Signal Transduction and Gene Regulation in Bacteria

大肠杆菌调节蛋白EnvZ和OmpR在响应于介质渗透压的外膜蛋白OmpF/OmpC的表达中起关键作用。EnvZ蛋白可能是一种位于膜上的渗透传感器（或信号转导器），它表现出OmpR蛋白特异性的激酶和磷酸酶活性。为了研究EnvZ分子的跨膜区段（命名为TM 1和TM 2）在跨膜信号传导中的功能重要性，一组EnvZ突变体，每个突变体在跨膜区域内具有氨基酸取代，其特征在于它们的体内表型和体外催化活性。其中之一，其特征在于，有一个氨基酸的变化（Pro-41丝氨酸或亮氨酸）在TM 1，似乎是有缺陷的磷酸酶活性，但不是在其激酶活性。该EnvZ突变体赋予OmpF^-/OmpC-组成型表型。对于这种EnvZ（P41 S或P41 L）突变体，分离出一组基因内抑制子，每个都具有OmpF^+/OmpC^+的野生型表型。这些抑制子突变体被揭示在TM 1或TM 2内具有额外的氨基酸变化。进一步证明，抑制子之一EnvZ（TM 2中的Arg-180至Trp）能够通过分子间互补抑制由EnvZ（P41 S）引起的体内表型和体外催化活性的缺陷。这些结果最好解释为意味着EnvZ分子作为寡聚体（最可能是二聚体）起作用，并且跨膜片段之间的紧密分子间相互作用对于响应于外部渗透刺激的跨膜信号传导至关重要。

The E.coli regulatory proteins, EnvZ and OmpR, are crucially involved in expression of the outer membrane proteins OmpF/OmpC in response to the medium osmolarity. The EnvZ protein is presumably a membrane-located osmotic sensor (or signal transducer), which exhibits both kinase and phosphatase activities specific for the OmpR protein. To examine the functional importance of the membrane-spanning segments (named TM1 and TM2) of EnvZ molecules in transmembrane signaling, a set of EnvZ mutants, each having amino acid substitutions within the membrane-spanning regions, was characterized in terms of both their in vivo phenotype and invitro catalytic activities. One of them, characterized further, has an amino acid change (Pro-41 to Ser or Leu) in TM1, and appeared to be defective in its phosphatase activity but not in its kinase activity. This EnvZ mutant conferred a phenotype of OmpF^-/OmpC-constitutive. For this EnvZ(P41S or P41L) mutant, a set of intragenic suppressors, each exhibting a wild-type phenotype of OmpF^+/OmpC^+, was isolated. These suppresor mutants were revealed to have an additional amino acid change within either TM1 or TM2. It was further demonstrated that one of the suppressors, EnvZ(Arg-180 to Trp in TM2), was able to suppress the defects in both the in vivo phenotype and in vitro catlytic activities caused by EnvZ(P41S), through an intermolecular complementation. These results are best interpreted as meaning that EnvZ molecules function an an oligomer (most likely a dimer), and that an intimate intermolecular interaction between the membrane-spanning segments is crucial fro transmembrane signaling in response to and external osmotic stimulus.