Lloyd J R, Cole J A, Macaskie L E
School of Biological Sciences, University of Birmingham, Edgbaston, United Kingdom.
J Bacteriol. 1997 Mar;179(6):2014-21. doi: 10.1128/jb.179.6.2014-2021.1997.
Anaerobic, but not aerobic, cultures of Escherichia coli accumulated Tc(VII) and reduced it to a black insoluble precipitate. Tc was the predominant element detected when the precipitate was analyzed by proton-induced X-ray emission. Electron microscopy in combination with energy-dispersive X-ray analysis showed that the site of Tc deposition was intracellular. It is proposed that Tc precipitation was a result of enzymatically mediated reduction of Tc(VII) to an insoluble oxide. Formate was an effective electron donor for Tc(VII) reduction which could be replaced by pyruvate, glucose, or glycerol but not by acetate, lactate, succinate, or ethanol. Mutants defective in the synthesis of the transcription factor FNR, in molybdenum cofactor (molybdopterin guanine dinucleotide [MGD]) synthesis, or in formate dehydrogenase H synthesis were all defective in Tc(VII) reduction, implicating a role for the formate hydrogenlyase complex in Tc(VII) reduction. The following observations confirmed that the hydrogenase III (Hyc) component of formate hydrogenlyase in both essential and sufficient for Tc(VII) reduction: (i) dihydrogen could replace formate as an effective electron donor for Tc(VII) reduction by wild-type bacteria and mutants defective in MGD synthesis; (ii) the inability of fnr mutants to reduce Tc(VII) can be suppressed phenotypically by growth with 250 microM Ni2+ and formate; (iii) Tc(VII) reduction is defective in a hyc mutant; (iv) the ability to reduce Tc(VII) was repressed during anaerobic growth in the presence of nitrate, but this repression was counteracted by the addition of formate to the growth medium; (v) H2, but not formate, was an effective electron donor for a Sel- mutant which is unable to incorporate selenocysteine into any of the three known formate dehydrogenases of E. coli. This appears to be the first report of Hyc functioning as an H2-oxidizing hydrogenase or as a dissimilatory metal ion reductase in enteric bacteria.
大肠杆菌的厌氧培养物(而非需氧培养物)积累了Tc(VII)并将其还原为黑色不溶性沉淀。当通过质子诱导X射线发射分析沉淀时,Tc是检测到的主要元素。电子显微镜结合能量色散X射线分析表明,Tc沉积位点在细胞内。有人提出,Tc沉淀是酶介导的将Tc(VII)还原为不溶性氧化物的结果。甲酸盐是Tc(VII)还原的有效电子供体,可被丙酮酸、葡萄糖或甘油替代,但不能被乙酸盐、乳酸盐、琥珀酸盐或乙醇替代。在转录因子FNR合成、钼辅因子(钼蝶呤鸟嘌呤二核苷酸[MGD])合成或甲酸脱氢酶H合成方面存在缺陷的突变体在Tc(VII)还原方面均有缺陷,这表明甲酸氢化酶复合物在Tc(VII)还原中起作用。以下观察结果证实,甲酸氢化酶的氢化酶III(Hyc)组分对于Tc(VII)还原既是必需的也是充分的:(i)氢气可以替代甲酸盐作为野生型细菌和MGD合成缺陷突变体还原Tc(VII)的有效电子供体;(ii)fnr突变体无法还原Tc(VII)的表型可通过在含有250 microM Ni2+和甲酸盐的条件下生长得到抑制;(iii)Tc(VII)还原在hyc突变体中存在缺陷;(iv)在硝酸盐存在下厌氧生长期间,还原Tc(VII)的能力受到抑制,但通过向生长培养基中添加甲酸盐可抵消这种抑制作用;(v)H2而非甲酸盐是Sel-突变体还原Tc(VII)的有效电子供体,该突变体无法将硒代半胱氨酸掺入大肠杆菌三种已知甲酸脱氢酶中的任何一种。这似乎是关于Hyc在肠道细菌中作为H2氧化氢化酶或异化金属离子还原酶发挥作用的首次报道。