Garcia Darysbel, Watts Kylie J, Johnson Mark S, Taylor Barry L
Division of Microbiology and Molecular Genetics, School of Medicine, Loma Linda University, Loma Linda, CA, 92350, USA.
Mol Microbiol. 2016 Apr;100(1):156-72. doi: 10.1111/mmi.13308. Epub 2016 Feb 2.
The Escherichia coli aerotaxis receptor, Aer, monitors cellular oxygen and redox potential via FAD bound to a cytosolic PAS domain. Here, we show that Aer-PAS controls aerotaxis through direct, lateral interactions with a HAMP domain. This contrasts with most chemoreceptors where signals propagate along the protein backbone from an N-terminal sensor to HAMP. We mapped the interaction surfaces of the Aer PAS, HAMP and proximal signalling domains in the kinase-off state by probing the solvent accessibility of 129 cysteine substitutions. Inaccessible PAS-HAMP surfaces overlapped with a cluster of PAS kinase-on lesions and with cysteine substitutions that crosslinked the PAS β-scaffold to the HAMP AS-2 helix. A refined Aer PAS-HAMP interaction model is presented. Compared to the kinase-off state, the kinase-on state increased the accessibility of HAMP residues (apparently relaxing PAS-HAMP interactions), but decreased the accessibility of proximal signalling domain residues. These data are consistent with an alternating static-dynamic model in which oxidized Aer-PAS interacts directly with HAMP AS-2, enforcing a static HAMP domain that in turn promotes a dynamic proximal signalling domain, resulting in a kinase-off output. When PAS-FAD is reduced, PAS interaction with HAMP is relaxed and a dynamic HAMP and static proximal signalling domain convey a kinase-on output.
大肠杆菌趋氧性受体Aer通过与胞质PAS结构域结合的FAD监测细胞内的氧气和氧化还原电位。在此，我们表明Aer-PAS通过与HAMP结构域的直接横向相互作用来控制趋氧性。这与大多数化学感受器不同，在大多数化学感受器中，信号沿着蛋白质主链从N端传感器传播到HAMP。我们通过探测129个半胱氨酸取代的溶剂可及性，绘制了处于激酶关闭状态的Aer PAS、HAMP和近端信号结构域的相互作用表面。无法触及的PAS-HAMP表面与一组PAS激酶开启损伤以及使PASβ支架与HAMP AS-2螺旋交联的半胱氨酸取代重叠。我们提出了一个优化的Aer PAS-HAMP相互作用模型。与激酶关闭状态相比，激酶开启状态增加了HAMP残基的可及性（显然放松了PAS-HAMP相互作用），但降低了近端信号结构域残基的可及性。这些数据与交替的静态-动态模型一致，在该模型中，氧化的Aer-PAS直接与HAMP AS-2相互作用，使HAMP结构域处于静态，进而促进近端信号结构域处于动态，从而产生激酶关闭输出。当PAS-FAD被还原时，PAS与HAMP的相互作用被放松，动态的HAMP和静态的近端信号结构域传递激酶开启输出。
