retinoid-derived peroxyl radicals.