Hess J, Gentschev I, Goebel W, Jarchau T
Institut für Genetik und Mikrobiologie, Universität Würzburg, Federal Republic of Germany.
Mol Gen Genet. 1990 Nov;224(2):201-8. doi: 10.1007/BF00271553.
We studied the efficiency of the pHly152-derived haemolysin transport system using PhoA-HlyA fusion proteins and different constructs which provide HlyB/HlyD in trans. The optimal C-terminal HlyA signal consists of the last 60 amino acids. Longer stretches of HlyA do not improve the transport efficiency of PhoA-HlyA fusion proteins. The introduction of deletions and/or replacements in the 60 amino acid HlyA signal domain revealed at least three functional regions with different degrees of specificity. Amino acids 1-21 (numbered from the N-terminal part of the 60 amino acid HlyA signal), termed region I, could be replaced by a Pro-containing peptide. The other two regions II and III (amino acids 22-40 and 41-60, respectively) seem to interact directly with the HlyB/HlyD translocator since a PhoA fusion protein which contains either of the two regions was still secreted in a HlyB/HlyD-dependent mode, albeit at low efficiency. An efficient trans-complementing HlyB/HlyD system was only obtained from the pHLy152-encoded hly determinant when the regulatory hlyR element was provided in cis. Secretion of the PhoA-HlyA fusion protein did not interfere with the secretion of HlyA even when the fusion protein was induced to a high level. This suggests that the capacity of the HlyB/HlyD translocation system is high and not normally saturated by its natural HlyA substrate.
我们使用PhoA-HlyA融合蛋白以及能反式提供HlyB/HlyD的不同构建体，研究了pHly152衍生的溶血素转运系统的效率。最佳的C端HlyA信号由最后60个氨基酸组成。更长的HlyA片段并不能提高PhoA-HlyA融合蛋白的转运效率。在60个氨基酸的HlyA信号结构域中引入缺失和/或替换，揭示了至少三个具有不同特异性程度的功能区域。编号从60个氨基酸的HlyA信号的N端部分起的第1-21位氨基酸，称为区域I，可以被含脯氨酸的肽取代。另外两个区域II和III（分别为第22-40位和第41-60位氨基酸）似乎直接与HlyB/HlyD转运体相互作用，因为包含这两个区域之一的PhoA融合蛋白仍以HlyB/HlyD依赖的方式分泌，尽管效率较低。只有当顺式提供调控性hlyR元件时，才能从pHLy152编码的hly决定簇中获得高效的反式互补HlyB/HlyD系统。即使将PhoA-HlyA融合蛋白诱导到高水平，其分泌也不会干扰HlyA的分泌。这表明HlyB/HlyD转运系统的能力很强，通常不会被其天然HlyA底物饱和。
