Purification and biosynthesis of a derepressible periplasmic arylsulfatase from Chlamydomonas reinhardtii.
Purification and biosynthesis of a derepressible periplasmic arylsulfatase from Chlamydomonas reinhardtii.
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DOI：
10.1083/jcb.106.1.29
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发表时间：
1988-01
影响因子：
7.8
通讯作者：
Grossman, A
中科院分区：
文献类型：
--
作者：
de Hostos, E L;Togasaki, R K;Grossman, A
单细胞绿色衣藻莱茵衣藻通过产生芳基硫酸酯酶来响应硫酸盐剥夺（Lien，T.，和O.施赖纳1975.生物化学生物Biophys. Acta. 384：168-179; Schreiner，O.，1975.生物化学生物Biophys. Acta. 384：180-193）和通过发展更快地运输硫酸盐的能力（我们未发表的数据）。在将细胞转移到低硫酸盐培养基（小于或等于10 μ M硫酸盐）后3小时可检测到的芳基硫酸酯酶活性是由cw 15（C.莱因哈德氏菌我们已将去阻遏芳基硫酸酯酶纯化至均一，并产生了单特异性抗体，蛋白单体（67.6 kD）结合成二聚体，酶活性显示碱性pH最适和对硝基苯硫酸的Km为0.3 mM。芳基硫酸酯酶生物合成的研究表明，它是糖基化和合成的更高分子量的前体。成熟蛋白含有复杂的N-连接寡糖，初级翻译产物的表观分子量比去糖基化单体大约5 kD。由于编码芳基硫酸酯酶的可翻译RNA只能在硫酸盐饥饿后的细胞中检测到，因此该酶的积累可能在转录水平上受到调节，尽管也可能涉及转录后过程。
The unicellular green alga Chlamydomonas reinhardtii responds to sulfate deprivation by producing an arylsulfatase (Lien, T., and O. Schreiner. 1975. Biochim. Biophys. Acta. 384:168-179; Schreiner, O., 1975. Biochim. Biophys. Acta. 384:180-193) and by developing the capacity to transport sulfate more rapidly (our unpublished data). The arylsulfatase activity, detectable 3 h after the transfer of the cells to low sulfate medium (less than or equal to 10 microM sulfate), is a periplasmic protein released into the culture medium by cw15, a cell wall-less mutant of C. reinhardtii. We have purified the derepressible arylsulfatase to homogeneity and have raised monospecific antibodies to it. The protein monomer (67.6 kD) associates into a dimer, and the enzyme activity shows an alkaline pH optimum and a Km of 0.3 mM for p- nitrophenylsulfate. Studies focused on arylsulfatase biosynthesis demonstrate that it is glycosylated and synthesized as a higher molecular mass precursor. The mature protein contains complex N-linked oligosaccharides and the primary translation product has an apparent molecular mass approximately 5 kD larger than the deglycosylated monomer. Since translatable RNA encoding the arylsulfatase can only be detected in cells after sulfate starvation, it is likely that accumulation of the enzyme is regulated at the level of transcription, although posttranscriptional processes may also be involved.