首页> 外文学位 >N-chain glucose processing and proper beta-1,3-glucan biosynthesis are required for normal cell wall beta-1,6-glucan levels in Saccharomyces cerevisiae.
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N-chain glucose processing and proper beta-1,3-glucan biosynthesis are required for normal cell wall beta-1,6-glucan levels in Saccharomyces cerevisiae.

机译:酿酒酵母中正常的细胞壁β-1,6-葡聚糖水平需要N链葡萄糖加工和适当的β-1,3-葡聚糖生物合成。

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摘要

CWH41 is required for beta-1,6-glucan biosynthesis and encodes glucosidase I, an enzyme involved in protein N-chain glucose processing. Therefore, the effects of N-chain glucosylation and processing on beta-1,6-glucan biosynthesis were examined, and it was shown that incomplete N-chain glucose processing results in loss of beta-1,6-glucan. To explore the involvement of other N-chain-dependent events with beta-1,6-glucan synthesis, the S. cerevisiae KRE5 and CNE1 genes were investigated, which encode homologs of the 'quality control' components UDP-Glc:glycoprotein glucosyltransferase and calnexin, respectively. The essential activity of Kre5p was found to be separate from its possible role as a UDP-Glc:glycoprotein glucosyltransferase. A ∼30% decrease in beta-1,6-glucan was observed upon disruption of CNE1, a phenotype which is additive with other beta-1,6-glucan synthetic mutants. Analysis of the cell wall anchorage of alpha-agglutinin suggests the existence of two beta-1,6-glucan biosynthetic pathways, one N-chain dependent, the other involving protein glycosylphosphatidylinositol modification.; Fks1p and Fks2p are related proteins thought to be catalytic subunits of the beta-1,3-glucan synthase. The fks1Delta mutant was partial K1 killer toxin resistant and showed a 30% reduction in alkali-soluble beta-1,3-glucan that was accompanied by a modest reduction in beta-1,6-glucan. The gas1Delta mutant lacking a 1,3-beta-glucanosyltransferase displayed a similar reduction in alkali-soluble beta-1,3-glucan but did not share the beta-1,6-glucan defect, indicating that beta-1,6-glucan reduction is not a general phenotype among beta-1,3-glucan biosynthetic mutants. FKS2 overexpression suppressed the killer toxin phenotype of fks1Delta mutants, implicating Fks2p in the biosynthesis of the residual beta-1,6-glucan present in fks1Delta cells. Eight out of twelve fks1tsfks2Delta mutants had altered beta-glucan levels at the permissive temperature: the FKS1F1258Y N1520D allele was severely affected in both polymers and displayed a 55% reduction in beta-1,6-glucan, while the in vitro hyperactive FKS1T6051 M761T allele increased both beta-glucan levels. These beta-1,6-glucan phenotypes may be due to altered availability of, and structural changes in, the beta-1,3-glucan polymer, which might serve as a beta-1,6-glucan acceptor at the cell surface. Alternatively, Fks1p and Fks2p could actively participate in the biosynthesis of both polymers as beta-glucan transporters. beta-1,6-Glucan deficient mutants had reduced in vitro glucan synthase activity and mislocalized Fks1p and Fks2p, possibly contributing to the observed beta-1,6-glucan defects.
机译:CWH41是β-1,6-葡聚糖生物合成所必需的,它编码葡糖苷酶I(一种参与蛋白质N链葡萄糖加工的酶)。因此,研究了N链糖基化和加工对β-1,6-葡聚糖生物合成的影响,结果表明,不完整的N链葡萄糖加工会导致β-1,6-葡聚糖的损失。为了探索其他N链依赖性事件与beta-1,6-葡聚糖合成的关系,研究了酿酒酵母KRE5和CNE1基因,它们编码“质量控制”成分UDP-Glc:糖蛋白葡糖基转移酶和钙粘蛋白。发现Kre5p的基本活性与其作为UDP-Glc:糖蛋白葡萄糖基转移酶的可能作用是分开的。当CNE1断裂时,可观察到β-1,6-葡聚糖减少了约30%,CNE1是与其他β-1,6-葡聚糖合成突变体相加的表型。对α-凝集素的细胞壁锚定的分析表明存在两个β-1,6-葡聚糖生物合成途径,一个是N链依赖性的,另一个涉及蛋白质糖基磷脂酰肌醇的修饰。 Fks1p和Fks2p是相关的蛋白质,被认为是β-1,3-葡聚糖合酶的催化亚基。 fks1Delta突变体对K1杀伤毒素具有部分抗性,并显示碱溶性β-1,3-葡聚糖减少了30%,同时β-1,6-葡聚糖也有所减少。缺少1,3-β-葡糖基糖基转移酶的gas1Delta突变体在碱溶性β-1,3-葡聚糖中显示出相似的减少,但没有共享β-1,6-葡聚糖缺陷,表明β-1,6-葡聚糖在β-1,3-葡聚糖生物合成突变体中,减少不是普遍的表型。 FKS2过表达抑制了fks1Delta突变体的杀手毒素表型,将Fks2p牵涉到fks1Delta细胞中残留β-1,6-葡聚糖的生物合成中。在允许的温度下,十二个fks1tsfks2Delta突变体中的八个已经改变了β-葡聚糖的水平:两种聚合物中FKS1F1258Y N1520D等位基因均受到严重影响,β-1,6-葡聚糖减少了55%,而体外FKS1T6051 M761T等位基因活跃增加了两个β-葡聚糖的水平。这些β-1,6-葡聚糖表型可能是由于β-1,3-葡聚糖聚合物的可利用性和结构变化所致,它可能在细胞表面充当β-1,6-葡聚糖受体。另外,Fks1p和Fks2p可以作为β-葡聚糖转运蛋白积极参与两种聚合物的生物合成。 beta-1,6-葡聚糖缺陷型突变体降低了体外葡聚糖合酶的活性和Fks1p和Fks2p的错误定位,可能有助于观察到的beta-1,6-葡聚糖缺陷。

著录项

  • 作者单位

    McGill University (Canada).;

  • 授予单位 McGill University (Canada).;
  • 学科 Biology Molecular.; Biology Cell.; Chemistry Biochemistry.
  • 学位 Ph.D.
  • 年度 2002
  • 页码 173 p.
  • 总页数 173
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类 分子遗传学;细胞生物学;生物化学;
  • 关键词

  • 入库时间 2022-08-17 11:46:06

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