Fontaine T, D'Hulst C, Maddelein M L, Routier F, Pépin T M, Decq A, Wieruszeski J M, Delrue B, Van den Koornhuyse N, Bossu J P
Laboratoire de Chimie Biologique, Université des Sciences et Technologies de Lille Flandres-Artois, Villeneuve, d'Ascq, France.
J Biol Chem. 1993 Aug 5;268(22):16223-30.
Low starch mutants of Chlamydomonas reinhardtii were isolated after x-ray mutagenesis of wild-type strain 137C. The mutants accumulated 20-40% of the normal amount and displayed a 2-fold decrease of the total glycogen-primed soluble starch synthase activity. Three different mutant alleles of the st-3 gene were isolated that were characterized by similar defects and displayed a net increase in amylose content. Amylose-primed synthesis of glucan in native gels revealed a complete wipe out of one of the soluble starch synthases. Zymograms and kinetic analyses performed both in the mutant and in partially purified wild type extracts reveal at least two distinct activities that are partly analogous to higher plant soluble starch synthases I and II (SSI and II). The st-3 mutants were defective for SSII. Methylation and debranching of the purified amylopectin fraction clearly show a decrease in the number of intermediate size glucans (dp8 to 50) and an absolute and relative increase of very short glucans (dp2 to 7). These results suggest that a soluble starch synthase may be necessary for the synthesis or maintenance of intermediate size glucans that are the main component of the branched clusters of amylopectin.
对莱茵衣藻野生型菌株137C进行X射线诱变后，分离出了低淀粉突变体。这些突变体积累的淀粉量为正常量的20%-40%，并且总糖原引发的可溶性淀粉合酶活性降低了2倍。分离出了st-3基因的三个不同突变等位基因，它们具有相似的缺陷，并且直链淀粉含量有净增加。在天然凝胶中直链淀粉引发的葡聚糖合成显示其中一种可溶性淀粉合酶完全缺失。在突变体和部分纯化的野生型提取物中进行的酶谱分析和动力学分析表明，至少有两种不同的活性，它们部分类似于高等植物可溶性淀粉合酶I和II（SSI和II）。st-3突变体的SSII有缺陷。纯化的支链淀粉级分的甲基化和去分支分析清楚地表明，中等大小葡聚糖（dp8至50）的数量减少，非常短的葡聚糖（dp2至7）的绝对和相对含量增加。这些结果表明，可溶性淀粉合酶对于合成或维持作为支链淀粉分支簇主要成分的中等大小葡聚糖可能是必需的。
