Yoshimura Hideaki, Yoshioka Shiro, Kobayashi Katsuaki, Ohta Takehiro, Uchida Takeshi, Kubo Minoru, Kitagawa Teizo, Aono Shigetoshi
Okazaki Institute for Integrative Bioscience, National Institutes of Natural Sciences, 5-1 Higashiyama, Myodaiji, Okazaki, Aichi 444-8787, Japan.
Biochemistry. 2006 Jul 11;45(27):8301-7. doi: 10.1021/bi060315c.
HemAT from Bacillus subtilis (HemAT-Bs) is a heme-based O2 sensor protein that acts as a signal transducer responsible for aerotaxis. HemAT-Bs discriminates its physiological effector, O2, from other gas molecules to generate the aerotactic signal, but the detailed mechanism of the selective O2 sensing is not obvious. In this study, we measured electronic absorption, electron paramagnetic resonance (EPR), and resonance Raman spectra of HemAT-Bs to elucidate the mechanism of selective O2 sensing by HemAT-Bs. Resonance Raman spectroscopy revealed the presence of a hydrogen bond between His86 and the heme propionate only in the O2-bound form, in addition to that between Thr95 and the heme-bound O2. The disruption of this hydrogen bond by the mutation of His86 caused the disappearance of a conformer with a direct hydrogen bond between Thr95 and the heme-bound O2 that is present in WT HemAT-Bs. On the basis of these results, we propose a model for selective O2 sensing by HemAT-Bs as follows. The formation of the hydrogen bond between His86 and the heme propionate induces a conformational change of the CE-loop and the E-helix by which Thr95 is located at the proper position to form the hydrogen bond with the heme-bound O2. This stepwise conformational change would be essential to selective O2 sensing and signal transduction by HemAT-Bs.
来自枯草芽孢杆菌的HemAT（HemAT-Bs）是一种基于血红素的氧气传感蛋白，作为负责趋氧性的信号转导器。HemAT-Bs能够将其生理效应物氧气与其他气体分子区分开来，以产生趋氧信号，但选择性氧气传感的详细机制尚不清楚。在本研究中，我们测量了HemAT-Bs的电子吸收光谱、电子顺磁共振（EPR）光谱和共振拉曼光谱，以阐明HemAT-Bs选择性氧气传感的机制。共振拉曼光谱显示，除了Thr95与血红素结合的氧气之间存在氢键外，仅在氧气结合形式下，His86与血红素丙酸酯之间也存在氢键。His86突变导致该氢键断裂，使得野生型HemAT-Bs中存在的Thr95与血红素结合的氧气之间具有直接氢键的构象体消失。基于这些结果，我们提出了HemAT-Bs选择性氧气传感的模型如下。His86与血红素丙酸酯之间氢键的形成诱导了CE环和E螺旋的构象变化，通过这种变化，Thr95位于与血红素结合的氧气形成氢键的合适位置。这种逐步的构象变化对于HemAT-Bs的选择性氧气传感和信号转导至关重要。
