Corpas F J, Barroso J B, Sandalio L M, Distefano S, Palma J M, Lupiáñez J A, Del Río L A
Departamento de Bioquímica, Biología Celular y Molecular de Plantas, Estación Experimental del Zaidín, CSIC, Apartado 419, E-18080 Granada, Spain.
Biochem J. 1998 Mar 1;330 ( Pt 2)(Pt 2):777-84. doi: 10.1042/bj3300777.
The presence of the two NADP-dependent dehydrogenases of the pentose phosphate pathway has been investigated in plant peroxisomes from pea (Pisum sativum L.) leaves. Both enzymes, glucose-6-phosphate dehydrogenase (G6PDH; EC 1.1.1.49) and 6-phosphogluconate dehydrogenase (6PGDH; EC 1.1.1.44), were present in the matrix of leaf peroxisomes, and their kinetic properties were studied. G6PDH and 6PGDH showed a typical Michaelis-Menten kinetic saturation curve, and had specific activities of 12.4 and 29.6 mU/mg protein, respectively. The Km values of G6PDH and 6PGDH for glucose 6-phosphate and for 6-phosphogluconate were 107.3 and 10.2 microM, respectively. Dithiothreitol did not inhibit G6PDH activity. By isoelectric focusing of peroxisomal matrices, the G6PDH activity was resolved into three isoforms with isoelectric points of 5.55, 5.30 and 4.85. The isoelectric point of peroxisomal 6PGDH was 5.10. Immunoblot analyses of peroxisomal matrix with an antibody against yeast G6PDH revealed a single cross-reactive band of 56 kDa. Post-embedment, EM immunogold labelling of G6PDH confirmed that this enzyme was localized in the peroxisomal matrices, the thylakoid membrane and matrix of chloroplasts, and the cytosol. The presence of the two oxidative enzymes of the pentose phosphate pathway in plant peroxisomes implies that these organelles have the capacity to reduce NADP+ to NADPH for its re-utilization in the peroxisomal metabolism. NADPH is particularly required for the ascorbate-glutathione cycle, which has been recently demonstrated in plant peroxisomes [Jiménez, Hernández, del Río and Sevilla (1997) Plant Physiol. 114, 275-284] and represents an important antioxidant protection system against H2O2 generated in peroxisomes.
已对豌豆（Pisum sativum L.）叶片的植物过氧化物酶体中戊糖磷酸途径的两种依赖NADP的脱氢酶的存在情况进行了研究。葡萄糖-6-磷酸脱氢酶（G6PDH；EC 1.1.1.49）和6-磷酸葡萄糖酸脱氢酶（6PGDH；EC 1.1.1.44）这两种酶均存在于叶片过氧化物酶体的基质中，并对它们的动力学特性进行了研究。G6PDH和6PGDH呈现典型的米氏动力学饱和曲线，其比活性分别为12.4和29.6 mU/mg蛋白质。G6PDH和6PGDH对葡萄糖6-磷酸和6-磷酸葡萄糖酸的Km值分别为107.3和10.2 microM。二硫苏糖醇不抑制G6PDH活性。通过对过氧化物酶体基质进行等电聚焦，G6PDH活性被解析为三种等电点分别为5.55、5.30和4.85的同工型。过氧化物酶体6PGDH的等电点为5.10。用抗酵母G6PDH抗体对过氧化物酶体基质进行免疫印迹分析，显示出一条56 kDa的单一交叉反应带。包埋后，G6PDH的电镜免疫金标记证实该酶定位于过氧化物酶体基质、叶绿体类囊体膜和基质以及细胞质溶胶中。植物过氧化物酶体中戊糖磷酸途径的两种氧化酶的存在意味着这些细胞器有能力将NADP+还原为NADPH，以供其在过氧化物酶体代谢中重新利用。NADPH对于抗坏血酸-谷胱甘肽循环尤为必需，最近已在植物过氧化物酶体中得到证实[Jiménez, Hernández, del Río和Sevilla（1997年）Plant Physiol. 114, 275 - 284]，并且代表了一种针对过氧化物酶体中产生的H2O2的重要抗氧化保护系统。
