Singh I, Pahan K, Dhaunsi G S, Lazo O, Ozand P
Department of Pediatrics, Medical University of South Carolina, Charleston 29425.
J Biol Chem. 1993 May 15;268(14):9972-9.
The subcellular site of oxidation of [1-14C]phytanic acid to pristanic acid and CO2 was examined by measurement of the release of 14CO2 in different organelles from human and rat tissues prepared by isopycnic density gradient centrifugation in Nycodenz. The activity of phytanic acid oxidation in human tissues (liver and cultured skin fibroblasts) paralleled that of the peroxisomal marker catalase. We also observed that Nycodenz (commonly used gradient material for isolation of subcellular organelles) has a strong inhibitory effect on the alpha-oxidation of phytanic acid. This inhibition is reversible and can be decreased or eliminated by dialysis of isolated organelles against isotonic solution. The dialysis of endoplasmic reticulum, mitochondrial, and peroxisomal fractions from human liver and cultured skin fibroblasts for 2 h against isotonic solution increased the specific activity of phytanic acid oxidation by 1.3-, 1.3-, and 5-21-fold, respectively, after removal of Nycodenz as compared with nondialyzed samples. After dialysis, the rate of oxidation of phytanic acid in peroxisomes from human liver and cultured skin fibroblasts was 4-26 times higher than that in mitochondria and 43-130 times than that in the endoplasmic reticulum, suggesting that, in human tissues, phytanic acid is oxidized to pristanic acid in peroxisomes. On the other hand, the oxidation of phytanic acid in rat liver paralleled the distribution of the mitochondrial marker cytochrome-c oxidase. The 18-fold higher rate of oxidation in dialyzed mitochondria (198.6 +/- 4.20 pmol/h/mg of protein) than in peroxisomes (11.0 +/- 0.5 pmol/h/mg of protein) demonstrates that, in rodents, phytanic acid is oxidized in mitochondria. 2-[5-(4-Chlorophenyl)pentyl]oxiran-2-carboxylic acid, an inhibitor of carnitine palmitoyltransferase I and mitochondrial fatty acid oxidation, inhibits the oxidation of phytanic acid in rat tissues (liver and cultured skin fibroblasts), whereas it has no effect on the oxidation of phytanic acid in human tissues (liver and cultured skin fibroblasts). The higher specific activity of phytanic acid oxidation in peroxisomes compared with that in mitochondria and the endoplasmic reticulum from human tissues and the inhibition of phytanic acid oxidation by 2-[5-(4-chlorophenyl)pentyl]oxiran-2-carboxylic acid in rat tissues (but not human tissues) demonstrate clearly that, in human tissues, phytanic acid is predominantly oxidized in peroxisomes.
通过测量在Nycodenz中进行等密度梯度离心制备的人和大鼠组织不同细胞器中(^{14}CO_2)的释放量，研究了([1-^{14}C])植烷酸氧化为降植烷酸和(CO_2)的亚细胞位点。人组织（肝脏和培养的皮肤成纤维细胞）中植烷酸氧化活性与过氧化物酶体标记物过氧化氢酶的活性平行。我们还观察到，Nycodenz（常用于分离亚细胞器的梯度材料）对植烷酸的α-氧化有很强的抑制作用。这种抑制是可逆的，通过将分离的细胞器用等渗溶液透析可以降低或消除。将人肝脏和培养的皮肤成纤维细胞的内质网、线粒体和过氧化物酶体组分用等渗溶液透析2小时后，与未透析的样品相比，去除Nycodenz后植烷酸氧化的比活性分别提高了1.3倍、1.3倍和5 - 21倍。透析后，人肝脏和培养的皮肤成纤维细胞过氧化物酶体中植烷酸的氧化速率比线粒体高4 - 26倍，比内质网高43 - 130倍，这表明在人体组织中，植烷酸在过氧化物酶体中氧化为降植烷酸。另一方面，大鼠肝脏中植烷酸的氧化与线粒体标记物细胞色素c氧化酶的分布平行。透析后的线粒体中氧化速率（198.6±4.20 pmol/h/mg蛋白质）比过氧化物酶体中高18倍（11.0±0.5 pmol/h/mg蛋白质），这表明在啮齿动物中，植烷酸在线粒体中氧化。2 - [5 - (4 - 氯苯基)戊基]环氧乙烷 - 2 - 羧酸是肉碱棕榈酰转移酶I和线粒体脂肪酸氧化的抑制剂，它抑制大鼠组织（肝脏和培养的皮肤成纤维细胞）中植烷酸的氧化，而对人体组织（肝脏和培养的皮肤成纤维细胞）中植烷酸的氧化没有影响。与人体组织线粒体和内质网相比，过氧化物酶体中植烷酸氧化的比活性更高，以及2 - [5 - (4 - 氯苯基)戊基]环氧乙烷 - 2 - 羧酸对大鼠组织（而非人体组织）中植烷酸氧化的抑制作用清楚地表明，在人体组织中，植烷酸主要在过氧化物酶体中氧化。
