Hayashi M, De Bellis L, Alpi A, Nishimura M
Department of Cell Biology, National Institute for Basic Biology, Okazaki, Japan.
Plant Cell Physiol. 1995 Jun;36(4):669-80.
Two different aconitases are known to be expressed after the germination of oil-seed plants. One is a mitochondrial aconitase that is involved in the tricarboxylic acid cycle. The other participates in the glyoxylate cycle, playing a role in gluconeogenesis from stored oil. We isolated and characterized a cDNA for an aconitase from etiolated pumpkin cotyledons. The cDNA was 3,145 bp long and capable of encoding a protein of 98 kDa. N-terminal and C-terminal amino acid sequences deduced from the cDNA did not contain mitochondrial or glyoxysomal targeting signals. A search of protein databases suggested that the cDNA encoded a cytosolic aconitase. Immunoblotting analysis with a specific antibody against the aconitase expressed in Escherichia coli revealed that developmental changes in the amount of the aconitase were correlated with changes in levels of other enzymes of the glyoxylate cycle during growth of seedlings. Further analysis by subcellular fractionation and immunofluorescence microscopy revealed that aconitase was present only the cytosol and mitochondria. No glyoxysomal aconitase was found in etiolated cotyledons even though all the other enzymes of the glyoxylate cycle are known to be localized in glyoxysomes. Taken together, the data suggest that the cytosolic aconitase participates in the glyoxylate cycle with four glyoxysomal enzymes.
已知在油料植物种子萌发后会表达两种不同的乌头酸酶。一种是参与三羧酸循环的线粒体乌头酸酶。另一种参与乙醛酸循环，在从储存的油脂进行糖异生过程中发挥作用。我们从黄化南瓜子叶中分离并鉴定了一种乌头酸酶的cDNA。该cDNA长3145 bp，能够编码一种98 kDa的蛋白质。从该cDNA推导的N端和C端氨基酸序列不包含线粒体或乙醛酸体靶向信号。对蛋白质数据库的搜索表明，该cDNA编码一种胞质乌头酸酶。用针对在大肠杆菌中表达的乌头酸酶的特异性抗体进行免疫印迹分析表明，在幼苗生长过程中，乌头酸酶量的发育变化与乙醛酸循环中其他酶水平的变化相关。通过亚细胞分级分离和免疫荧光显微镜进一步分析表明，乌头酸酶仅存在于细胞质和线粒体中。尽管已知乙醛酸循环的所有其他酶都定位于乙醛酸体中，但在黄化子叶中未发现乙醛酸体乌头酸酶。综上所述，数据表明胞质乌头酸酶与四种乙醛酸体酶一起参与乙醛酸循环。
