Enterobactin synthase polypeptides of Escherichia coli are present in an osmotic-shock-sensitive cytoplasmic locality
Enterobactin synthase polypeptides of Escherichia coli are present in an osmotic-shock-sensitive cytoplasmic locality
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DOI：
10.1099/00221287-143-1-147
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发表时间：
1997-01-01
期刊：
影响因子：
--
通讯作者：
Earhart, CF
中科院分区：
文献类型：
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作者：
Hantash, FM;Ammerlaan, M;Earhart, CF
大肠杆菌合成铁载体肠杆菌素（Ent）的末端反应需要EntD、E、F和B/G多肽。体外结果未提供支持该提议的证据：（i）Ent合酶活性在膜不存在或被高浓度中性去污剂破坏的条件下正常发生，和（ii）对参与Ent合成的提取物进行的免疫沉淀实验未能检测到Ent多肽之间的任何关联，然而Western印迹分析显示EntE，F和B/G通过渗透压休克和冻融处理从细胞中释放出来，但不通过细胞转化为原生质球而释放出来。这些结果表明，EntE、F和B/G属于Beacham D组蛋白。给定的D组Ent蛋白的休克性不受EntB/G或EntD的存在的影响，并且对于EntB/G，N-末端足以通过渗透压休克释放。D组蛋白的行为通常归因于它们与细胞质膜的结合（部分的、松散的或瞬时的）;因此，结果是Ent合酶在体内与膜相互作用的间接证据。至少这些数据表明，EntE、F和B/G在E中是区室化的。由于铁载体和表面活性剂的其他生物合成酶与这些Ent蛋白有关，因此表明整个蛋白质类可能在体内被隔离。
The terminal reactions in the synthesis of the siderophore enterobactin (Ent) by Escherichia coli require the EntD, E, F and B/G polypeptides The idea that these molecules form a complex (Ent synthase) that is membrane-associated was re-evaluated. In vitro results provided no evidence in support of the proposal: (i) Ent synthase activity occurred normally under conditions where membrane was either absent or disrupted by high concentrations of neutral detergents, and (ii) immunoprecipitation experiments conducted on extracts engaged in Ent synthesis failed to detect any association among the Ent polypeptides however Western blot analyses showed that EntE, F and B/G were released from cells by osmotic shock and freeze/thaw treatment but not by conversion of cells to spheroplasts These results demonstrated that EntE, F and B/G belong to the Beacham group D class of proteins. The shockability of a given group D Ent protein was unaffected by the absence of either EntB/G or EntD and, for EntB/G, the N-terminus was sufficient for release by osmotic shock. The behaviour of group D proteins is generally attributed to their association (partial, loose or transient) with cytoplasmic membrane; therefore, the results are indirect evidence that Ent synthase interacts with membrane in vivo. At the very least the data indicate that EntE, F and B/G are compartmentalized in E. coli and, because other biosynthetic enzymes for siderophores and surfactants are related to these Ent proteins, suggest that this entire protein class may be sequestered in vivo.