Bame K J, Lidholt K, Lindahl U, Esko J D
Department of Biochemistry, School of Medicine, University of Alabama, Birmingham 35294.
J Biol Chem. 1991 Jun 5;266(16):10287-93.
A previous study identified a Chinese hamster ovary cell mutant, pgsE-606, which is defective in the N-sulfotransferase that catalyzes one of the initial polymer-modification reactions in the biosynthesis of heparan sulfate (Bame, K. J., and Esko, J. D. (1989) J. Biol. Chem. 264, 8059-8065). The structure of heparan sulfate generated by these cells reflects a 3-5-fold reduction in enzyme activity. The mutant produces heparan sulfate with half the content of N-sulfated glucosamine residues of that produced by wild-type cells and a more sparse distribution of N-sulfated residues. The present study demonstrates corresponding reductions in the proportion of 6-O-sulfated glucosamine residues (41% reduction) and the content of L-iduronic acid (51% reduction). The amount of 2-O-sulfated L-iduronic acid declines more dramatically (from 25% of total L-iduronic acid in the wild type to 8.4% in the mutant). Enzymatic assay of mixed O-sulfotransferases showed that the mutant has more activity than the wild type. Previous studies on the biosynthesis of heparin/heparan sulfate in cell-free systems point to a pivotal role of N-sulfation in determining the extent of the subsequent polymer-modification reactions. The present study shows that this concept also applies to heparan sulfate biosynthesis in the intact cell.
先前的一项研究鉴定出一种中国仓鼠卵巢细胞突变体pgsE - 606，它在N - 磺基转移酶方面存在缺陷，该酶催化硫酸乙酰肝素生物合成中最初的聚合物修饰反应之一（巴姆，K. J.，和埃斯科，J. D.（1989年）《生物化学杂志》264卷，8059 - 8065页）。这些细胞产生的硫酸乙酰肝素的结构反映出酶活性降低了3至5倍。该突变体产生的硫酸乙酰肝素中N - 硫酸化葡糖胺残基的含量是野生型细胞产生的一半，且N - 硫酸化残基的分布更为稀疏。本研究表明6 - O - 硫酸化葡糖胺残基的比例相应降低（降低了41%），L - 艾杜糖醛酸的含量也降低（降低了51%）。2 - O - 硫酸化L - 艾杜糖醛酸的量下降更为显著（从野生型中占总L - 艾杜糖醛酸的25%降至突变体中的8.4%）。对混合O - 磺基转移酶的酶活性测定表明，该突变体比野生型具有更高的活性。先前在无细胞系统中对肝素/硫酸乙酰肝素生物合成的研究指出，N - 硫酸化在决定后续聚合物修饰反应的程度方面起着关键作用。本研究表明这一概念也适用于完整细胞中的硫酸乙酰肝素生物合成。
