Mössbauer spectroscopy as a tool for the study of activation/inactivation of the transcription regulator FNR in whole cells of Escherichia coli

The global regulator FNR (for fumarate nitrate reduction) controls the transcription of >100 genes whose products facilitate adaptation of Escherichia coli to growth under O(2)-limiting conditions. Previous Mössbauer studies have shown that anaerobically purified FNR contains a [4Fe-4S](2+) cluster that, on exposure to oxygen, is converted into a [2Fe-2S](2+) cluster, a process that decreases DNA binding by FNR. Using (57)Fe Mössbauer spectroscopy of E. coli cells containing overexpressed FNR, we show here that the same cluster conversion also occurs in vivo on exposure to O(2). Furthermore, the data show that a significant amount of the [4Fe-4S](2+) cluster is regenerated when the cells are shifted back to an anaerobic environment. The present study also demonstrates that (57)Fe Mössbauer spectroscopy can be employed to study the in vivo behavior of (overexpressed) proteins. The use of this technique to study other iron-containing cell components is discussed.

全局调控因子FNR（fumarate nitrate reduction，延胡索酸硝酸盐还原调控因子）可调控百余个基因的转录，这些基因的编码产物可帮助大肠杆菌（Escherichia coli）适应氧气受限环境下的生长。此前的穆斯堡尔光谱（Mössbauer spectroscopy）研究表明，厌氧纯化获得的FNR含有[4Fe-4S](2+)簇，当暴露于氧气时，该簇会转化为[2Fe-2S](2+)簇，这一过程会降低FNR与DNA的结合能力。本研究通过对过表达FNR的大肠杆菌细胞开展(57)Fe标记的穆斯堡尔光谱分析，证实该簇的转化在暴露于氧气的活体内同样会发生。此外，实验数据显示，当细胞被重新转移至厌氧环境时，大量的[4Fe-4S](2+)簇可实现再生。本研究同时证实，(57)Fe标记的穆斯堡尔光谱可用于研究（过表达）蛋白质在活体内的行为特征。本文还讨论了该技术在研究其他含铁细胞组分方面的应用前景。