Baldet P, Alban C, Axiotis S, Douce R
Unité mixte C.N.R.S./Rhône-Poulenc (Unité Associée au Centre National de la Recherche Scientifique, U.M.R. 41), Lyon, France.
Arch Biochem Biophys. 1993 May 15;303(1):67-73. doi: 10.1006/abbi.1993.1256.
Cytosol and vacuoles from protoplasts, chloroplasts, and mitochondria from green pea (Pisum sativum) leaves were purified and examined for their biotin content. The bulk of free biotin was shown to be exclusively associated with the cytosolic fraction at a concentration of about 4 pmol/mg protein and no bound biotin was detected. The bulk of bound biotin (biotin-containing carboxylases) was associated with the soluble fraction of chloroplasts and mitochondria at a concentration of about 1.2 and 13 microM, respectively. No free biotin was detected in these organelles. Western blot analysis of total, chloroplastic, and mitochondrial polypeptides, using horseradish peroxidase-labeled streptavidin, revealed three biotin-containing polypeptides with molecular mass of 220,000, 76,000 and 34,000. All were detected in the total pea leaf extract, but the M(r) 76,000 and the M(r) 34,000 biotinylated polypeptides were only detected in mitochondria and chloroplasts, respectively. 3-Methylcrotonyl-coenzyme A carboxylase and acetyl-coenzyme A carboxylase activities were measured in these two compartments, respectively. Previously, it has been shown that the M(r) 76,000 polypeptide was the biotinylated subunit of the mitochondrial 3-methylcrotonyl-coenzyme A carboxylase. In this paper, the origin and putative function of free biotin located in cytosol are discussed.
对来自绿色豌豆（Pisum sativum）叶片的原生质体的胞质溶胶和液泡、叶绿体和线粒体进行了纯化，并检测了它们的生物素含量。结果表明，大部分游离生物素仅与胞质溶胶部分相关，浓度约为4 pmol/mg蛋白质，未检测到结合生物素。大部分结合生物素（含生物素的羧化酶）分别以约1.2和13 microM的浓度与叶绿体和线粒体的可溶部分相关。在这些细胞器中未检测到游离生物素。使用辣根过氧化物酶标记的链霉亲和素对总多肽、叶绿体多肽和线粒体多肽进行蛋白质免疫印迹分析，发现了三种分子量分别为220,000、76,000和34,000的含生物素多肽。在豌豆叶片总提取物中均检测到了这三种多肽，但分子量为76,000和34,000的生物素化多肽分别仅在线粒体和叶绿体中检测到。分别在这两个区室中测量了3-甲基巴豆酰辅酶A羧化酶和乙酰辅酶A羧化酶的活性。此前已表明，分子量为76,000的多肽是线粒体3-甲基巴豆酰辅酶A羧化酶的生物素化亚基。本文讨论了位于胞质溶胶中的游离生物素的来源和假定功能。
