Yu Y, Mu H H, Wasserman B P, Carman G M
Department of Food Science, Cook College, New Jersey Agricultural Experiment Station, Rutgers University, 65 Dudley Road, New Brunswick, New Jersey 08901, USA.
Plant Physiol. 2001 Jan;125(1):351-9. doi: 10.1104/pp.125.1.351.
Amyloplast is the site of starch synthesis in the storage tissue of maize (Zea mays). The amyloplast stroma contains an enriched group of proteins when compared with the whole endosperm. Proteins with molecular masses of 76 and 85 kD have been identified as starch synthase I and starch branching enzyme IIb, respectively. A 112-kD protein was isolated from the stromal fraction by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and subjected to tryptic digestion and amino acid sequence analysis. Three peptide sequences showed high identity to plastidic forms of starch phosphorylase (SP) from sweet potato, potato, and spinach. SP activity was identified in the amyloplast stromal fraction and was enriched 4-fold when compared with the activity in the whole endosperm fraction. Native and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analyses showed that SP activity was associated with the amyloplast stromal 112-kD protein. In addition, antibodies raised against the potato plastidic SP recognized the amyloplast stromal 112-kD protein. The amyloplast stromal 112-kD SP was expressed in whole endosperm isolated from maize harvested 9 to 24 d after pollination. Results of affinity electrophoresis and enzyme kinetic analyses showed that the amyloplast stromal 112-kD SP preferred amylopectin over glycogen as a substrate in the synthetic reaction. The maize shrunken-4 mutant had reduced SP activity due to a decrease of the amyloplast stromal 112-kD enzyme.
造粉体是玉米（Zea mays）贮藏组织中淀粉合成的场所。与整个胚乳相比，造粉体基质含有一组丰富的蛋白质。分子量为76和85 kD的蛋白质分别被鉴定为淀粉合酶I和淀粉分支酶IIb。通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳从基质组分中分离出一种112-kD的蛋白质，并对其进行胰蛋白酶消化和氨基酸序列分析。三个肽序列与甘薯、马铃薯和菠菜中淀粉磷酸化酶（SP）的质体形式具有高度同源性。在造粉体基质组分中鉴定出SP活性，与整个胚乳组分中的活性相比，其活性提高了4倍。天然和十二烷基硫酸钠-聚丙烯酰胺凝胶电泳分析表明，SP活性与造粉体基质112-kD蛋白质相关。此外，针对马铃薯质体SP产生的抗体识别造粉体基质112-kD蛋白质。造粉体基质112-kD SP在授粉后9至24天收获的玉米分离的整个胚乳中表达。亲和电泳和酶动力学分析结果表明，在合成反应中，造粉体基质112-kD SP以支链淀粉为底物优于糖原。由于造粉体基质112-kD酶的减少，玉米皱缩-4突变体的SP活性降低。
