Saccharomyces cerevisiae cell wall chitin, the Kluyveromyces lactis zymocin receptor
Saccharomyces cerevisiae cell wall chitin, the Kluyveromyces lactis zymocin receptor
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DOI：
10.1002/yea.776
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发表时间：
2001-10-01
期刊：
影响因子：
2.6
通讯作者：
Schaffrath, R
中科院分区：
文献类型：
--
作者：
Jablonowski, D;Fichtner, L;Schaffrath, R
乳酸克鲁维酵母分泌的外切酶素会导致敏感的酵母细胞（包括酿酒酵母）将生长抑制在细胞周期的 G1 期。尽管其具有异三聚体（αβγ）结构，但其最小亚基γ-毒素的细胞内表达单独负责G(1)阻滞。然而，α亚基具有几丁质酶活性，这对于细胞外部的全酶素作用至关重要。在这里，我们表明，通过外源应用粗制几丁质制剂，可以将敏感酵母细胞从酶素处理中拯救出来，这支持了这样的观点，即几丁质聚合物可以与敏感酵母细胞表面存在的几丁质竞争与酶素的结合。与此一致，可以使用固定化几丁质基质通过亲和层析纯化holozymocin。 PCR 介导的几丁质合成 (CHS) 基因删除表明，大多数（如果不是全部）遗传情况会导致几丁质合酶 III 活性完全丢失 (chs Delta)、输出受阻 (ells7 Delta) 或激活减少 (chs4 Delta)，并伴有错误定位（chs4 Delta chs5 Delta；chs4 Delta chs6 Delta；chs4 Delta chs5 Delta chs6 Delta） (CSIII)，使细胞对 exozymocin 的抑制作用产生抵抗。相比之下，分别编码 CSI 和 CSII 的 CHS1 和 CHS2 中的缺失对酶素敏感性没有影响。因此，CSIII聚合几丁质（几乎占细胞几丁质资源的90%）似乎是酶素与敏感细胞初始相互作用所需的碳水化合物受体。版权所有 (C) 2001 John Wiley & Sons, Ltd.
The exozymocin secreted by Kluyveromyces lactis causes sensitive yeast cells, including Saccharomyces cerevisiae, to arrest growth in the G, phase of the cell cycle. Despite its heterotrimeric (alpha beta gamma) structure, intracellular expression of its smallest subunit, the gamma -toxin, is alone responsible for the G(1) arrest. The alpha subunit, however, has a chitinase activity that is essential for holozymocin action from the cell exterior. Here we show that sensitive yeast cells can be rescued from zymocin treatment by exogenously applying crude chitin preparations, supporting the idea that chitin polymers can compete for binding to zymocin with chitin present on the surface of sensitive yeast cells. Consistent with this, holozymocin can be purified by way of affinity chromatography using an immobilized chitin matrix. PCR-mediated deletions of chitin synthesis (CHS) genes show that most, if not all, genetic scenarios that lead to complete loss (chs Delta), blocked export (ells7 Delta) or reduced activation (chs4 Delta), combined with mislocalization (chs4 Delta chs5 Delta; chs4 Delta chs6 Delta; chs4 Delta chs5 Delta chs6 Delta) of chitin synthase III activity (CSIII), render cells refractory to the inhibitory effects of exozymocin. In contrast, deletions in CHS1 and CHS2, which code for CSI and CSII, respectively, have no effect on zymocin sensitivity. Thus, CSIII-polymerized chitin, which amounts to almost 90%,, of the cell's chitin resources, appears to be the carbohydrate receptor required for the initial interaction of zymocin with sensitive cells. Copyright (C) 2001 John Wiley & Sons, Ltd.