Topography of cell wall lytic enzyme in Chlamydomonas reinhardtii: form and location of the stored enzyme in vegetative cell and gamete.
Topography of cell wall lytic enzyme in Chlamydomonas reinhardtii: form and location of the stored enzyme in vegetative cell and gamete.
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DOI：
10.1083/jcb.104.2.321
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发表时间：
1987-02
期刊：
影响因子：
--
通讯作者：
Hayashi K
中科院分区：
文献类型：
--
作者：
Matsuda Y;Saito T;Yamaguchi T;Koseki M;Hayashi K
衣藻细胞壁的裂解酶(配子壁自溶素)负责反交配子交配过程中细胞壁的脱落。本文报道了细胞内裂解酶的一些形态特征。营养细胞和配子细胞都含有相同的壁裂解酶。纯化的酶是一种糖蛋白，凝胶过滤表观分子量为67kD，SDS-PAGE表观分子量为62kD，对金属离子螯合剂和SH阻断剂敏感。这些特性与配子交配后释放到介质中的配子壁自溶素相同。然而，酶的储存形式在这两种细胞类型中被证明是非常不同的。在营养细胞中，裂解酶以不溶解的形式存在于细胞匀浆中，只有通过超声或冻融细胞才能将活性释放到可溶部分，而配子总是在细胞匀浆的可溶部分产生裂解活性。当营养细胞缺乏氮素时，随着交配能力的获得，酶的储存形式从营养状态转变为配子状态。在配子中加入氮素，在失去交配能力的同时，将其转化为营养状态。我们还发现，用外源加酶处理营养细胞或配子获得的原生质体在细胞匀浆中几乎没有酶活性，这表明裂解酶储存在质膜之外。当壁缺陷突变体CW-15的去壁配子或相反交配型的配子混合在一起时，它们正常交配，但裂解酶的释放几乎可以忽略不计。当去壁营养细胞孵化时，裂解酶在细胞壁再生几乎完成后再次积累。
Chlamydomonas lytic enzyme of the cell wall (gamete wall-autolysin) is responsible for shedding of cell walls during mating of opposite mating- type gametes. This paper reports some topographic aspects of lytic enzyme in cells. Both vegetative and gametic cells contain the same wall lytic enzyme. The purified enzyme is a glycoprotein with an apparent molecular mass of 67 kD by gel filtration and 62 kD by SDS PAGE, and is sensitive to metal ion chelators and SH-blocking agents. These properties are the same as those of the gamete wall-autolysin released into the medium by mating gametes. However, the storage form of the enzyme proves to be quite different between the two cell types. In vegetative cells, the lytic enzyme is found in an insoluble form in cell homogenates and activity is released into the soluble fraction only by sonicating the homogenates or freeze-thawing the cells, whereas gametes always yield lytic activity in the soluble fractions of cell homogenates. When vegetative cells are starved for nitrogen, the storage form of enzyme shifts from its vegetative state to gametic state in parallel with the acquisition of mating ability. Adding nitrogen to gametes converts it to the vegetative state concurrently with the loss of mating ability. We also show that protoplasts obtained by treatment of vegetative cells or gametes with exogenously added enzyme have little activity of enzyme in the cell homogenates, suggesting that lytic enzyme is stored outside the plasmalemma. When the de-walled gametes or gametes of the wall-deficient mutant, cw-15, of opposite mating types are mixed together, they mate normally but the release of lytic enzyme into the medium is practically negligible. When the de-walled vegetative cells are incubated, the lytic enzyme is again accumulated in the cells after the wall regeneration is almost complete.