A specific targeting domain in mature exotoxin A is required for its extracellular secretion from Pseudomonas aeruginosa.
A specific targeting domain in mature exotoxin A is required for its extracellular secretion from Pseudomonas aeruginosa.
复制标题
成熟外毒素 A 中的特定靶向结构域是其从铜绿假单胞菌分泌到细胞外所必需的。
DOI：
10.1002/j.1460-2075.1996.tb00373.x
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发表时间：
1996
期刊：
影响因子：
--
通讯作者：
Stephen Loryl
中科院分区：
文献类型：
--
作者：
Hong;Stephen Loryl
许多革兰氏阴性细菌，包括铜绿假单胞菌，主动分泌一种周质蛋白到周围培养基中。一个推定的细胞外靶向信号内的一个这样的蛋白质，外毒素A，进行了研究。构建了一系列与β-内酰胺酶融合的外毒素A截短物。在其N末端携带成熟外毒素A的120、255、355或整个613个残基的杂合蛋白是稳定的，并分泌到细胞外培养基中。携带成熟外毒素A的残基1-30和1-60的杂合蛋白是不稳定的;然而，它们可以在短的标记期后在细胞内完全检测到。构建了仅携带外毒素A的N-末端残基1-3和区域60-120的β-内酰胺酶杂合体。它也被分泌到培养基中，表明一个特定的60个氨基酸的结构域包含必要的靶向信息的外毒素A跨外膜易位。杂合蛋白的分泌与过客蛋白无关，因为也分泌了类似的外毒素A-鼠白细胞介素4杂合蛋白。氨基酸60和120之间的细胞外靶向信号富含反平行β折叠。先前已显示其参与外毒素A与真核细胞受体的相互作用。在三维视图中，靶向区域位于毒素表面上，在那里它很容易接近细胞外分泌机制的组分。
A number of Gram‐negative bacteria, including Pseudomonas aeruginosa, actively secrete a subset of periplasmic proteins into their surrounding medium. The presence of a putative extracellular targeting signal within one such protein, exotoxin A, was investigated. A series of exotoxin A truncates, fused to beta‐lactamase, was constructed. Hybrid proteins, which carry at their N‐ termini 120, 255, 355 or the entire 613 residues of the mature exotoxin A, were stable and were secreted into the extracellular medium. Hybrid proteins which carry residues 1–30 and 1–60 of the mature exotoxin A were unstable; however, they could be detected entirely within the cells after a short labeling period. A hybrid with beta‐lactamase was constructed which carried only the N‐terminal residues 1–3 and region 60–120 of exotoxin A. It was also secreted into the culture medium, suggesting that a specific 60 amino acid domain contains the necessary targeting information for translocation of exotoxin A across the outer membrane. The secretion of the hybrid proteins is independent of the passenger protein, since a similar exotoxin A‐murine interleukin 4 hybrid protein was also secreted. The extracellular targeting signal between amino acids 60 and 120 is rich in anti‐parallel beta‐sheets. It has been shown previously to be involved in the interaction of the exotoxin A with the receptors of the eukaryotic cells. In the three‐ dimensional view, the targeting region is on the toxin surface where it is easily accessible to the components of the extracellular secretion machinery.