Structure and function of the heme-based gas sensor proteins

基于血红素的气体传感器蛋白的结构和功能

基本信息

批准号：
14380303
负责人：
AONO Shigetoshi
金额：
$ 9.73万
依托单位：
Okazaki National Research Institutes
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2003
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14380303/
关键词：
Sensor Protein Aerotaxis Control System Hemoprotein Oxygen Sensor protein

项目摘要

HemAT-Bs is a heme-containing signal transducer protein responsible for aerotaxis of Bacillus subtilis. The recombinant HemAT-Bs expressed in Escherichia coli was purified as the oxy form in which oxygen was bound to the ferrous heme. Oxygen binding and dissociation rate constants were determined to be kon = 32 μM^<-1> s^<-1> and koff = 23^<-1>, respectively, revealing that HemAT-Bs has a moderate oxygen affinity similar to that of sperm whale myoglobin (Mb). The rate constant for autoxidation at 37 ℃ was 0.06 h^<-1>, which is also close to that of Mb. Although the electronic absorption spectra of HemAT-Bs were similar to those of Mb, HemAT-Bs showed some unique characteristics in its resonance Raman spectra. Oxygen-bound HemAT-Bs gave the Fe-0_2 band at a noticeably low frequency (560 cm^<-1>), which suggests a unique hydrogen bonding between a distal amino acid residue and the proximal atom of the bound oxygen molecule. Deoxy HemAT-Bs gave the Fe-His band at a higher frequency (225 cm<-1>) than those of ordinary His-coordinated deoxy heme proteins. CO-bound HemAT-Bs gave the Fe-CO and C-O bands at 494 and 1964 cm1, respectively, which fall on the same C-O versus Fe-CO correlation line as that of Mb. Based on these results, the structural and functional properties of HemAT-Bs has been elucidated.In this work, a new CooA homologue from C.hydrogenoformans (Ch-CooA) was also studied. Ch-CooA also contains a protoheme as the active site for sensing CO. The heme in Ch-CooA is six-coordinate, low spin state in the ferric, ferrous and CO-bound forms. The heme environmental structure of Ch-CooA was elucidated by mutagenesis and spectroscopic studies.

HEMAT-BS是负责枯草芽孢杆菌空气的含血红素信号传感器蛋白。在大肠杆菌中表达的重组hemat-bs被纯化为氧与亚铁血红素结合的氧形式。确定氧结合和解离速率常数为KON =32μM^<-1> S^<-1>和Koff = 23^<-1>，表明Hemat-BS的现代氧亲和力与精子鲸鱼肌红蛋白（MB）相似。 37℃时自氧化的速率常数为0.06 H^<-1>，也接近MB的速率常数。尽管HEMAT-BS的电子损失光谱与MB的电子损失光谱相似，但HEMAT-BS在其谐振拉曼光谱中显示出一些独特的特征。结合氧气的HEMAT-BS以明显的低频（560 cm^<-1>）的形式给出了Fe-0_2带，这表明远端氨基酸保留和结合氧分子的代理之间存在独特的氢键。脱氧Hemat-BS比普通的His协调脱氧血红素蛋白的Fe-His带的频率更高（225 cm <-1>）。共同绑定的Hemat-BS分别在494和1964 CM1下给出了Fe-CO和C-O频段，该频段与MB相同的C-O与Fe-CO相关线相同。基于这些结果，已经阐明了HEMAT-BS的结构和功能特性。在这项工作中，也研究了来自C. hydrogenogenogenogenogenogenogenogenogenogentoy（CH-COOA）的新的COOA同源物。 CH-COOA还包含一个原始素作为传感CO的活性位点。CH-COOA中的血红素是六坐标，低旋转状态，在铁，亚铁和共同的形式中。通过诱变和光谱研究阐明了CH-COOA的血红素环境结构。